# ent

id: cord-280621-tph5n7ak
type: ORG
entity:
  Veterinary Immunology and Immunopathology Isolation of Fidelity Variants of RNA Viruses and Characterization of Virus Mutation Frequency EasyModeller


id: cord-000318-wk2u2a9w
type: CELL_LINE
entity:
  trimer Human T cell Leukaemia virus types I and II Fields Virology Mechanisms of leukemogenesis induced by bovine leukemia virus prospects for novel antiretroviral therapies in human Detection and isolation of type C retrovirus particles


id: cord-000318-wk2u2a9w
type: DISEASE
entity:
  human T lymphotropic virus type 1 transmembrane glycoprotein helical hairpin structure Retrovirus envelope domain


id: cord-000318-wk2u2a9w
type: DISEASE
entity:
  human immunodeficiency virus type 1 Rev transactivator essential residues near the amino terminus HTLV I


id: cord-001109-xs7df6a7
type: ORG
entity:
  the Institutional Animal Care and Use Committee University of Pennsylvania Animal Welfare Assurance Number


id: cord-001257-t21l6i3f
type: PROTEIN
entity:
  anti Flag antibody 1 10 000 dilution and AP conjugated anti mouse antibody 1 10 000 His 6 AtRH2 or His 6 AtRH5 proteins


id: cord-001541-5d64esp4
type: PROTEIN
entity:
  nucleoprotein N polymeraseassociated phosphoprotein P matrix protein M glycoprotein G and RNA dependent RNA polymerase L 17 The genome features


id: cord-001541-5d64esp4
type: PROTEIN
entity:
  genus Novirhabdovirus Infectious haematopoietic necrosis virus ADB93801 Viral hemorrhagic septicaemia virus BAH57327 Hirame rhabdovirus ACO87999 and Snakehead rhabdovirus NP050585


id: cord-001541-5d64esp4
type: ORG
entity:
  Thomas G Paradkar Prasad N Holmes Edward C Tesh Robert B Vasilakis Nikos title Evolution of Genome Size and Complexity in


id: cord-001541-5d64esp4
type: WORK_OF_ART
entity:
  The Evolution and Emergence of RNA Viruses Virus Taxonomy Classification and Nomenclature of Viruses Ninth report of the International Committee on Taxonomy of Viruses


id: cord-001541-5d64esp4
type: WORK_OF_ART
entity:
  the Global Burden of Disease Study Virus Taxonomy Ninth Report of the International Committee on Taxonomy of Viruses Adelaide River


id: cord-001859-d62iuk72
type: PROTEIN
entity:
  goat anti mouse IgG Alexa Flour 488 goat anti mouse IgG Alexa Flour 488 goat anti rabbit IgG Alexa Flour 546 donkey anti mouse IgG


id: cord-001859-d62iuk72
type: DNA
entity:
  KSHV ORF50 promoter Merkel Cell Polyomavirus Small T Antigen Mediates Microtubule Destabilization To Promote Cell


id: cord-001859-d62iuk72
type: WORK_OF_ART
entity:
  Hsp70 Isoforms Are Essential for the Formation of Kaposi s Sarcoma Associated Herpesvirus Replication and Transcription Compartments


id: cord-001859-d62iuk72
type: ORG
entity:
  Lytic Reactivation and Represents a Novel Anti KSHV Target Interaction Network of Proteins Associated


id: cord-001859-d62iuk72
type: ORG
entity:
  C Terminal Signal Prevents Secretion of Luminal ER Proteins Functional Analysis of Hsp70 Inhibitors Adenosine


id: cord-000933-nn9gj0z1
type: PROTEIN
entity:
  Millipore anti F monoclonal ab43812 abcam anti P rabbit polyclonal 3 V Biosciences Menlo Park USA anti ZO 1 goat anti mouse goat anti rabbit


id: cord-312743-9e4yufo5
type: PROTEIN
entity:
  Anti A Antibody Titer Unexpected Receptor Functional Mimicry Elucidates Activation of Coronavirus Fusion Functional


id: cord-312743-9e4yufo5
type: WORK_OF_ART
entity:
  Serum Sickness to Xenosialitis Past Present and Future Significance of the Non human Sialic Acid Neu5Gc Absence of Neu5Gc and Presence of Anti Neu5Gc Antibodies in Humans


id: cord-000729-iq30z094
type: DNA
entity:
  17 mer defined primer sequence 59 GTTTCCCAGTAGGTCTCNNN NNNNN 39 and SuperScript III Reverse Transcriptase Invitrogen 8


id: cord-001636-khrx6rnh
type: PRODUCT
entity:
  Whole Genome Deep Sequencing of HIV 1 Reveals the Impact of Early Minor Variants Upon Immune Recognition During Acute Infection Mutational


id: cord-000018-amvlm09p
type: DNA
entity:
  human SOCS 3 siRNA sense 59 CCA AGA ACC UGC GCA UCC AdTdT 39 human SOCS 3 siRNA anti sense 59 UGG AUG CGC AGG UUC UUG GdTdT 39 see Table 1


id: cord-003441-810r5q03
type: PROTEIN
entity:
  retinoic acidinducible protein I RIG I mitochondrial antiviral signaling protein MAVS Tumor necrosis factor TNF receptor associated factor 3 TRAF3 TANK binding kinase 1 TBK1


id: cord-266078-h53zpjab
type: CELL_LINE
entity:
  B515 Foxp3 B710 CXCR3 G575 Ki67 G610 CTA 4 G666 CD127 G780 KLRG1 R660 NS4b Tet R710 CD45 R780 CD44 UV395 CD8 UV730 CD3 V450 CD73 V610 CD4 V655 CD25 V510 live dead Cells


id: cord-002835-qaogpxy9
type: PROTEIN
entity:
  nucleic acid binding transcription factor activity 34 7 structural molecule activity 25 or binding 22 2


id: cord-002835-qaogpxy9
type: PROTEIN
entity:
  mTORinteracting protein DEPTOR peripherin PRPH phosphatidylethanolamine binding protein 1 PEBP1 prohibitin PHB stomatin


id: cord-002835-qaogpxy9
type: WORK_OF_ART
entity:
  Proteomic Analysis of Human Brain Microvascular Endothelial Cells Reveals Differential Protein Expression in Response to Enterovirus 71 Infection Requirement


id: cord-287219-qxkwjkif
type: PROTEIN
entity:
  albumin ALB amylase AMY alanine aminotransferase ALT aspartate aminotransferase AST alkaline phosphatase ALP gamma glutamyltransferase GGT


id: cord-287219-qxkwjkif
type: WORK_OF_ART
entity:
  a Piccolo Point Of Care Blood Analyzer Abaxis Sunnyvale CA USA Flow Cytometry for Circulating Cell Populations


id: cord-287219-qxkwjkif
type: DISEASE
entity:
  HIV 1 influenza virus respiratory syncytial virus measles virus herpes simplex virus type 2 hepatitis C virus


id: cord-302340-pw2xqhwa
type: ORG
entity:
  the Centers for Disease Control and Prevention or the Agency for Toxic Substances and Disease Registry


id: cord-290253-hxxizipk
type: PROTEIN
entity:
  nuclear Adh SOD Amyrel RpL32 A random starting tree was used with a relaxed uncorrelated lognormal molecular clock Each of the partitions used a HKY substitution model


id: cord-002423-1u44tdrj
type: TAXON
entity:
  Flaviviridae Hepadnaviridae Herpesviridae Orthomyxoviridae Papillomaviridae Paramyxoviridae Parvoviridae


id: cord-001676-68y733y3
type: PROTEIN
entity:
  mouse anti STAT1 phospho Tyr701 mAb ab29045 abcam rabbit anti IRF3 phospho Ser396 mAb 4947 Cell Signaling and mouse anti βactin A2228


id: cord-001676-68y733y3
type: PROTEIN
entity:
  HRP conjugated antimouse IgG antibody GE Healthcare and HRP conjugated anti rabbit IgG antibody GE Healthcare Mouse lungs


id: cord-001676-68y733y3
type: WORK_OF_ART
entity:
  the Fundamental Guidelines for Proper Conduct of Animal Experiment and Related Activities in Academic Research Institutions


id: cord-001676-68y733y3
type: DISEASE
entity:
  H1N1 pandemic influenza virus 13 14 In contrast seasonal influenza viruses typically replicate less efficiently elicit more restrained inflammatory responses and are usually not associated with lethal infections


id: cord-000103-3zh8jmc2
type: PROTEIN
entity:
  Human Respirovirus Virulence Factor date 2009 09 18 journal PLoS Pathog DOI 10 1371 journal ppat 1000587 sha fe2000f280297c40bc53ce95d703a9ca6aac19fd doc_id 103 cord_uid 3zh8jmc2 A


id: cord-000103-3zh8jmc2
type: PROTEIN
entity:
  CT Upstate phospho eIF4E Ser209 Cell Signaling eIF4E Cell Signaling phospho S6 235 236 Ser235 236 Cell Signaling S6


id: cord-000246-mlhd9zbs
type: DISEASE
entity:
  T cell recognition T cell receptor recognition of a super bulged major histocompatibility complex class I bound peptide The fidelity occasional promiscuity and versatility of T cell receptor recognition Primary CTL response magnitude in mice is determined by the extent of naive T cell recruitment


id: cord-265947-7j9qqi8w
type: DNA
entity:
  monovalent MUNANA 2 4 Methylumbelliferyl α D N acetylneuraminic acid substrate Fig 1A S4A and S4B Fig


id: cord-265947-7j9qqi8w
type: PROTEIN
entity:
  glycophorin A A single sample method for determination of carbohydrate and protein contents glycoprotein bands


id: cord-001992-m4k20i7g
type: DNA
entity:
  primer L 845 5 GCAGG ACTTGAGGGCTATGA 3 Superscript III 18080 044 Invitrogen RNAse Out 10777 019 Invitrogen and 5 μL of RNA


id: cord-001992-m4k20i7g
type: PROTEIN
entity:
  Platinum Pfx DNA polymerase 11708 013 Invitrogen and primers L126 5 ATAG TACAAACAGGGCCGAAATCC 3 and L764 5 TTTGTTGGGTTCAGAGATAAGTG T 3


id: cord-001992-m4k20i7g
type: DNA
entity:
  TaqMan probe S 2295 5 6FAM CTG CAGGTTTCTCGC MGBNFQ 3 or L 5582 5 6FAM CTGAAGAATACCACCTATTAT ACCA MGBNFQ 3 and 12 5 μL


id: cord-001992-m4k20i7g
type: LAW
entity:
  Cellular Factors Required for Lassa Virus Budding Parallels Between Cytokinesis and Retroviral Budding A Role for the ESCRT Machinery Human ESCRT


id: cord-001992-m4k20i7g
type: ORG
entity:
  Biochemical Composition of Lymphocytic Choriomeningitis Virus Interfering Particles Equilibrium Sedimentation of Virus


id: cord-001992-m4k20i7g
type: ORG
entity:
  Iodinated Compounds Structural Components and Replication of Arenaviruses Properties of Defective Lymphocytic Choriomeningitis Virus Arenavirus Defective Interfering


id: cord-001992-m4k20i7g
type: WORK_OF_ART
entity:
  the Secretory Pathway Ubiquitin Depletion and Dominant Negative VPS4 Inhibit Rhabdovirus Budding without Affecting Alphavirus Budding Old World Arenaviruses


id: cord-001992-m4k20i7g
type: ORG
entity:
  Arrestin Related Proteins Implications for PPXY Dependent Budding Assessment of the Risk of Ebola Virus Transmission


id: cord-001992-m4k20i7g
type: ORG
entity:
  Dept of Health and Human Services Public Health Service Centers for Disease Control and Prevention National Institutes of Health Genomic Features of Attenuated


id: cord-001769-2sdg5ll7
type: ORG
entity:
  the Third Military Medical University Institutional Animal Care and Use Committee Supporting Information S1 Text Reagents


id: cord-253783-3a1qde41
type: ORG
entity:
  Byron Caughey National Institute of Allergy and Infectious Diseases National Institutes of Health Rocky Mountain Laboratories


id: cord-002608-zn7tm1ww
type: DNA
entity:
  primer pairs SINV nt9300F 5 GCACCGCCATCAAGCAATGTGTGGC 3 SINV nt9300R 5 CAATTTC CCTTGGGCCGTGTGGTCG 3 SINV nt10100F 5 TGTTCCAAATGTGCCACAGATACCG 3 SINV nt10100R 5 AATGT ACTCTTGGTTGGTGGAAGGC 3 SINV nt10400F 5 CAGCAGATTGCGCGTCTGACCATGC 3 SINV nt10400R 5 GACTCC GTTCACGTACACATCTAGG 3


id: cord-002608-zn7tm1ww
type: CHEMICAL
entity:
  amino terminal peptides of alfalfa mosaic virus coat protein The EMBO journal Coat protein binding sites on RNA 1 of alfalfa mosaic virus Removal of the N terminal part of alfalfa mosaic virus coat protein


id: cord-002142-tdgu9sr9
type: WORK_OF_ART
entity:
  An In Vivo Selection Identifies Listeria monocytogenes Genes Required to Sense the Intracellular Environment and Activate Virulence Factor Expression


id: cord-262753-jld1ygxt
type: PROTEIN
entity:
  primary antibodies rabbit anti eIF3a Cell Signaling 3411 rabbit anti eIF4E Cell Signaling 9742 rabbit anti 4E BP1 Cell Signaling 9452 rabbit anti phospho 4E BP1 Ser65 Cell Signaling 9451 rabbit anti phospho 4E BP1 Thr37 46 Cell Signaling 2855 mouse anti actin Millipore MAB1501 mouse anti actin


id: cord-262753-jld1ygxt
type: DISEASE
entity:
  influenza A virus a segmented negative strand RNA virus that replicates in the nucleus 50 of the total mRNA in the cytoplasm is viral 48 In this case however the viral endonuclease PA X degrades cellular mRNA which further contributes to the dominance of viral mRNA 66 67 For vaccinia virus a DNA virus that replicates entirely within the cytoplasm degradation


id: cord-000354-05lnj3w0
type: PROTEIN
entity:
  Spe4262 5 9GCCTTTCTTTATGTTTTTGGCACTAGTCATTTTACCG ATGTCACTCAG Spe4263 59 CTGAGTGACATCGGTAA AATGACTAGTGCCAAAAACATAAAGAAAGGC


id: cord-000354-05lnj3w0
type: GGP
entity:
  cytochalasin D cytochalasin B Blebbistatin 17 AA geldanamycin ML 7 ML 9 PP 2 5 N ethyl N isopropyl amiloride EIPA IPA 3


id: cord-268388-kkhuzf3p
type: ORG
entity:
  MERS CoV International Severe Acute Respiratory Emerging Infection Consortium ISARIC Clinical Decision Making Tool for Treatment of MERS CoV


id: cord-321874-eenaixp0
type: DISEASE
entity:
  Human immunodeficiency viruses avian influenza viruses and the recently described Middle East respiratory syndrome coronavirus


id: cord-013526-6fip93l2
type: DNA
entity:
  BTV1 S7 322F 5 0 GACGCCAGAGATACCTTTTAC 3 0 BTV1 S7 478R 5 0 CTTGAATCA TATCCGGACCAC 3 0 or segment 10 specific primers BTV1 S10 249F 5 CATTCGCATCG TACGCAGAA 3 BTV1 S10 464R 5 GCTTAAACGCCACGCTCATA 3


id: cord-013526-6fip93l2
type: ORG
entity:
  Extracellular Vesicle Heterogeneity Subpopulations Isolation Techniques and Diverse Functions in Cancer Progression Lysosome


id: cord-013481-3zwq67do
type: WORK_OF_ART
entity:
  The Significance of Type I Interferons in the Pathogenesis and Therapy of Human Immunodeficiency Virus 1 Infection


id: cord-013481-3zwq67do
type: WORK_OF_ART
entity:
  Brief Report Inflammatory Colonic Innate Lymphoid Cells Are Increased During Untreated HIV 1 Infection


id: cord-001655-uqw74ra0
type: PROTEIN
entity:
  cord 001655 uqw74ra0 authors Stenglein Mark D Jacobson Elliott R Chang Li Wen Sanders Chris Hawkins Michelle G Guzman David S M


id: cord-001655-uqw74ra0
type: DNA
entity:
  Meta Genomic Sequence Data G3 GenesGenomesGenetics MAFFT Multiple Sequence Alignment Software Version 7


id: cord-001655-uqw74ra0
type: DISEASE
entity:
  influenza A viruses HIV drug resistance Drug resistance virus fitness and HIV 1 mutagenesis Vaccine derived polioviruses and the endgame


id: cord-003169-bdw5ke4i
type: ORG
entity:
  fowl plague virus Mutation of the Second Sialic Acid Binding Site Resulting in Reduced Neuraminidase Activity


id: cord-003169-bdw5ke4i
type: DISEASE
entity:
  influenza A H3N2 viruses from 1968 to 2009 and its correspondence to haemagglutinin evolution Single hemagglutinin mutations


id: cord-001385-rb5vwolt
type: GGP
entity:
  TNFa Forward CTTGTGGCAGGGGCCACCAC Reverse CCATGCCGTTGGCCAGGAGG MCP 1 Forward TCACCTGCTGCTACTCATTCACCA Reverse AGCACAGACCTCTCTCTTGAGCTT


id: cord-254735-8reu45yz
type: CHEMICAL
entity:
  N terminal peptide trimming Structural basis for antigenic peptide precursor processing by the endoplasmic reticulum aminopeptidase ERAP1 Structure of aminopeptidase N from Escherichia coli


id: cord-286072-kgpvdb42
type: PROTEIN
entity:
  NF κB activator TBK1 TANK binding kinase 1 TRAF3 tumor necrosis factor receptor associated factor 3 TYK2 tyrosine kinase 2


id: cord-286072-kgpvdb42
type: ORG
entity:
  Plasmacytoid Dendritic Cells Development Regulation and Function Cell Cell Sensing of Viral Infection


id: cord-002530-ao7kenbx
type: DNA
entity:
  mouse anti CMV IE1 2 monoclonal antibody MAB8131 Millipore mouse anti CMV pp52 monoclonal antibody CH16 Santa Cruz Biotechnology mouse anti CMV pp28 monoclonal antibody CH19 Santa Cruz Biotechnology rabbit anti VCP polyclonal antibody H 120 Santa Cruz Biotechnology


id: cord-002530-ao7kenbx
type: PROTEIN
entity:
  mouse anti CMV IE2 monoclonal antibody 12E2 Santa Cruz Biotechnology rabbit anti VCP polyclonal antibody H 120 Santa Cruz Biotechnology


id: cord-315316-w7cn9iqp
type: PROTEIN
entity:
  goat anti mouse AlexaFluor 568 Donkey anti goat goat anti mouse and goat anti rabbit HRP conjugated antibodies


id: cord-286137-4cbh3u3z
type: WORK_OF_ART
entity:
  Prolonged Influenza Virus Shedding and Emergence of Antiviral Resistance in Immunocompromised Patients and Ferrets Oseltamivir


id: cord-278569-yr06jwm1
type: DISEASE
entity:
  influenza strain A HK 2 68 was performed as described 11 Mice were monitored once every 24 hours for symptoms of illness


id: cord-010762-c01wgg4v
type: DNA
entity:
  Full Length Japanese Encephalitis Virus NS5 Reveals a Conserved Methyltransferase Polymerase Interface A structural


id: cord-275307-d7htyfcl
type: GGP
entity:
  n 61 SOX n 178 B Frequency distribution histogram of log likelihood scores for the 20 nt surrounding


id: cord-314451-mqnqjn0c
type: DNA
entity:
  primer sets 59 44 59 TGATCCTCTGCAACCTGAGC and 39 SMAS MuM À 59 NNNCGTCTCACGGTAATAGTACCGTTGTCTGC and 59 SMAS MuM


id: cord-314451-mqnqjn0c
type: GGP
entity:
  aspartate aminotransferase alanine aminotransferase gamma glutamyl transferase creatine kinase urea nitrogen


id: cord-271692-60nlid3c
type: DNA
entity:
  hantavirus S segment 103 sequences 1201 bp M segment 71 sequences 3024 bp L segment 30 sequences 6519 bp partial L segment 32 330 bp mt cyt b gene 97 sequences 1140 bp Phylogenetic trees


id: cord-321957-ybtk9cp1
type: PROTEIN
entity:
  anti PKCδ Cell Signaling 9616 rabbit primary antibody Anti VEEV capsid BEI Resources NR 9403 goat primary antibody


id: cord-321957-ybtk9cp1
type: DNA
entity:
  gov_geo_query_acc cgi 3Facc 3DGSE132125 d Dw IBAg c OAG1LQNACBDguGvBeNj18Swhr9TMTjS x4O_KuapPgY r HAbXqcvDsSEpsf Q7 8oxF9_XjrPTRA0maMiRyITXKtM m mn_ESwz7wg_rIgqHybbAzs0w2PxZfCkymQnJ poVc3Tk s zwhEXJsjw_vFKC4S3I6Li0BN9V1KI7IUXIIaYN53S88 e


id: cord-321957-ybtk9cp1
type: DNA
entity:
  Integrated DNA Technologies primer pairs Table 2 and Taq Man probe 6 carboxyfluorescein TCGTCCGCACTGACATCTGTTGC carboxytetramethylrhodamine


id: cord-293164-i50cgkvq
type: PROTEIN
entity:
  virus Bat 09 mH1mN1 Bat09 mH1mN1 PB2 701D Bat09 mH1mN1 PB2 627K701D Bat09 mH1mN1 PB2 158G701D Bat09 mH1mN1 PB2 158G PR8


id: cord-293164-i50cgkvq
type: CHEMICAL
entity:
  N1ss H17ps H1ss N10ps N1ss H17ps H1ss N10ps N1ss H17ps H1ss N10ps N1ss H17ps H1ss N10ps N1ss H17ps H1ss N10ps N1ss H17ps H1ss N10ps N1ss H18ps H1ss N10ps N1ss H17ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H18ps H1ss N11ps N1ss H17ps H1ss N11ps


id: cord-000578-jhetyd9t
type: PROTEIN
entity:
  yeast 25 pyruvate decarboxylase Pdc1p 24 Cdc34p E2 ubiquitin conjugating enzyme 24 25 26 eukaryotic translation elongation factor 1A eEF1A 27 28 eukaryotic translation elongation factor 1Bgamma eEF1Bc 29


id: cord-000578-jhetyd9t
type: PROTEIN
entity:
  anti His 6 antibody 1 5 000 dilution and AP conjugated anti mouse antibody 1 5 000 DED1 66HA protein


id: cord-001082-sufwsu77
type: PROTEIN
entity:
  PV oncosuppression Primary antibodies Sec13 Sec23 calnexin Abcam ab22595 gm130 rab1A Sta Cruz Biotechnologies sc26541 rab6


id: cord-001082-sufwsu77
type: PROTEIN
entity:
  CKIIaE81A dominant negative 36 37 ERM family proteins FL EzT566A dominant negative FL RdxT564A dominant negative FL RdxT564E constitutiveactive FL Rdxdl P dominant


id: cord-001082-sufwsu77
type: WORK_OF_ART
entity:
  Bär Séverine Rommelaere Jean Nüesch Jürg P F title Vesicular Transport of Progeny Parvovirus Particles through ER


id: cord-260949-w2xuf15h
type: PROTEIN
entity:
  factor ANT adenine nucleotide translocase CK creatine kinase CypD cyclophilin D Cyt c cytochrome c HK hexokinase OXPHOS


id: cord-260949-w2xuf15h
type: PROTEIN
entity:
  mitochondrial matrix proteins e g cyclophilin D CypD IMS proteins e g creatine kinase CK OM proteins e g peripheral type benzodiazepine receptor PBR


id: cord-260949-w2xuf15h
type: CHEBI
entity:
  factor ANT adenine nucleotide translocase CK creatine kinase CypD cyclophilin D Cyt c cytochrome c HK


id: cord-263239-andje0wu
type: PROTEIN
entity:
  mouse monoclonal anti GM130 BD Biosciences rabbit polyclonal anti TGN46 Novus Biologicals mouse monoclonal anti ERGIC53 Enzo Life Sciences rabbit polyclonal anti Sec13


id: cord-263239-andje0wu
type: PROTEIN
entity:
  Singapore Singapore rabbit polyclonal anti calreticulin Sigma rabbit polyclonal anti HA Santa Cruz mouse monoclonal anti HA Abcam mouse monoclonal anti C Myc Sigma rabbit polyclonal anti myc Thermo Scientific mouse anti


id: cord-263239-andje0wu
type: PROTEIN
entity:
  primary antibodies rabbit polyclonal anti PI4KA Cell Signaling rabbit polyclonal anti PI4KB Upstate rabbit polyclonal anti OSBP ProteinTech rabbit polyclonal anti EMCV capsid kind gift


id: cord-000660-tsvzg0ax
type: DNA
entity:
  NCBI Entrez Nucleotide database Ifit2 NM_008332 Ifit1 NM_008331 Ifit3 NM_010501 Ifnb1 NM_010510 Ifnar1 NM_010508 Figure S1 Survival of wt and Ifit2 2 2


id: cord-000660-tsvzg0ax
type: CHEMICAL
entity:
  IFN alpha beta gene induction Essential role of IPS 1 in innate immune responses against RNA viruses


id: cord-000660-tsvzg0ax
type: CHEMICAL
entity:
  interferon alpha beta or gamma using oligonucleotide arrays Interferon inducible antiviral effectors A


id: cord-304747-ojyxs3cp
type: CELL_LINE
entity:
  NP_040785 budgerigar NP_848011 crow YP_529825 murine NP_041268 murine pneumotropic NP_041235 WU complete genome sequences


id: cord-304747-ojyxs3cp
type: DISEASE
entity:
  rhinoviruses coronaviruses influenzaviruses parainfluenzaviruses respiratory syncytial virus and adenoviruses


id: cord-304747-ojyxs3cp
type: DISEASE
entity:
  human rhinoviruses paramyxoviruses coronaviruses and bocavirus during acute respiratory tract infections


id: cord-323568-s0wmll4q
type: PROTEIN
entity:
  IBV spike ectodomain S1 receptor binding subunit S2 membrane fusion subunit GCN4 His 6 GCN4 trimerization tag


id: cord-303403-9th2jiq6
type: PROTEIN
entity:
  Biotech HRP conjugated goat anti mouse IgG H L CoWin Bioscience HRP conjugated goat anti rabbit IgG Santa Cruz PE conjugated goat anti mouse IgG


id: cord-002706-m3y35ozx
type: WORK_OF_ART
entity:
  Capsid Phosphorylation State and Hepadnavirus Virion Secretion Hepatitis B Virus Core Protein Phosphorylation Sites Affect Capsid Stability and Transient Exposure of the C


id: cord-267149-5twx9y5c
type: CELL_LINE
entity:
  human mouse rat feline canine Calomys callosus Cc MACV host Calomys musculinus Cm JUNV host or Viruses that cause hemorrhagic fever in humans which are only found in clade B


id: cord-325825-0lyt8gfq
type: PROTEIN
entity:
  chromoboxes b homeoboxes c general transcription factors d nuclear receptors e proteasome family members


id: cord-325825-0lyt8gfq
type: DISEASE
entity:
  herpes simplex virus type 1 compartment dynamics Construction and characterization of bacterial artificial chromosomes


id: cord-002125-anp238gu
type: WORK_OF_ART
entity:
  The Structure of Immature Virus Like Rous Sarcoma Virus Gag Particles Reveals a Structural Role for the p10 Domain in Assembly


id: cord-325326-2bbqz4o7
type: ORG
entity:
  the US Army Medical Research Institute of Infectious Disease Institutional Animal Care and Use Committee


id: cord-325192-italbsed
type: ORG
entity:
  influenza virus membrane fusion Fusion of Mature HIV 1 Particles Leads to Complete Release of a Gag GFP


id: cord-342825-s8ddek2f
type: GGP
entity:
  UbcH8 5 0 AACCTGTCCAGCGATGATGC 3 0 sense and 5 0 TGGTGCAAGGCTTCCAGTTC 3 0 anti sense for Herc5 5 0 GGGATGAA AGTGCTGAGGAG 3 0


id: cord-342825-s8ddek2f
type: DISEASE
entity:
  influenza B virus vaccinia virus and HCMV HCMV also encodes a tegument protein pUL48 that contains a deubiquitinating protease domain however its activity was specific for ubiquitin and did not cleave ISG15 90 In this study


id: cord-342825-s8ddek2f
type: DISEASE
entity:
  herpes simplex virus type 1 gB and virus egress require functional biogenesis of multivesicular bodies Herpes simplex


id: cord-350640-sz6xj5o3
type: CHEMICAL
entity:
  amyloid precursor protein processing The modification of mammalian membrane polyunsaturated fatty acid


id: cord-338400-30vl2hks
type: PROTEIN
entity:
  bilirubin alkaline phosphatase alanine transferase aspartate aminotransferase gamma glutamyltransferase


id: cord-338400-30vl2hks
type: PROTEIN
entity:
  helicase C pfam 00271 NS4A pfam 01006 NS4B pfam 01001 NS5A domain 1a zinc finger pfam 08300 domain 1b pfam 08301


id: cord-356115-vblgotjn
type: PROTEIN
entity:
  cysteine proteinase ExoN exonuclease HEL superfamily 1 helicase MT S adenosylmethioninedependent 29 O methyl transferase NeU endoribonuclease PL1 papain


id: cord-356115-vblgotjn
type: DISEASE
entity:
  infectious cycle the MHV A59 replicase transcriptase complex amplifies the genomic RNA and synthesizes subgenomic mRNAs Amplification of the genomic RNA involves full length negative strand templates and the synthesis of subgenomic mRNA involves subgenomelength negative strand templates 24 25 The structures engaged in the replication


id: cord-346053-mk1mzc5z
type: CL
entity:
  stem rust of wheat Recent research in medical epidemiology has elucidated the impact of pathogen ecology in environmental reservoirs on the evolution of novel or enhanced pathogen virulence In contrast the evolution of virulence in plant pathogens has been investigated from a predominantly agro centric


id: cord-264225-vzcfeh7t
type: DISEASE
entity:
  human immunodeficiency virus Analysis of human APOBEC3H haplotypes and anti human immunodeficiency virus type 1


id: cord-351952-lhhjax3s
type: ORG
entity:
  Standard Operating Procedure COVID 19 Virus Testing in NHS Laboratories Centers for Disease Control and Prevention Interim Guidelines for Collecting Handling and Testing Clinical Specimens for COVID 19


id: cord-289879-fh7ic0kp
type: DISEASE
entity:
  influenza A H1N1 pandemic public health laboratory and clinical perspectives Activation of influenza A viruses by trypsin treatment Enhancement of the infectivity of influenza A


id: cord-343221-e29of29o
type: GGP
entity:
  MHV A59 6 0 MHV H277A 5 6 p 0 024 left panel peak MHV A59 6 6 MHV H277A 6 0 p 0 016 right panel and significance


id: cord-343221-e29of29o
type: ORG
entity:
  Myeloid Cells Is Dependent on Basal Oas Gene Expression and Independent of Virus Induced Interferon Activation


id: cord-343221-e29of29o
type: ORG
entity:
  ORF1a Virology R Development Core Team A Language and Environment for Statistical Computing R Foundation for Statistical Computing Nonlinear


id: cord-355610-7xy4s483
type: PROTEIN
entity:
  mouse anti c Myc antibody Roche Alexa 488 conjugated goat anti mouse antibody Invitrogen and PE conjugated streptavidin Invitrogen


id: cord-325624-6anybxnk
type: CL
entity:
  stem and spinal cord grey matter were identified as Iba 1 positive microglia or infiltrating monocytes


id: cord-325624-6anybxnk
type: CL
entity:
  stem and spinal cord The enhanced demyelination phenotype is thus distinct from demyelination attributed to enhanced white matter infection by recombinant neuronotropic MHV variants


id: cord-355839-o0m71kvw
type: PROTEIN
entity:
  nuclear factor kappa B kinase subunit epsilon IRF3 interferon regulatory factor 3 IRF7 interferon regulatory factor 7 JNK c Jun N terminal kinase MAPK mitogen activated protein kinase


id: cord-355839-o0m71kvw
type: PROTEIN
entity:
  receptor TRAF3 tumor necrosis factor receptor associated factor 3 TRAF6 tumor necrosis factor receptor associated factor 6 TRAM toll


id: cord-355839-o0m71kvw
type: GGP
entity:
  kappa B kinase subunit epsilon IRF3 interferon regulatory factor 3 IRF7 interferon regulatory factor 7 JNK c Jun N terminal kinase MAPK mitogen activated protein kinase MAVS mitochondrial


id: cord-355839-o0m71kvw
type: ORG
entity:
  Immunopharmacol Identification of Respiratory Syncytial Virus Nonstructural Protein 2 Residues Essential for Exploitation of the Host Ubiquitin System and Inhibition of Innate Immune Responses Mutation


id: cord-355839-o0m71kvw
type: CHEMICAL
entity:
  IFN interferon IFNAR interferon alpha beta receptor IKKε inhibitor of nuclear factor kappa B kinase subunit epsilon IPO13 importin 13 IRF3 interferon regulatory factor 3 ISG IFN


id: cord-353337-o302vxqm
type: DNA
entity:
  eIF 4γ A General Chemical Ligation Approach Towards Isopeptide Linked Ubiquitin and Ubiquitin Like Assay Reagents Molecular basis of


id: cord-353337-o302vxqm
type: GGP
entity:
  IFN α β transcription 72 73 Unfortunately it has not been determined whether PL pro PLP2 cleaves RLR


id: cord-353337-o302vxqm
type: WORK_OF_ART
entity:
  Viral Reorganization of the Secretory Pathway Generates Distinct Organelles for RNA Replication Picornavirus


id: cord-328947-3l9ydspz
type: PROTEIN
entity:
  secondary messenger molecule 2 3 cyclic guanosine monophosphate GMP adenosine monophosphate AMP cyclic GMP AMP


id: cord-328947-3l9ydspz
type: GGP
entity:
  Pathog DOI 10 1371 journal ppat 1008999 sha b6f5c34d2e0a2a6acb4e4650c318bf9d7dc61be5 doc_id 328947 cord_uid 3l9ydspz


id: cord-260729-b12v3c8c
type: PROTEIN
entity:
  imgenex com mouse anti phospho STAT1 Clone ST1P 11A5 mouse IgG2a j Zymed Laboratories http www invitrogen com and rabbit anti IFN b Chemicon http


id: cord-260729-b12v3c8c
type: DISEASE
entity:
  acute respiratory syndrome coronavirus Severe acute respiratory syndrome coronavirus open reading frame ORF 3b ORF 6


id: cord-353703-u86ggw11
type: WORK_OF_ART
entity:
  The PERK Arm of the Unfolded Protein Response Negatively Regulates Transmissible Gastroenteritis Virus Replication by


id: cord-351548-jvl63652
type: DNA
entity:
  flank putative introns M116 F CTTCATCGGATTCGGAGGC R TGTTGTTGTCGACGTCTGATGTG m71 m75 F ATC TCCTCTGCCTCCGACCTC R CGATGTCATCTTGGAA TCCGACGA m72 m75 F CCGGATACGACCGTCAGC R CGATGTCATCTTGGAATCCGACGA


id: cord-351548-jvl63652
type: GGP
entity:
  Cxcl10 chemokine C X C motif ligand 10 8 5 Ccl5 chemokine C C motif ligand 5 RANTES 8 5 Trank1 tetratricopeptide repeat


id: cord-324928-cpryxa6p
type: DNA
entity:
  1 T position 3 4 TATT positions 21 25 and 14 ACAGCCGACCAATT positions 32 45 nucleotides Fig 10A Therefore


id: cord-324928-cpryxa6p
type: ORG
entity:
  RNA A Chikungunya Virus trans Replicase System Reveals the Importance of Delayed Nonstructural Polyprotein Processing for Efficient Replication Complex Formation


id: cord-324928-cpryxa6p
type: ORG
entity:
  a Novel Mechanism Leading to Increased Production of Interferon Inducing RNAs mBio Infection Pattern of Mayaro Virus


id: cord-324928-cpryxa6p
type: ORG
entity:
  Diptera Culicidae and Transmission Potential of the Virus in Mixed Infections With Chikungunya Virus Arbovirus


id: cord-331802-wo462anq
type: DNA
entity:
  RNA binding 15 16 RNA replication 17 18 19 membrane anchoring and rearrangement 20 21 autophagy inhibition 22 encapsidation


id: cord-334947-pa0p5dif
type: PROTEIN
entity:
  antimutator 483Y variant P 0 0006 Mann Whitney u test and significantly higher for the 483A G and W mutator variants


id: cord-334947-pa0p5dif
type: WORK_OF_ART
entity:
  Virus Molecular Determinants of Unique Vector Specificity Reside in Non Structural Protein 3 Conservation


id: cord-348799-qu4zin3o
type: CELL_TYPE
entity:
  human peripheral blood T lymphocyte Jurkat Jurkat human peripheral blood monocytic THP 1 cells THP 1


id: cord-348799-qu4zin3o
type: DNA
entity:
  ORF 5 GATCACTAATACGACTCACTATAGGTGGG CCTCAAAGGGTGAGGTTTTAGAGCTAGAAAT 3 In vitro Transcription T7 Kit 6140 TaKaRa


id: cord-348799-qu4zin3o
type: DISEASE
entity:
  Venezuelan equine encephalitis viruses Rhabdoviridae Vesicular Stomatitis virus VSV Orthomyxoviridae Influenza virus Retroviridae Human Immunodeficiency Type 1 virus Hepadnaviridae Hepatitis B


id: cord-353353-njvalb44
type: DNA
entity:
  144 bp 5 TGTC AGATGGTGTCAACAGTC AAA 3 and 5 TCATGGCGCACTTTCACATT 3 a 137 bp 5 ACAAATCTACAGCCAA ATTCCAAA 3 and 5 GTAGGGTATGCCT TTCTGGTCA A 3


id: cord-353353-njvalb44
type: CELL_LINE
entity:
  house HFL human embryonic lung fibroblast 3T3 mouse embryonic fibroblast ATCC CCL 92 cells RD human embryo rhabdomyosarcoma ATCC CCL 136 RK3E rat kidney ATCC CRL 1895


# bib

id: cord-000005-zt6i3o86
author: Bischof, Larry J.
date: 2008-10-10
txt: ./txt/cord-000005-zt6i3o86.txt
cache: ./cache/cord-000005-zt6i3o86.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2553261/
doi: 10.1371/journal.ppat.1000176
summary:
  elegans and mammalian cells to PFTs. We demonstrate for the first time that the ire-1 -xbp-1 arm of the UPR (and to a lesser extent the atf-6 arm) is functionally important for defense against a pathogenic attack since loss of this pathway leads to animals hypersensitive to PFT, but not to other toxic insults. As shown, a strong and specific increase in GFP expression in the intestine can be seen in the presence of the PFT ( Figure 1B , middle panel), consistent with activation of the ire-1-xbp-1 pathway by Cry5B. Consistent with activation of the ire-1-xbp-1 pathway by p38 MAPK in response to PFT but not unfolded proteins, the full induction of both mRNAs by Cry5B, but not tunicamycin, is dependent on sek-1 MAPKK. Specifically, we find that bacterial pore-forming toxins (PFTs) activate the ire-1-xbp-1 branch of the ER Unfolded Protein Response (UPR) in C.

title:
  Activation of the Unfolded Protein Response Is Required for Defenses against Bacterial Pore-Forming Toxin In Vivo

abstract:
  Pore-forming toxins (PFTs) constitute the single largest class of proteinaceous bacterial virulence factors and are made by many of the most important bacterial pathogens. Host responses to these toxins are complex and poorly understood. We find that the endoplasmic reticulum unfolded protein response (UPR) is activated upon exposure to PFTs both in Caenorhabditis elegans and in mammalian cells. Activation of the UPR is protective in vivo against PFTs since animals that lack either the ire-1-xbp-1 or the atf-6 arms of the UPR are more sensitive to PFT than wild-type animals. The UPR acts directly in the cells targeted by the PFT. Loss of the UPR leads to a normal response against unrelated toxins or a pathogenic bacterium, indicating its PFT-protective role is specific. The p38 mitogen-activated protein (MAPK) kinase pathway has been previously shown to be important for cellular defenses against PFTs. We find here that the UPR is one of the key downstream targets of the p38 MAPK pathway in response to PFT since loss of a functional p38 MAPK pathway leads to a failure of PFT to properly activate the ire-1-xbp-1 arm of the UPR. The UPR-mediated activation and response to PFTs is distinct from the canonical UPR-mediated response to unfolded proteins both in terms of its activation and functional sensitivities. These data demonstrate that the UPR, a fundamental intracellular pathway, can operate in intrinsic cellular defenses against bacterial attack.


id: cord-000007-h8upyzb1
title: Prevention of Cytotoxic T Cell Escape Using a Heteroclitic Subdominant Viral T Cell Determinant
author: Butler, Noah S.
date: 2008-10-24
txt: ./txt/cord-000007-h8upyzb1.txt
cache: ./cache/cord-000007-h8upyzb1.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2563037/
doi: 10.1371/journal.ppat.1000186
summary:
  Recombinant viruses encoding this engineered determinant primed CTL responses that also reacted to the wildtype epitope with significantly higher functional avidity, and protected against selection of virus mutated at a second CTL determinant and consequent disease progression in persistently infected mice. Enhancement strategies, which result in augmented responses to the native, subdominant epitope, have been described for both MHC class I and class II-restricted determinants, whereby the most common approaches involve generating a series of conserved and non-conserved mutations at MHC anchor residues, followed by an empiric determination of whether each individual substitution augments T cell effector function [12] . Immunization with the modified peptide resulted in an improved response to the native S598 epitope, demonstrating a true heteroclitic effect and suggesting that this strategy may have clinical applications for reducing viral titer and preventing CTL escape during chronic infections. Seven days following virus infection, brains were harvested from mice and the frequencies of epitope-specific CD8 T cells were determined by ex vivo stimulation and intracellular cytokine staining as described above.

abstract:
  High affinity antigen-specific T cells play a critical role during protective immune responses. Epitope enhancement can elicit more potent T cell responses and can subsequently lead to a stronger memory pool; however, the molecular basis of such enhancement is unclear. We used the consensus peptide-binding motif for the Major Histocompatibility Complex molecule H-2K(b) to design a heteroclitic version of the mouse hepatitis virus-specific subdominant S598 determinant. We demonstrate that a single amino acid substitution at a secondary anchor residue (Q to Y at position 3) increased the stability of the engineered determinant in complex with H-2K(b). The structural basis for this enhanced stability was associated with local alterations in the pMHC conformation as a result of the Q to Y substitution. Recombinant viruses encoding this engineered determinant primed CTL responses that also reacted to the wildtype epitope with significantly higher functional avidity, and protected against selection of virus mutated at a second CTL determinant and consequent disease progression in persistently infected mice. Collectively, our findings provide a basis for the enhanced immunogenicity of an engineered determinant that will serve as a template for guiding the development of heteroclitic T cell determinants with applications in prevention of CTL escape in chronic viral infections as well as in tumor immunity.


id: cord-000018-amvlm09p
title: Influenza A Virus Inhibits Type I IFN Signaling via NF-κB-Dependent Induction of SOCS-3 Expression
author: Pauli, Eva-K.
date: 2008-11-07
txt: ./txt/cord-000018-amvlm09p.txt
cache: ./cache/cord-000018-amvlm09p.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2572141/
doi: 10.1371/journal.ppat.1000196
summary:
  Our study was based on the observation that in cells that were infected with influenza A virus and subsequently stimulated with IFNα/β, phosphorylation of the signal transducer and activator of transcription protein 1 (STAT1) was strongly reduced. This direct viral induction of SOCS-3 mRNA and protein expression appears to be relevant for suppression of the antiviral response since in SOCS-3 deficient cells a sustained phosphorylation of STAT1 correlated with elevated expression of type I IFN-dependent genes. These results are consistent with data gained from previous experiments in Vero cells ( Figure 1E ) and indicate that neither induction of SOCS-3 mRNA nor inhibition of STAT phosphorylation is dependent on virus-induced type I IFN expression. To answer the question whether enhanced STAT phosphorylation in SOCS-3 deficient cells would also lead to enhanced expression of ISGs, total RNA was isolated at different time points p.i. from infected wild type and knock out cells and monitored for induction of SP110, IRF-1 and OAS1 ( Figure 7E, 7F and 7G ).

abstract:
  The type I interferon (IFN) system is a first line of defense against viral infections. Viruses have developed various mechanisms to counteract this response. So far, the interferon antagonistic activity of influenza A viruses was mainly observed on the level of IFNβ gene induction via action of the viral non-structural protein 1 (NS1). Here we present data indicating that influenza A viruses not only suppress IFNβ gene induction but also inhibit type I IFN signaling through a mechanism involving induction of the suppressor of cytokine signaling-3 (SOCS-3) protein. Our study was based on the observation that in cells that were infected with influenza A virus and subsequently stimulated with IFNα/β, phosphorylation of the signal transducer and activator of transcription protein 1 (STAT1) was strongly reduced. This impaired STAT1 activation was not due to the action of viral proteins but rather appeared to be induced by accumulation of viral 5′ triphosphate RNA in the cell. SOCS proteins are potent endogenous inhibitors of Janus kinase (JAK)/STAT signaling. Closer examination revealed that SOCS-3 but not SOCS-1 mRNA levels increase in an RNA- and nuclear factor kappa B (NF-κB)-dependent but type I IFN-independent manner early in the viral replication cycle. This direct viral induction of SOCS-3 mRNA and protein expression appears to be relevant for suppression of the antiviral response since in SOCS-3 deficient cells a sustained phosphorylation of STAT1 correlated with elevated expression of type I IFN-dependent genes. As a consequence, progeny virus titers were reduced in SOCS-3 deficient cells or in cells were SOCS-3 expression was knocked-down by siRNA. These data provide the first evidence that influenza A viruses suppress type I IFN signaling on the level of JAK/STAT activation. The inhibitory effect is at least in part due to the induction of SOCS-3 gene expression, which results in an impaired antiviral response.


id: cord-000025-6zrv687k
author: Chachu, Karen A.
date: 2008-12-12
txt: ./txt/cord-000025-6zrv687k.txt
cache: ./cache/cord-000025-6zrv687k.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2587711/
doi: 10.1371/journal.ppat.1000236
summary:
  In the absence of a cell culture system for HuNV, virus like particles (VLPs) that assemble when the viral capsid protein is expressed have been important for evaluating norovirus immune responses [20] [21] [22] [23] . To determine the role of T cells in clearance of acute MNV infection we inoculated WT, MHC Class II-/-, and MHC Class I6b2M-/-mice orally with MNV1.CW3 and measured viral titers in the distal ileum and MLN three, five, seven and 21 days post-infection ( Figure 4B-4E) . Together, these data from primary challenges of non-immune mice lacking antigen presenting molecules or depleted of specific T cell subsets demonstrated that CD4 T cells are important for efficient MNV clearance in the distal ileum especially at days three Lack of any of the three components of the adaptive response: B cells, CD4 T cells, or CD8 T cells significantly diminished vaccine effects generated by either live virus or VP1 capsid protein immunization, and delayed viral clearance during primary infection.

title:
  Immune Mechanisms Responsible for Vaccination against and Clearance of Mucosal and Lymphatic Norovirus Infection

abstract:
  Two cardinal manifestations of viral immunity are efficient clearance of acute infection and the capacity to vaccinate against secondary viral exposure. For noroviruses, the contributions of T cells to viral clearance and vaccination have not been elucidated. We report here that both CD4 and CD8 T cells are required for efficient clearance of primary murine norovirus (MNV) infection from the intestine and intestinal lymph nodes. Further, long-lasting protective immunity was generated by oral live virus vaccination. Systemic vaccination with the MNV capsid protein also effectively protected against mucosal challenge, while vaccination with the capsid protein of the distantly related human Lordsdale virus provided partial protection. Fully effective vaccination required a broad immune response including CD4 T cells, CD8 T cells, and B cells, but the importance of specific immune cell types varied between the intestine and intestinal lymph nodes. Perforin, but not interferon gamma, was required for clearance of MNV infection by adoptively transferred T lymphocytes from vaccinated hosts. These studies prove the feasibility of both mucosal and systemic vaccination against mucosal norovirus infection, demonstrate tissue specificity of norovirus immune cells, and indicate that efficient vaccination strategies should induce potent CD4 and CD8 T cell responses.


id: cord-000103-3zh8jmc2
author: Caignard, Grégory
date: 2009-09-18
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2736567/
doi: 10.1371/journal.ppat.1000587
title:
  Differential Regulation of Type I Interferon and Epidermal Growth Factor Pathways by a Human Respirovirus Virulence Factor

abstract:
  A number of paramyxoviruses are responsible for acute respiratory infections in children, elderly and immuno-compromised individuals, resulting in airway inflammation and exacerbation of chronic diseases like asthma. To understand the molecular pathogenesis of these infections, we searched for cellular targets of the virulence protein C of human parainfluenza virus type 3 (hPIV3-C). We found that hPIV3-C interacts directly through its C-terminal domain with STAT1 and GRB2, whereas C proteins from measles or Nipah viruses failed to do so. Binding to STAT1 explains the previously reported capacity of hPIV3-C to block type I interferon signaling, but the interaction with GRB2 was unexpected. This adaptor protein bridges Epidermal Growth Factor (EGF) receptor to MAPK/ERK pathway, a signaling cascade recently found to be involved in airway inflammatory response. We report that either hPIV3 infection or transient expression of hPIV3-C both increase cellular response to EGF, as assessed by Elk1 transactivation and phosphorylation levels of ERK1/2, 40S ribosomal subunit protein S6 and translation initiation factor 4E (eIF4E). Furthermore, inhibition of MAPK/ERK pathway with U0126 prevented viral protein expression in infected cells. Altogether, our data provide molecular basis to explain the role of hPIV3-C as a virulence factor and determinant of pathogenesis and demonstrate that Paramyxoviridae have evolved a single virulence factor to block type I interferon signaling and to boost simultaneous cellular response to growth factors.


id: cord-000127-tkxpjlyn
title: Immunoglobulin Superfamily Virus Receptors and the Evolution of Adaptive Immunity
author: Dermody, Terence S.
date: 2009-11-26
txt: ./txt/cord-000127-tkxpjlyn.txt
cache: ./cache/cord-000127-tkxpjlyn.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2777377/
doi: 10.1371/journal.ppat.1000481
abstract: nan
summary:
  The immunoglobulin superfamily (IgSF) of molecules contains several members that are expressed at the cell surface, bind diverse ligands, and contribute to a variety of cellular activities, including adhesion and immune responses. While structural information about com-plex formation is lacking for the IgSF receptors carcinoembryonic antigen-related cell adhesion molecule, nectin-1, nectin-2, and signaling lymphocyte-activation molecule (SLAM), biochemical studies also implicate their respective D1 domains in virus binding [11] [12] [13] [14] . The evolution of JAM family members prior to the biochemical means to efficiently and extensively diversify antigen receptor genes, along with the structural similarities in the binding surfaces of virus receptors and immunoglobulins, provides strong support for the contention that viruses and perhaps other pathogens that engage IgSF receptors contributed to the selection of humoral mediators of adaptive immunity.


id: cord-000164-094d0rn6
title: IPS-1 Is Essential for the Control of West Nile Virus Infection and Immunity
author: Suthar, Mehul S.
date: 2010-02-05
txt: ./txt/cord-000164-094d0rn6.txt
cache: ./cache/cord-000164-094d0rn6.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2816698/
doi: 10.1371/journal.ppat.1000757
summary:
  Infection of cultured bone-marrow (BM) derived dendritic cells (DCs), macrophages (Macs), and primary cortical neurons showed that the IPS-1-dependent RLR signaling was essential for triggering IFN defenses and controlling virus replication in these key target cells of infection. Intriguingly, infected IPS-1(−/−) mice displayed uncontrolled inflammation that included elevated systemic type I IFN, proinflammatory cytokine and chemokine responses, increased numbers of inflammatory DCs, enhanced humoral responses marked by complete loss of virus neutralization activity, and increased numbers of virus-specific CD8+ T cells and non-specific immune cell proliferation in the periphery and in the CNS. To define the role of IPS-1 in controlling virus replication and innate immunity in myeloid cells, we analyzed WNV infection and host responses in primary bone marrow-derived DC and Mw recovered from wild type and IPS-1 2/2 mice.

abstract:
  The innate immune response is essential for controlling West Nile virus (WNV) infection but how this response is propagated and regulates adaptive immunity in vivo are not defined. Herein, we show that IPS-1, the central adaptor protein to RIG-I-like receptor (RLR) signaling, is essential for triggering of innate immunity and for effective development and regulation of adaptive immunity against pathogenic WNV. IPS-1(−/−) mice exhibited increased susceptibility to WNV infection marked by enhanced viral replication and dissemination with early viral entry into the CNS. Infection of cultured bone-marrow (BM) derived dendritic cells (DCs), macrophages (Macs), and primary cortical neurons showed that the IPS-1-dependent RLR signaling was essential for triggering IFN defenses and controlling virus replication in these key target cells of infection. Intriguingly, infected IPS-1(−/−) mice displayed uncontrolled inflammation that included elevated systemic type I IFN, proinflammatory cytokine and chemokine responses, increased numbers of inflammatory DCs, enhanced humoral responses marked by complete loss of virus neutralization activity, and increased numbers of virus-specific CD8+ T cells and non-specific immune cell proliferation in the periphery and in the CNS. This uncontrolled inflammatory response was associated with a lack of regulatory T cell expansion that normally occurs during acute WNV infection. Thus, the enhanced inflammatory response in the absence of IPS-1 was coupled with a failure to protect against WNV infection. Our data define an innate/adaptive immune interface mediated through IPS-1-dependent RLR signaling that regulates the quantity, quality, and balance of the immune response to WNV infection.


id: cord-000232-boto4h8x
title: Bid Regulates the Pathogenesis of Neurotropic Reovirus
author: Danthi, Pranav
date: 2010-07-01
txt: ./txt/cord-000232-boto4h8x.txt
cache: ./cache/cord-000232-boto4h8x.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2895667/
doi: 10.1371/journal.ppat.1000980
summary:
  Blockade of NF-kB signaling, which diminishes apoptosis induction by reovirus [8, 28] , prevents cleavage of Bid. In comparison to wild-type mice, Bid-deficient mice display diminished susceptibility to reovirus-induced CNS disease following either peroral (PO) or intracranial (IC) inoculation. To directly test whether Bid is required for apoptosis induction following reovirus infection, we compared reovirus-induced apoptosis in wild-type and Bid-deficient MEFs. For these experiments, MEFs were infected with T3D, and apoptosis was assessed by chemiluminescent measurement of the activity of caspase-3 and caspase-7, which serve as effector caspases for both the extrinsic and intrinsic apoptotic pathways ( Figure 2A ). Increase in caspase-3/7 activity following treatment of each cell type with a broad-spectrum protein kinase inhibitor, staurosporine, was equivalent (,5-fold), demonstrating that although Bid-deficient cells possess functional death-signaling pathways, they resist apoptosis induction by reovirus. Analogous to treatment with TNFa, a control NF-kB agonist, reovirus infection resulted in equivalent (,2-to 3-fold) activation of NF-kB-driven gene expression in wild-type and Bid-deficient cells ( Figure 3A ).

abstract:
  Reovirus infection leads to apoptosis in both cultured cells and the murine central nervous system (CNS). NF-κB-driven transcription of proapoptotic cellular genes is required for the effector phase of the apoptotic response. Although both extrinsic death-receptor signaling pathways and intrinsic pathways involving mitochondrial injury are implicated in reovirus-induced apoptosis, mechanisms by which either of these pathways are activated and their relationship to NF-κB signaling following reovirus infection are unknown. The proapoptotic Bcl-2 family member, Bid, is activated by proteolytic cleavage following reovirus infection. To understand how reovirus integrates host signaling circuits to induce apoptosis, we examined proapoptotic signaling following infection of Bid-deficient cells. Although reovirus growth was not affected by the absence of Bid, cells lacking Bid failed to undergo apoptosis. Furthermore, we found that NF-κB activation is required for Bid cleavage and subsequent proapoptotic signaling. To examine the functional significance of Bid-dependent apoptosis in reovirus disease, we monitored fatal encephalitis caused by reovirus in the presence and absence of Bid. Survival of Bid-deficient mice was significantly enhanced in comparison to wild-type mice following either peroral or intracranial inoculation of reovirus. Decreased reovirus virulence in Bid-null mice was accompanied by a reduction in viral yield. These findings define a role for NF-κB-dependent cleavage of Bid in the cell death program initiated by viral infection and link Bid to viral virulence.


id: cord-000246-mlhd9zbs
author: Valkenburg, Sophie A.
date: 2010-08-12
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2920842/
doi: 10.1371/journal.ppat.1001039
title:
  Protective Efficacy of Cross-Reactive CD8(+) T Cells Recognising Mutant Viral Epitopes Depends on Peptide-MHC-I Structural Interactions and T Cell Activation Threshold

abstract:
  Emergence of a new influenza strain leads to a rapid global spread of the virus due to minimal antibody immunity. Pre-existing CD8(+) T-cell immunity directed towards conserved internal viral regions can greatly ameliorate the disease. However, mutational escape within the T cell epitopes is a substantial issue for virus control and vaccine design. Although mutations can result in a loss of T cell recognition, some variants generate cross-reactive T cell responses. In this study, we used reverse genetics to modify the influenza NP(336–374) peptide at a partially-solvent exposed residue (N->A, NPN3A mutation) to assess the availability, effectiveness and mechanism underlying influenza-specific cross-reactive T cell responses. The engineered virus induced a diminished CD8(+) T cell response and selected a narrowed T cell receptor (TCR) repertoire within two Vβ regions (Vβ8.3 and Vβ9). This can be partially explained by the H-2D(b)NPN3A structure that showed a loss of several contacts between the NPN3A peptide and H-2D(b), including a contact with His155, a position known to play an important role in mediating TCR-pMHC-I interactions. Despite these differences, common cross-reactive TCRs were detected in both the naïve and immune NPN3A-specific TCR repertoires. However, while the NPN3A epitope primes memory T-cells that give an equivalent recall response to the mutant or wild-type (wt) virus, both are markedly lower than wt->wt challenge. Such decreased CD8(+) responses elicited after heterologous challenge resulted in delayed viral clearance from the infected lung. Furthermore, mice first exposed to the wt virus give a poor, low avidity response following secondary infection with the mutant. Thus, the protective efficacy of cross-reactive CD8(+) T cells recognising mutant viral epitopes depend on peptide-MHC-I structural interactions and functional avidity. Our study does not support vaccine strategies that include immunization against commonly selected cross-reactive variants with mutations at partially-solvent exposed residues that have characteristics comparable to NPN3A.


id: cord-000318-wk2u2a9w
author: Lamb, Daniel
date: 2011-02-03
txt: ./txt/cord-000318-wk2u2a9w.txt
cache: ./cache/cord-000318-wk2u2a9w.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3033372/
doi: 10.1371/journal.ppat.1001268
summary:
  Our data indicate that, through electrostatic interactions with a chloride ion, the asparagine residues promote assembly and profoundly stabilize the fusion-active structures that are required for viral envelope-mediated membrane fusion. Moreover, the BLV TM structure also reveals a charge-surrounded hydrophobic pocket on the central coiled coil and interactions with basic residues that cluster around this pocket are critical to membrane fusion and form a target for peptide inhibitors of envelope function. Charge-surrounded pockets and electrostatic interactions with small ions are common among class-1 fusion proteins, suggesting that small molecules that specifically target such motifs should prevent assembly of the trimer-of-hairpins and be of value as therapeutic inhibitors of viral entry. The BLV crystal structure and our analysis of HTLV-1 and BLV envelope-mediated membrane fusion reveal an important role for electrostatic interactions in binding of the LHR to the grooves of the coiled coil.

title:
  Charge-Surrounded Pockets and Electrostatic Interactions with Small Ions Modulate the Activity of Retroviral Fusion Proteins

abstract:
  Refolding of viral class-1 membrane fusion proteins from a native state to a trimer-of-hairpins structure promotes entry of viruses into cells. Here we present the structure of the bovine leukaemia virus transmembrane glycoprotein (TM) and identify a group of asparagine residues at the membrane-distal end of the trimer-of-hairpins that is strikingly conserved among divergent viruses. These asparagines are not essential for surface display of pre-fusogenic envelope. Instead, substitution of these residues dramatically disrupts membrane fusion. Our data indicate that, through electrostatic interactions with a chloride ion, the asparagine residues promote assembly and profoundly stabilize the fusion-active structures that are required for viral envelope-mediated membrane fusion. Moreover, the BLV TM structure also reveals a charge-surrounded hydrophobic pocket on the central coiled coil and interactions with basic residues that cluster around this pocket are critical to membrane fusion and form a target for peptide inhibitors of envelope function. Charge-surrounded pockets and electrostatic interactions with small ions are common among class-1 fusion proteins, suggesting that small molecules that specifically target such motifs should prevent assembly of the trimer-of-hairpins and be of value as therapeutic inhibitors of viral entry.


id: cord-000354-05lnj3w0
author: de Vries, Erik
date: 2011-03-31
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3068995/
doi: 10.1371/journal.ppat.1001329
title:
  Dissection of the Influenza A Virus Endocytic Routes Reveals Macropinocytosis as an Alternative Entry Pathway

abstract:
  Influenza A virus (IAV) enters host cells upon binding of its hemagglutinin glycoprotein to sialylated host cell receptors. Whereas dynamin-dependent, clathrin-mediated endocytosis (CME) is generally considered as the IAV infection pathway, some observations suggest the occurrence of an as yet uncharacterized alternative entry route. By manipulating entry parameters we established experimental conditions that allow the separate analysis of dynamin-dependent and -independent entry of IAV. Whereas entry of IAV in phosphate-buffered saline could be completely inhibited by dynasore, a specific inhibitor of dynamin, a dynasore-insensitive entry pathway became functional in the presence of fetal calf serum. This finding was confirmed with the use of small interfering RNAs targeting dynamin-2. In the presence of serum, both IAV entry pathways were operational. Under these conditions entry could be fully blocked by combined treatment with dynasore and the amiloride derivative EIPA, the hallmark inhibitor of macropinocytosis, whereas either drug alone had no effect. The sensitivity of the dynamin-independent entry pathway to inhibitors or dominant-negative mutants affecting actomyosin dynamics as well as to a number of specific inhibitors of growth factor receptor tyrosine kinases and downstream effectors thereof all point to the involvement of macropinocytosis in IAV entry. Consistently, IAV particles and soluble FITC-dextran were shown to co-localize in cells in the same vesicles. Thus, in addition to the classical dynamin-dependent, clathrin-mediated endocytosis pathway, IAV enters host cells by a dynamin-independent route that has all the characteristics of macropinocytosis.


id: cord-000409-lpf9lpky
author: Chen, Yongwen
date: 2011-07-07
txt: ./txt/cord-000409-lpf9lpky.txt
cache: ./cache/cord-000409-lpf9lpky.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3131267/
doi: 10.1371/journal.ppat.1001347
summary:
  Fluorescence double-staining revealed that FGL2 and PD-1 were not co-expressed on the same cells, while quantitative RT-PCR demonstrated that higher levels of IFN-γ and TNF-α mRNA transcription occurred in PD-1-deficient mice in response to MHV-3 infection. In all, these results demonstrated that PD-1 deficiency led to enhanced pathological damage by MHV-3 in the liver, spleen, lymph node and thymus, where higher levels of PD-1-positive cells were found after infection. Results showed that the expression of FGL2 was principally associated with CD31-positive endothelial cells, CD68-positive macrophages and CD11c-positive DCs. Surprisingly, significantly higher levels of FGL2 were observed after infection in all of PD-1-deficient organs, including the liver, thymus, spleen, lymph nodes, and serum than that in those from WT littermates. However, the number of Foxp3-positive cells in the liver, spleen, lymph node or thymus was not significantly different between PD-1-deficient mice and their WT littermates after 72 h of MHV-3 infection (Fig. S3) .

title:
  Programmed Death (PD)-1-Deficient Mice Are Extremely Sensitive to Murine Hepatitis Virus Strain-3 (MHV-3) Infection

abstract:
  The inhibitory receptor programmed death-1 (PD-1) has the capacity to maintain peripheral tolerance and limit immunopathological damage; however, its precise role in fulminant viral hepatitis (FH) has yet to be described. Here, we investigated the functional mechanisms of PD-1 as related to FH pathogenesis induced by the murine hepatitis virus strain-3 (MHV-3). High levels of PD-1-positive CD4(+), CD8(+) T cells, NK cells and macrophages were observed in liver, spleen, lymph node and thymus tissues following MHV-3 infection. PD-1-deficient mice exhibited significantly higher expression of the effector molecule which initiates fibrinogen deposition, fibrinogen-like protein 2 (FGL2), than did their wild-type (WT) littermates. As a result, more severe tissue damage was produced and mortality rates were higher. Fluorescence double-staining revealed that FGL2 and PD-1 were not co-expressed on the same cells, while quantitative RT-PCR demonstrated that higher levels of IFN-γ and TNF-α mRNA transcription occurred in PD-1-deficient mice in response to MHV-3 infection. Conversely, in vivo blockade of IFN-γ and TNF-α led to efficient inhibition of FGL2 expression, greatly attenuated the development of tissue lesions, and ultimately reduced mortality. Thus, the up-regulation of FGL2 in PD-1-deficient mice was determined to be mediated by IFN-γ and TNF-α. Taken together, our results suggest that PD-1 signaling plays an essential role in decreasing the immunopathological damage induced by MHV-3 and that manipulation of this signal might be a useful strategy for FH immunotherapy.


id: cord-000578-jhetyd9t
title: A Co-Opted DEAD-Box RNA Helicase Enhances Tombusvirus Plus-Strand Synthesis
author: Kovalev, Nikolay
date: 2012-02-16
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3280988/
doi: 10.1371/journal.ppat.1002537
abstract:
  Replication of plus-strand RNA viruses depends on recruited host factors that aid several critical steps during replication. In this paper, we show that an essential translation factor, Ded1p DEAD-box RNA helicase of yeast, directly affects replication of Tomato bushy stunt virus (TBSV). To separate the role of Ded1p in viral protein translation from its putative replication function, we utilized a cell-free TBSV replication assay and recombinant Ded1p. The in vitro data show that Ded1p plays a role in enhancing plus-strand synthesis by the viral replicase. We also find that Ded1p is a component of the tombusvirus replicase complex and Ded1p binds to the 3′-end of the viral minus-stranded RNA. The data obtained with wt and ATPase deficient Ded1p mutants support the model that Ded1p unwinds local structures at the 3′-end of the TBSV (−)RNA, rendering the RNA compatible for initiation of (+)-strand synthesis. Interestingly, we find that Ded1p and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), which is another host factor for TBSV, play non-overlapping functions to enhance (+)-strand synthesis. Altogether, the two host factors enhance TBSV replication synergistically by interacting with the viral (−)RNA and the replication proteins. In addition, we have developed an in vitro assay for Flock house virus (FHV), a small RNA virus of insects, that also demonstrated positive effect on FHV replicase activity by the added Ded1p helicase. Thus, two small RNA viruses, which do not code for their own helicases, seems to recruit a host RNA helicase to aid their replication in infected cells.


id: cord-000660-tsvzg0ax
title: Interferon-Induced Ifit2/ISG54 Protects Mice from Lethal VSV Neuropathogenesis
author: Fensterl, Volker
date: 2012-05-17
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3355090/
doi: 10.1371/journal.ppat.1002712
abstract:
  Interferon protects mice from vesicular stomatitis virus (VSV) infection and pathogenesis; however, it is not known which of the numerous interferon-stimulated genes (ISG) mediate the antiviral effect. A prominent family of ISGs is the interferon-induced with tetratricopeptide repeats (Ifit) genes comprising three members in mice, Ifit1/ISG56, Ifit2/ISG54 and Ifit3/ISG49. Intranasal infection with a low dose of VSV is not lethal to wild-type mice and all three Ifit genes are induced in the central nervous system of the infected mice. We tested their potential contributions to the observed protection of wild-type mice from VSV pathogenesis, by taking advantage of the newly generated knockout mice lacking either Ifit2 or Ifit1. We observed that in Ifit2 knockout (Ifit2 (−/−)) mice, intranasal VSV infection was uniformly lethal and death was preceded by neurological signs, such as ataxia and hind limb paralysis. In contrast, wild-type and Ifit1 (−/−) mice were highly protected and survived without developing such disease. However, when VSV was injected intracranially, virus replication and survival were not significantly different between wild-type and Ifit2(−/−) mice. When administered intranasally, VSV entered the central nervous system through the olfactory bulbs, where it replicated equivalently in wild-type and Ifit2 (−/−) mice and induced interferon-β. However, as the infection spread to other regions of the brain, VSV titers rose several hundred folds higher in Ifit2 (−/−) mice as compared to wild-type mice. This was not caused by a broadened cell tropism in the brains of Ifit2 (−/−) mice, where VSV still replicated selectively in neurons. Surprisingly, this advantage for VSV replication in the brains of Ifit2(−/−) mice was not observed in other organs, such as lung and liver. Pathogenesis by another neurotropic RNA virus, encephalomyocarditis virus, was not enhanced in the brains of Ifit2 (−/−) mice. Our study provides a clear demonstration of tissue-, virus- and ISG-specific antiviral action of interferon.


id: cord-000729-iq30z094
title: Cedar Virus: A Novel Henipavirus Isolated from Australian Bats
author: Marsh, Glenn A.
date: 2012-08-02
txt: ./txt/cord-000729-iq30z094.txt
cache: ./cache/cord-000729-iq30z094.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3410871/
doi: 10.1371/journal.ppat.1002836
summary:
  The genome size (18,162 nt) and organization of CedPV is very similar to that of HeV and NiV; its nucleocapsid protein displays antigenic cross-reactivity with henipaviruses; and it uses the same receptor molecule (ephrinB2) for entry during infection. Preliminary challenge studies with CedPV in ferrets and guinea pigs, both susceptible to infection and disease with known henipaviruses, confirmed virus replication and production of neutralizing antibodies although clinical disease was not observed. As part of our on-going field studies on HeV genetic diversity and infection dynamics in the Queensland flying fox populations, urine samples were collected on a regular basis for PCR and virus isolation. CedPV is more closely related to HeV and NiV than henipavirus-like sequences detected in African bats [26, 32] as shown in a phylogenetic tree based on the only sequences available of a 550-nt L gene fragment (Fig. S5) .

abstract:
  The genus Henipavirus in the family Paramyxoviridae contains two viruses, Hendra virus (HeV) and Nipah virus (NiV) for which pteropid bats act as the main natural reservoir. Each virus also causes serious and commonly lethal infection of people as well as various species of domestic animals, however little is known about the associated mechanisms of pathogenesis. Here, we report the isolation and characterization of a new paramyxovirus from pteropid bats, Cedar virus (CedPV), which shares significant features with the known henipaviruses. The genome size (18,162 nt) and organization of CedPV is very similar to that of HeV and NiV; its nucleocapsid protein displays antigenic cross-reactivity with henipaviruses; and it uses the same receptor molecule (ephrin- B2) for entry during infection. Preliminary challenge studies with CedPV in ferrets and guinea pigs, both susceptible to infection and disease with known henipaviruses, confirmed virus replication and production of neutralizing antibodies although clinical disease was not observed. In this context, it is interesting to note that the major genetic difference between CedPV and HeV or NiV lies within the coding strategy of the P gene, which is known to play an important role in evading the host innate immune system. Unlike HeV, NiV, and almost all known paramyxoviruses, the CedPV P gene lacks both RNA editing and also the coding capacity for the highly conserved V protein. Preliminary study indicated that CedPV infection of human cells induces a more robust IFN-β response than HeV.


id: cord-000933-nn9gj0z1
author: Krzyzaniak, Magdalena Anna
date: 2013-04-11
txt: ./txt/cord-000933-nn9gj0z1.txt
cache: ./cache/cord-000933-nn9gj0z1.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3623752/
doi: 10.1371/journal.ppat.1003309
summary:
  To study the entry process in human tissue culture cells (HeLa, A549), we used fluorescence microscopy and developed quantitative, FACS-based assays to follow virus binding to cells, endocytosis, intracellular trafficking, membrane fusion, and infection. To determine the pathway of RSV entry into HeLa and A549 cells, we developed quantitative fluorescence-activated cell sorting (FACS) assays and complemented them with confocal microscopy to monitor cell binding of RSV, endocytosis, fusion, and infection. Since we did not detect free capsids that would stain only for N or P (data not shown), we used the presence of the capsid antigens to distinguish between intact RSVs and VLPs. When purified virus preparations were incubated with HeLa cells at 4uC, immunoblotting after SDS-PAGE showed that more than half of the input N and P associated with the cells indicating that RSV binding in the cold was efficient (Fig. 1C) .

title:
  Host Cell Entry of Respiratory Syncytial Virus Involves Macropinocytosis Followed by Proteolytic Activation of the F Protein

abstract:
  Respiratory Syncytial Virus (RSV) is a highly pathogenic member of the Paramyxoviridae that causes severe respiratory tract infections. Reports in the literature have indicated that to infect cells the incoming viruses either fuse their envelope directly with the plasma membrane or exploit clathrin-mediated endocytosis. To study the entry process in human tissue culture cells (HeLa, A549), we used fluorescence microscopy and developed quantitative, FACS-based assays to follow virus binding to cells, endocytosis, intracellular trafficking, membrane fusion, and infection. A variety of perturbants were employed to characterize the cellular processes involved. We found that immediately after binding to cells RSV activated a signaling cascade involving the EGF receptor, Cdc42, PAK1, and downstream effectors. This led to a series of dramatic actin rearrangements; the cells rounded up, plasma membrane blebs were formed, and there was a significant increase in fluid uptake. If these effects were inhibited using compounds targeting Na(+)/H(+) exchangers, myosin II, PAK1, and other factors, no infection was observed. The RSV was rapidly and efficiently internalized by an actin-dependent process that had all hallmarks of macropinocytosis. Rather than fusing with the plasma membrane, the viruses thus entered Rab5-positive, fluid-filled macropinosomes, and fused with the membranes of these on the average 50 min after internalization. Rab5 was required for infection. To find an explanation for the endocytosis requirement, which is unusual among paramyxoviruses, we analyzed the fusion protein, F, and could show that, although already cleaved by a furin family protease once, it underwent a second, critical proteolytic cleavage after internalization. This cleavage by a furin-like protease removed a small peptide from the F1 subunits, and made the virus infectious.


id: cord-001082-sufwsu77
author: Bär, Séverine
date: 2013-09-19
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3777860/
doi: 10.1371/journal.ppat.1003605
title:
  Vesicular Transport of Progeny Parvovirus Particles through ER and Golgi Regulates Maturation and Cytolysis

abstract:
  Progeny particles of non-enveloped lytic parvoviruses were previously shown to be actively transported to the cell periphery through vesicles in a gelsolin-dependent manner. This process involves rearrangement and destruction of actin filaments, while microtubules become protected throughout the infection. Here the focus is on the intracellular egress pathway, as well as its impact on the properties and release of progeny virions. By colocalization with cellular marker proteins and specific modulation of the pathways through over-expression of variant effector genes transduced by recombinant adeno-associated virus vectors, we show that progeny PV particles become engulfed into COPII-vesicles in the endoplasmic reticulum (ER) and are transported through the Golgi to the plasma membrane. Besides known factors like sar1, sec24, rab1, the ERM family proteins, radixin and moesin play (an) essential role(s) in the formation/loading and targeting of virus-containing COPII-vesicles. These proteins also contribute to the transport through ER and Golgi of the well described analogue of cellular proteins, the secreted Gaussia luciferase in absence of virus infection. It is therefore likely that radixin and moesin also serve for a more general function in cellular exocytosis. Finally, parvovirus egress via ER and Golgi appears to be necessary for virions to gain full infectivity through post-assembly modifications (e.g. phosphorylation). While not being absolutely required for cytolysis and progeny virus release, vesicular transport of parvoviruses through ER and Golgi significantly accelerates these processes pointing to a regulatory role of this transport pathway.


id: cord-001109-xs7df6a7
author: Tapia, Karla
date: 2013-10-31
txt: ./txt/cord-001109-xs7df6a7.txt
cache: ./cache/cord-001109-xs7df6a7.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3814336/
doi: 10.1371/journal.ppat.1003703
summary:
  We demonstrate that these defective viral genomes (DVGs) are generated naturally during respiratory infections in vivo even in mice lacking the type I IFN receptor, and their appearance coincides with the production of cytokines during infections with Sendai virus (SeV) or influenza virus. To further investigate the cellular mechanisms responsible for the efficient activation of the antiviral response by SeV DVGs, we evaluated the phosphorylation of transcription factors that are critical for the expression of type I IFNs in cells infected with equivalent amounts of infectious particles of a SeV strain Cantell stock containing high levels of copy-back DVGs (SeV Cantell HD) or with SeV Cantell depleted of DVGs (SeV Cantell LD). Based on the potent ability of SeV stocks containing a high content of copy-back DVGs to induce the host response to infection in vitro [23, 24, 25, 28] (Fig. 1 ) and on our prior reports of strong host responses to DVGs regardless of the presence of functional virus-encoded antagonists [23, 24] , we hypothesized that DVGs that arise in situ during viral infections provide essential stimuli to initiate an antiviral immune response.

title:
  Defective Viral Genomes Arising In Vivo Provide Critical Danger Signals for the Triggering of Lung Antiviral Immunity

abstract:
  The innate immune response to viruses is initiated when specialized cellular sensors recognize viral danger signals. Here we show that truncated forms of viral genomes that accumulate in infected cells potently trigger the sustained activation of the transcription factors IRF3 and NF-κB and the production type I IFNs through a mechanism independent of IFN signaling. We demonstrate that these defective viral genomes (DVGs) are generated naturally during respiratory infections in vivo even in mice lacking the type I IFN receptor, and their appearance coincides with the production of cytokines during infections with Sendai virus (SeV) or influenza virus. Remarkably, the hallmark antiviral cytokine IFNβ is only expressed in lung epithelial cells containing DVGs, while cells within the lung that contain standard viral genomes alone do not express this cytokine. Together, our data indicate that DVGs generated during viral replication are a primary source of danger signals for the initiation of the host immune response to infection.


id: cord-001257-t21l6i3f
author: Kovalev, Nikolay
date: 2014-04-17
txt: ./txt/cord-001257-t21l6i3f.txt
cache: ./cache/cord-001257-t21l6i3f.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3990711/
doi: 10.1371/journal.ppat.1004051
summary:
  Novel pro-viral function of AtRH2, AtRH5 and the yeast Dbp3p and Fal1p is to bind to the viral replication enhancer present in the tombusvirus minus-strand RNA To test if AtRH2, AtRH5 and the yeast Dbp3p and Fal1p play a comparable role with the previously analyzed yeast Ded1p (human DDX3-like) and Dbp2p (human p68-like) and the ortologous AtRH20 DEAD-box helicases [30, 49] , first we performed in vitro RNA binding experiments with affinity-purified recombinant helicase proteins. The replication process leads to the production of abundant (+)RNA progeny, while the (2)RNA templates are likely sequestered in dsRNA forms within the VRCs. The presented in vitro data based on the solubilized/purified tombusvirus replicase and the CFE assay containing the membrane-bound VRC indicate that the eIF4AIIIlike RNA helicases can mainly stimulate TBSV (+)-strand synthesis, while their effects on (2)RNA synthesis have not been observed (not shown).

title:
  The Expanding Functions of Cellular Helicases: The Tombusvirus RNA Replication Enhancer Co-opts the Plant eIF4AIII-Like AtRH2 and the DDX5-Like AtRH5 DEAD-Box RNA Helicases to Promote Viral Asymmetric RNA Replication

abstract:
  Replication of plus-strand RNA viruses depends on recruited host factors that aid several critical steps during replication. Several of the co-opted host factors bind to the viral RNA, which plays multiple roles, including mRNA function, as an assembly platform for the viral replicase (VRC), template for RNA synthesis, and encapsidation during infection. It is likely that remodeling of the viral RNAs and RNA-protein complexes during the switch from one step to another requires RNA helicases. In this paper, we have discovered a second group of cellular RNA helicases, including the eIF4AIII-like yeast Fal1p and the DDX5-like Dbp3p and the orthologous plant AtRH2 and AtRH5 DEAD box helicases, which are co-opted by tombusviruses. Unlike the previously characterized DDX3-like AtRH20/Ded1p helicases that bind to the 3′ terminal promoter region in the viral minus-strand (−)RNA, the other class of eIF4AIII-like RNA helicases bind to a different cis-acting element, namely the 5′ proximal RIII(−) replication enhancer (REN) element in the TBSV (−)RNA. We show that the binding of AtRH2 and AtRH5 helicases to the TBSV (−)RNA could unwind the dsRNA structure within the RIII(−) REN. This unique characteristic allows the eIF4AIII-like helicases to perform novel pro-viral functions involving the RIII(−) REN in stimulation of plus-strand (+)RNA synthesis. We also show that AtRH2 and AtRH5 helicases are components of the tombusvirus VRCs based on co-purification experiments. We propose that eIF4AIII-like helicases destabilize dsRNA replication intermediate within the RIII(−) REN that promotes bringing the 5′ and 3′ terminal (−)RNA sequences in close vicinity via long-range RNA-RNA base pairing. This newly formed RNA structure promoted by eIF4AIII helicase together with AtRH20 helicase might facilitate the recycling of the viral replicases for multiple rounds of (+)-strand synthesis, thus resulting in asymmetrical viral replication.


id: cord-001385-rb5vwolt
author: Reuven, Eliran Moshe
date: 2014-08-14
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4133399/
doi: 10.1371/journal.ppat.1004248
title:
  The HIV-1 Envelope Transmembrane Domain Binds TLR2 through a Distinct Dimerization Motif and Inhibits TLR2-Mediated Responses

abstract:
  HIV-1 uses a number of means to manipulate the immune system, to avoid recognition and to highjack signaling pathways. HIV-1 infected cells show limited Toll-Like Receptor (TLR) responsiveness via as yet unknown mechanisms. Using biochemical and biophysical approaches, we demonstrate that the trans-membrane domain (TMD) of the HIV-1 envelope (ENV) directly interacts with TLR2 TMD within the membrane milieu. This interaction attenuates TNFα, IL-6 and MCP-1 secretion in macrophages, induced by natural ligands of TLR2 both in in vitro and in vivo models. This was associated with decreased levels of ERK phosphorylation. Furthermore, mutagenesis demonstrated the importance of a conserved GxxxG motif in driving this interaction within the membrane milieu. The administration of the ENV TMD in vivo to lipotechoic acid (LTA)/Galactosamine-mediated septic mice resulted in a significant decrease in mortality and in tissue damage, due to the weakening of systemic macrophage activation. Our findings suggest that the TMD of ENV is involved in modulation of the innate immune response during HIV infection. Furthermore, due to the high functional homology of viral ENV proteins this function may be a general character of viral-induced immune modulation.


id: cord-001541-5d64esp4
title: Evolution of Genome Size and Complexity in the Rhabdoviridae
author: Walker, Peter J.
date: 2015-02-13
txt: ./txt/cord-001541-5d64esp4.txt
cache: ./cache/cord-001541-5d64esp4.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4334499/
doi: 10.1371/journal.ppat.1004664
summary:
  We demonstrate that remarkable changes in genome size and complexity have occurred in rhabdoviruses in a clade-specific manner, primarily by extension and insertion of additional transcriptional units in the structural protein gene junctions, followed by occasional losses. All rhabdovirus genomes contained the five canonical structural protein genes (N, P, M, G and L); however, there was remarkable diversity in the number and location of other long ORFs. Across the data set, we identified 179 additional ORFs 180 nt in length of which 142 shared no detectable protein sequence similarity with any other protein in our data set or with those in public databases (S2 Table) . Interestingly, although substantial variation in the length of gene junctions was observed in several genera (including ephemeroviruses and lyssaviruses), most variation in genome size occurred as the result of the presence of new, non-canonical ORFs in the regions between the structural protein genes (Table 1) .

abstract:
  RNA viruses exhibit substantial structural, ecological and genomic diversity. However, genome size in RNA viruses is likely limited by a high mutation rate, resulting in the evolution of various mechanisms to increase complexity while minimising genome expansion. Here we conduct a large-scale analysis of the genome sequences of 99 animal rhabdoviruses, including 45 genomes which we determined de novo, to identify patterns of genome expansion and the evolution of genome complexity. All but seven of the rhabdoviruses clustered into 17 well-supported monophyletic groups, of which eight corresponded to established genera, seven were assigned as new genera, and two were taxonomically ambiguous. We show that the acquisition and loss of new genes appears to have been a central theme of rhabdovirus evolution, and has been associated with the appearance of alternative, overlapping and consecutive ORFs within the major structural protein genes, and the insertion and loss of additional ORFs in each gene junction in a clade-specific manner. Changes in the lengths of gene junctions accounted for as much as 48.5% of the variation in genome size from the smallest to the largest genome, and the frequency with which new ORFs were observed increased in the 3’ to 5’ direction along the genome. We also identify several new families of accessory genes encoded in these regions, and show that non-canonical expression strategies involving TURBS-like termination-reinitiation, ribosomal frame-shifts and leaky ribosomal scanning appear to be common. We conclude that rhabdoviruses have an unusual capacity for genomic plasticity that may be linked to their discontinuous transcription strategy from the negative-sense single-stranded RNA genome, and propose a model that accounts for the regular occurrence of genome expansion and contraction throughout the evolution of the Rhabdoviridae.


id: cord-001636-khrx6rnh
author: Bordería, Antonio V.
date: 2015-05-05
txt: ./txt/cord-001636-khrx6rnh.txt
cache: ./cache/cord-001636-khrx6rnh.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4420505/
doi: 10.1371/journal.ppat.1004838
summary:
  The total number of minority variants within the P1 structural coding region also significantly increased over the passage series (Fig 1G) , yet there were no significant differences between the numbers observed in Hela and A549 cell passage. The mutations that arise from adaptive passage map to receptor binding sites in a cell-type specific manner Although the HeLa and A549 cell passage series presented similar mean genetic diversity at passage 40 (Fig 1E-1G , P = 0.314), we mined the deep sequence data for all minority variants above 1% frequency that might explain adaptation in each condition (Table 1 ). Given the different frequencies of minority variants observed at passage 40, we examined the patterns of emergence and sequential adaptation to novel environments by deep sequencing every second passage in the A549 cell series (Fig 3 and S1 Dataset).

title:
  Group Selection and Contribution of Minority Variants during Virus Adaptation Determines Virus Fitness and Phenotype

abstract:
  Understanding how a pathogen colonizes and adapts to a new host environment is a primary aim in studying emerging infectious diseases. Adaptive mutations arise among the thousands of variants generated during RNA virus infection, and identifying these variants will shed light onto how changes in tropism and species jumps can occur. Here, we adapted Coxsackie virus B3 to a highly permissive and less permissive environment. Using deep sequencing and bioinformatics, we identified a multi-step adaptive process to adaptation involving residues in the receptor footprints that correlated with receptor availability and with increase in virus fitness in an environment-specific manner. We show that adaptation occurs by selection of a dominant mutation followed by group selection of minority variants that together, confer the fitness increase observed in the population, rather than selection of a single dominant genotype.


id: cord-001655-uqw74ra0
author: Stenglein, Mark D.
date: 2015-05-20
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4438980/
doi: 10.1371/journal.ppat.1004900
title:
  Widespread Recombination, Reassortment, and Transmission of Unbalanced Compound Viral Genotypes in Natural Arenavirus Infections

abstract:
  Arenaviruses are one of the largest families of human hemorrhagic fever viruses and are known to infect both mammals and snakes. Arenaviruses package a large (L) and small (S) genome segment in their virions. For segmented RNA viruses like these, novel genotypes can be generated through mutation, recombination, and reassortment. Although it is believed that an ancient recombination event led to the emergence of a new lineage of mammalian arenaviruses, neither recombination nor reassortment has been definitively documented in natural arenavirus infections. Here, we used metagenomic sequencing to survey the viral diversity present in captive arenavirus-infected snakes. From 48 infected animals, we determined the complete or near complete sequence of 210 genome segments that grouped into 23 L and 11 S genotypes. The majority of snakes were multiply infected, with up to 4 distinct S and 11 distinct L segment genotypes in individual animals. This S/L imbalance was typical: in all cases intrahost L segment genotypes outnumbered S genotypes, and a particular S segment genotype dominated in individual animals and at a population level. We corroborated sequencing results by qRT-PCR and virus isolation, and isolates replicated as ensembles in culture. Numerous instances of recombination and reassortment were detected, including recombinant segments with unusual organizations featuring 2 intergenic regions and superfluous content, which were capable of stable replication and transmission despite their atypical structures. Overall, this represents intrahost diversity of an extent and form that goes well beyond what has been observed for arenaviruses or for viruses in general. This diversity can be plausibly attributed to the captive intermingling of sub-clinically infected wild-caught snakes. Thus, beyond providing a unique opportunity to study arenavirus evolution and adaptation, these findings allow the investigation of unintended anthropogenic impacts on viral ecology, diversity, and disease potential.


id: cord-001676-68y733y3
title: An Ultrasensitive Mechanism Regulates Influenza Virus-Induced Inflammation
author: Shoemaker, Jason E.
date: 2015-06-05
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4457877/
doi: 10.1371/journal.ppat.1004856
abstract:
  Influenza viruses present major challenges to public health, evident by the 2009 influenza pandemic. Highly pathogenic influenza virus infections generally coincide with early, high levels of inflammatory cytokines that some studies have suggested may be regulated in a strain-dependent manner. However, a comprehensive characterization of the complex dynamics of the inflammatory response induced by virulent influenza strains is lacking. Here, we applied gene co-expression and nonlinear regression analysis to time-course, microarray data developed from influenza-infected mouse lung to create mathematical models of the host inflammatory response. We found that the dynamics of inflammation-associated gene expression are regulated by an ultrasensitive-like mechanism in which low levels of virus induce minimal gene expression but expression is strongly induced once a threshold virus titer is exceeded. Cytokine assays confirmed that the production of several key inflammatory cytokines, such as interleukin 6 and monocyte chemotactic protein 1, exhibit ultrasensitive behavior. A systematic exploration of the pathways regulating the inflammatory-associated gene response suggests that the molecular origins of this ultrasensitive response mechanism lie within the branch of the Toll-like receptor pathway that regulates STAT1 phosphorylation. This study provides the first evidence of an ultrasensitive mechanism regulating influenza virus-induced inflammation in whole lungs and provides insight into how different virus strains can induce distinct temporal inflammation response profiles. The approach developed here should facilitate the construction of gene regulatory models of other infectious diseases.


id: cord-001700-c6elsnag
title: Protective mAbs and Cross-Reactive mAbs Raised by Immunization with Engineered Marburg Virus GPs
author: Fusco, Marnie L.
date: 2015-06-26
txt: ./txt/cord-001700-c6elsnag.txt
cache: ./cache/cord-001700-c6elsnag.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4482612/
doi: 10.1371/journal.ppat.1005016
summary:
  Here we present ten mAbs elicited by immunization of mice using recombinant mucin-deleted GPs from different Marburg virus (MARV) strains. Surprisingly, two of the mAbs raised against MARV GP also cross-react with the mucin-deleted GP cores of all tested ebolaviruses (Ebola, Sudan, Bundibugyo, Reston), but these epitopes are masked differently by the mucin-like domains themselves. To generate MARV GP-specific mAbs, BALB/c mice were immunized with GPΔmuc antigens from either MARV strain Ci67, Musoke, Angola, or Ravn ( Fig 1A) . To characterize the binding of mAbs, we performed enzyme-linked immunosorbent assays (ELISAs) with recombinant GPs from four MARV strains, and determined EC50 values for binding with different forms of MARV Ravn GP: GP, GPΔmuc, GPcl (Fig 2A) . Two of the highly cross-reactive MARV antibodies, mAbs 40G1 and 2D8, also exhibit binding to Ebola, Sudan, Bundibugyo and Reston virus mucin-deleted GPs by ELISA (Fig 5A) .

abstract:
  The filoviruses, which include the marburg- and ebolaviruses, have caused multiple outbreaks among humans this decade. Antibodies against the filovirus surface glycoprotein (GP) have been shown to provide life-saving therapy in nonhuman primates, but such antibodies are generally virus-specific. Many monoclonal antibodies (mAbs) have been described against Ebola virus. In contrast, relatively few have been described against Marburg virus. Here we present ten mAbs elicited by immunization of mice using recombinant mucin-deleted GPs from different Marburg virus (MARV) strains. Surprisingly, two of the mAbs raised against MARV GP also cross-react with the mucin-deleted GP cores of all tested ebolaviruses (Ebola, Sudan, Bundibugyo, Reston), but these epitopes are masked differently by the mucin-like domains themselves. The most efficacious mAbs in this panel were found to recognize a novel “wing” feature on the GP2 subunit that is unique to Marburg and does not exist in Ebola. Two of these anti-wing antibodies confer 90 and 100% protection, respectively, one hour post-exposure in mice challenged with MARV.


id: cord-001769-2sdg5ll7
author: Guo, Sheng
date: 2015-09-14
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4569300/
doi: 10.1371/journal.ppat.1005155
title:
  The NLRP3 Inflammasome and IL-1β Accelerate Immunologically Mediated Pathology in Experimental Viral Fulminant Hepatitis

abstract:
  Viral fulminant hepatitis (FH) is a severe disease with high mortality resulting from excessive inflammation in the infected liver. Clinical interventions have been inefficient due to the lack of knowledge for inflammatory pathogenesis in the virus-infected liver. We show that wild-type mice infected with murine hepatitis virus strain-3 (MHV-3), a model for viral FH, manifest with severe disease and high mortality in association with a significant elevation in IL-1β expression in the serum and liver. Whereas, the viral infection in IL-1β receptor-I deficient (IL-1R1 (-/-)) or IL-1R antagonist (IL-1Ra) treated mice, show reductions in virus replication, disease progress and mortality. IL-1R1 deficiency appears to debilitate the virus-induced fibrinogen-like protein-2 (FGL2) production in macrophages and CD45(+)Gr-1(high) neutrophil infiltration in the liver. The quick release of reactive oxygen species (ROS) by the infected macrophages suggests a plausible viral initiation of NLRP3 inflammasome activation. Further experiments show that mice deficient of p47 (phox), a nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunit that controls acute ROS production, present with reductions in NLRP3 inflammasome activation and subsequent IL-1β secretion during viral infection, which appears to be responsible for acquiring resilience to viral FH. Moreover, viral infected animals in deficiencies of NLRP3 and Caspase-1, two essential components of the inflammasome complex, also have reduced IL-1β induction along with ameliorated hepatitis. Our results demonstrate that the ROS/NLRP3/IL-1β axis institutes an essential signaling pathway, which is over activated and directly causes the severe liver disease during viral infection, which sheds light on development of efficient treatments for human viral FH and other severe inflammatory diseases.


id: cord-001859-d62iuk72
author: Baquero-Pérez, Belinda
date: 2015-11-20
txt: ./txt/cord-001859-d62iuk72.txt
cache: ./cache/cord-001859-d62iuk72.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4654589/
doi: 10.1371/journal.ppat.1005274
summary:
  Similar Hsc70 localization was seen during early lytic replication (12 h reactivation), when RTA protein was diffuse in the nucleus, Successful enrichment of the nuclear envelope region and associated KSHV RTCs in HEK-293T rKSHV.219 cells. These results clearly demonstrate that KSHV specifically redistributes the molecular chaperones, Hsc70 and iHsp70, from the cytoplasm to the nucleus, in contrast to Grp78, which coincides with the initial formation of KSHV RTCs. Treatment with the small molecule inhibitor VER-155008 abrogated viral protein synthesis at non-cytotoxic concentrations Members of the HSP70 chaperone family possess an N-terminal nucleotide binding domain with ATPase activity which is essential for their function. Therefore to ascertain whether Hsp70 isoforms could stabilise the essential KSHV lytic proteins RTA and ORF57, TREx BCBL1-RTA cells were reactivated for 24 h to allow sufficient viral protein expression followed by addition of DMSO control or VER-155008 in conjunction with cycloheximide (CHX) at 50 μg/ml to block de novo protein synthesis.

title:
  Hsp70 Isoforms Are Essential for the Formation of Kaposi’s Sarcoma-Associated Herpesvirus Replication and Transcription Compartments

abstract:
  Kaposi’s sarcoma-associated herpesvirus (KSHV) is an oncogenic herpesvirus associated with various AIDS-related malignancies. Like other herpesviruses, multiple processes required for KSHV lytic replication, including viral transcription, viral DNA synthesis and capsid assembly occur in virus-induced intranuclear structures, termed replication and transcription compartments (RTCs). Here we utilised a novel methodology, combining subcellular fractionation and quantitative proteomics, to identify cellular proteins which are recruited to KSHV-induced RTCs and thus play a key role in KSHV lytic replication. We show that several isoforms of the HSP70 chaperone family, Hsc70 and iHsp70, are redistributed from the cytoplasm into the nucleus coinciding with the initial formation of KSHV-induced RTCs. We demonstrate that nuclear chaperone foci are dynamic, initially forming adjacent to newly formed KSHV RTCs, however during later time points the chaperones move within KSHV RTCs and completely co-localise with actively replicating viral DNA. The functional significance of Hsp70 isoforms recruitment into KSHV RTCs was also examined using the specific Hsp70 isoform small molecule inhibitor, VER-155008. Intriguingly, results highlight an essential role of Hsp70 isoforms in the KSHV replication cycle independent of protein stability and maturation. Notably, inhibition of Hsp70 isoforms precluded KSHV RTC formation and RNA polymerase II (RNAPII) relocalisation to the viral genome leading to the abolishment of global KSHV transcription and subsequent viral protein synthesis and DNA replication. These new findings have revealed novel mechanisms that regulate KSHV lytic replication and highlight the potential of HSP70 inhibitors as novel antiviral agents.


id: cord-001992-m4k20i7g
author: Ziegler, Christopher M.
date: 2016-03-24
txt: ./txt/cord-001992-m4k20i7g.txt
cache: ./cache/cord-001992-m4k20i7g.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4806877/
doi: 10.1371/journal.ppat.1005501
summary:
  We show that neither the PPXY late domain encoded within the lymphocytic choriomeningitis virus (LCMV) matrix protein nor a functional endosomal sorting complex transport (ESCRT) pathway is absolutely required for the generation of standard infectious virus particles. Given the important role of the LCMV matrix protein and its late domain motif in regulating viral budding [10, 11] , we next investigated the specific effect these point mutations had on Z''s budding efficiency in a VLP release assay. To investigate the protein and genome composition of virions containing mutated late domains, an equivalent quantity of cell-free infectious virus particles from each rLCMV strain was concentrated for screening. Limiting dilutions of this UV-treated sample were applied to Vero E6 cells, followed by the addition of a fixed quantity of LCMV PFUs. As shown in Fig 5A, this allows for the determination of LCMV DI particle activity and is expressed as plaque interfering units 50 (PIU 50 ) per mL of a given sample.

title:
  The Lymphocytic Choriomeningitis Virus Matrix Protein PPXY Late Domain Drives the Production of Defective Interfering Particles

abstract:
  Arenaviruses cause severe diseases in humans but establish asymptomatic, lifelong infections in rodent reservoirs. Persistently-infected rodents harbor high levels of defective interfering (DI) particles, which are thought to be important for establishing persistence and mitigating virus-induced cytopathic effect. Little is known about what drives the production of DI particles. We show that neither the PPXY late domain encoded within the lymphocytic choriomeningitis virus (LCMV) matrix protein nor a functional endosomal sorting complex transport (ESCRT) pathway is absolutely required for the generation of standard infectious virus particles. In contrast, DI particle release critically requires the PPXY late domain and is ESCRT-dependent. Additionally, the terminal tyrosine in the PPXY motif is reversibly phosphorylated and our findings indicate that this posttranslational modification may regulate DI particle formation. Thus we have uncovered a new role for the PPXY late domain and a possible mechanism for its regulation.


id: cord-002125-anp238gu
author: Effantin, Grégory
date: 2016-07-11
txt: ./txt/cord-002125-anp238gu.txt
cache: ./cache/cord-002125-anp238gu.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4939959/
doi: 10.1371/journal.ppat.1005721
summary:
  Mature PFV particles have a distinct morphology with a capsid of constant dimension as well as a less ordered shell of density between the capsid and the membrane likely formed by the Gag N-terminal domain and the cytoplasmic part of the Env leader peptide gp18(LP). In order to obtain structural insight into PFV organization at medium resolution we used cryo-electron tomography (cryo-ET) and microscopy (cryo-EM) to analyze isolated wild-type particles and variants with mutations in PFV Gag and Env proteins. To gain more insights into the structure of the PFV glycoprotein, we use 3D reconstructions of iNAB and iFuse Env hexagonal assemblies of six trimers determined by subtomogram averaging as initial references for automatic particle selection of micrographs acquired by cryo-EM (S4 Fig) (see Material and Methods).

title:
  Cryo-electron Microscopy Structure of the Native Prototype Foamy Virus Glycoprotein and Virus Architecture

abstract:
  Foamy viruses (FV) belong to the genus Spumavirus, which forms a distinct lineage in the Retroviridae family. Although the infection in natural hosts and zoonotic transmission to humans is asymptomatic, FVs can replicate well in human cells making it an attractive gene therapy vector candidate. Here we present cryo-electron microscopy and (cryo-)electron tomography ultrastructural data on purified prototype FV (PFV) and PFV infected cells. Mature PFV particles have a distinct morphology with a capsid of constant dimension as well as a less ordered shell of density between the capsid and the membrane likely formed by the Gag N-terminal domain and the cytoplasmic part of the Env leader peptide gp18(LP). The viral membrane contains trimeric Env glycoproteins partly arranged in interlocked hexagonal assemblies. In situ 3D reconstruction by subtomogram averaging of wild type Env and of a Env gp48(TM)- gp80(SU) cleavage site mutant showed a similar spike architecture as well as stabilization of the hexagonal lattice by clear connections between lower densities of neighboring trimers. Cryo-EM was employed to obtain a 9 Å resolution map of the glycoprotein in its pre-fusion state, which revealed extensive trimer interactions by the receptor binding subunit gp80(SU) at the top of the spike and three central helices derived from the fusion protein subunit gp48(TM). The lower part of Env, presumably composed of interlaced parts of gp48(TM), gp80(SU) and gp18(LP) anchors the spike at the membrane. We propose that the gp48(TM) density continues into three central transmembrane helices, which interact with three outer transmembrane helices derived from gp18(LP). Our ultrastructural data and 9 Å resolution glycoprotein structure provide important new insights into the molecular architecture of PFV and its distinct evolutionary relationship with other members of the Retroviridae.


id: cord-002142-tdgu9sr9
author: Reniere, Michelle L.
date: 2016-07-14
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4945081/
doi: 10.1371/journal.ppat.1005741
title:
  An In Vivo Selection Identifies Listeria monocytogenes Genes Required to Sense the Intracellular Environment and Activate Virulence Factor Expression

abstract:
  Listeria monocytogenes is an environmental saprophyte and facultative intracellular bacterial pathogen with a well-defined life-cycle that involves escape from a phagosome, rapid cytosolic growth, and ActA-dependent cell-to-cell spread, all of which are dependent on the master transcriptional regulator PrfA. The environmental cues that lead to temporal and spatial control of L. monocytogenes virulence gene expression are poorly understood. In this study, we took advantage of the robust up-regulation of ActA that occurs intracellularly and expressed Cre recombinase from the actA promoter and 5’ untranslated region in a strain in which loxP sites flanked essential genes, so that activation of actA led to bacterial death. Upon screening for transposon mutants that survived intracellularly, six genes were identified as necessary for ActA expression. Strikingly, most of the genes, including gshF, spxA1, yjbH, and ohrA, are predicted to play important roles in bacterial redox regulation. The mutants identified in the genetic selection fell into three broad categories: (1) those that failed to reach the cytosolic compartment; (2) mutants that entered the cytosol, but failed to activate the master virulence regulator PrfA; and (3) mutants that entered the cytosol and activated transcription of actA, but failed to synthesize it. The identification of mutants defective in vacuolar escape suggests that up-regulation of ActA occurs in the host cytosol and not the vacuole. Moreover, these results provide evidence for two non-redundant cytosolic cues; the first results in allosteric activation of PrfA via increased glutathione levels and transcriptional activation of actA while the second results in translational activation of actA and requires yjbH. Although the precise host cues have not yet been identified, we suggest that intracellular redox stress occurs as a consequence of both host and pathogen remodeling their metabolism upon infection.


id: cord-002423-1u44tdrj
author: Geoghegan, Jemma L.
date: 2017-02-08
txt: ./txt/cord-002423-1u44tdrj.txt
cache: ./cache/cord-002423-1u44tdrj.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5319820/
doi: 10.1371/journal.ppat.1006215
summary:
  While this method does not explicitly model host-switching events, it does provide a simple means to compare multiple topologies of virus-host pairs, and accounts for differences in sample size and the fact that several viruses from a specific family can infect a single host species. Across the data set as a whole we found that all virus families displayed relatively large tree topological distances with nPH85 values of !0.6, suggesting that cross-species transmission is widespread, at least at the family-level (Fig 2; S3 Table) . As with the analysis of topological distances, this revealed that cross-species transmission was the most common evolutionary event in all virus families studied here, with co-divergence consistently less frequent (with the possible exception of the Hepadnaviridae-see below), and lineage duplication and extinction playing a much more minor role. To investigate the comparative prevalence of cross-species transmission among viruses we measured the congruence between virus and host phylogenetic trees using a normalized tree topological distance-based approach (nPH85, [14] ).

title:
  Comparative analysis estimates the relative frequencies of co-divergence and cross-species transmission within viral families

abstract:
  The cross-species transmission of viruses from one host species to another is responsible for the majority of emerging infections. However, it is unclear whether some virus families have a greater propensity to jump host species than others. If related viruses have an evolutionary history of co-divergence with their hosts there should be evidence of topological similarities between the virus and host phylogenetic trees, whereas host jumping generates incongruent tree topologies. By analyzing co-phylogenetic processes in 19 virus families and their eukaryotic hosts we provide a quantitative and comparative estimate of the relative frequency of virus-host co-divergence versus cross-species transmission among virus families. Notably, our analysis reveals that cross-species transmission is a near universal feature of the viruses analyzed here, with virus-host co-divergence occurring less frequently and always on a subset of viruses. Despite the overall high topological incongruence among virus and host phylogenies, the Hepadnaviridae, Polyomaviridae, Poxviridae, Papillomaviridae and Adenoviridae, all of which possess double-stranded DNA genomes, exhibited more frequent co-divergence than the other virus families studied here. At the other extreme, the virus and host trees for all the RNA viruses studied here, particularly the Rhabdoviridae and the Picornaviridae, displayed high levels of topological incongruence, indicative of frequent host switching. Overall, we show that cross-species transmission plays a major role in virus evolution, with all the virus families studied here having the potential to jump host species, and that increased sampling will likely reveal more instances of host jumping.


id: cord-002530-ao7kenbx
author: Lin, Yao-Tang
date: 2017-05-11
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5426786/
doi: 10.1371/journal.ppat.1006329
title:
  The host ubiquitin-dependent segregase VCP/p97 is required for the onset of human cytomegalovirus replication

abstract:
  The human cytomegalovirus major immediate early proteins IE1 and IE2 are critical drivers of virus replication and are considered pivotal in determining the balance between productive and latent infection. IE1 and IE2 are derived from the same primary transcript by alternative splicing and regulation of their expression likely involves a complex interplay between cellular and viral factors. Here we show that knockdown of the host ubiquitin-dependent segregase VCP/p97, results in loss of IE2 expression, subsequent suppression of early and late gene expression and, ultimately, failure in virus replication. RNAseq analysis showed increased levels of IE1 splicing, with a corresponding decrease in IE2 splicing following VCP knockdown. Global analysis of viral transcription showed the expression of a subset of viral genes is not reduced despite the loss of IE2 expression, including UL112/113. Furthermore, Immunofluorescence studies demonstrated that VCP strongly colocalised with the viral replication compartments in the nucleus. Finally, we show that NMS-873, a small molecule inhibitor of VCP, is a potent HCMV antiviral with potential as a novel host targeting therapeutic for HCMV infection.


id: cord-002542-f7l4ty2j
author: Jaworski, Elizabeth
date: 2017-05-05
txt: ./txt/cord-002542-f7l4ty2j.txt
cache: ./cache/cord-002542-f7l4ty2j.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5435362/
doi: 10.1371/journal.ppat.1006365
summary:
  Using a combination of longand short-read Next-Generation Sequencing, we have characterized the formation of DI-RNAs during serial passaging of Flock House virus (FHV) in cell-culture over a period of 30 days in order to elucidate the pathways and potential mechanisms of DI-RNA emergence and evolution. By combining these data, we aimed to determine the correlation of recombination events within single RNA virus genomes, characterize the distribution of defective RNA genomes, and determine the exact make-up of DI-RNAs during serial passaging of FHV in cell culture. This observation indicates the protection of cells from infection and/or CPE when supplied with a large dose of viral particles that contain a large proportion of DI-RNAs. In this manuscript, we sought to provide a thorough and comprehensive analysis of the frequency and identity of recombination events present during the serial passaging of Flock House virus in cell culture in order to elucidate the pathways and mechanism of DI-RNA emergence and evolution.

title:
  Parallel ClickSeq and Nanopore sequencing elucidates the rapid evolution of defective-interfering RNAs in Flock House virus

abstract:
  Defective-Interfering RNAs (DI-RNAs) have long been known to play an important role in virus replication and transmission. DI-RNAs emerge during virus passaging in both cell-culture and their hosts as a result of non-homologous RNA recombination. However, the principles of DI-RNA emergence and their subsequent evolution have remained elusive. Using a combination of long- and short-read Next-Generation Sequencing, we have characterized the formation of DI-RNAs during serial passaging of Flock House virus (FHV) in cell-culture over a period of 30 days in order to elucidate the pathways and potential mechanisms of DI-RNA emergence and evolution. For short-read RNAseq, we employed ‘ClickSeq’ due to its ability to sensitively and confidently detect RNA recombination events with nucleotide resolution. In parallel, we used the Oxford Nanopore Technologies’s (ONT) MinION to resolve full-length defective and wild-type viral genomes. Together, these accurately resolve both rare and common RNA recombination events, determine the correlation between recombination events, and quantifies the relative abundance of different DI-RNAs throughout passaging. We observe the formation of a diverse pool of defective RNAs at each stage of viral passaging. However, many of these ‘intermediate’ species, while present in early stages of passaging, do not accumulate. After approximately 9 days of passaging we observe the rapid accumulation of DI-RNAs with a correlated reduction in specific infectivity and with the Nanopore data find that DI-RNAs are characterized by multiple RNA recombination events. This suggests that intermediate DI-RNA species are not competitive and that multiple recombination events interact epistatically to confer ‘mature’ DI-RNAs with their selective advantage allowing for their rapid accumulation. Alternatively, it is possible that mature DI-RNA species are generated in a single event involving multiple RNA rearrangements. These insights have important consequences for our understanding of the mechanisms, determinants and limitations in the emergence and evolution of DI-RNAs.


id: cord-002608-zn7tm1ww
author: Sokoloski, Kevin J.
date: 2017-06-29
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5507600/
doi: 10.1371/journal.ppat.1006473
title:
  Identification of Interactions between Sindbis Virus Capsid Protein and Cytoplasmic vRNA as Novel Virulence Determinants

abstract:
  Alphaviruses are arthropod-borne viruses that represent a significant threat to public health at a global level. While the formation of alphaviral nucleocapsid cores, consisting of cargo nucleic acid and the viral capsid protein, is an essential molecular process of infection, the precise interactions between the two partners are ill-defined. A CLIP-seq approach was used to screen for candidate sites of interaction between the viral Capsid protein and genomic RNA of Sindbis virus (SINV), a model alphavirus. The data presented in this report indicates that the SINV capsid protein binds to specific viral RNA sequences in the cytoplasm of infected cells, but its interaction with genomic RNA in mature extracellular viral particles is largely non-specific in terms of nucleotide sequence. Mutational analyses of the cytoplasmic viral RNA-capsid interaction sites revealed a functional role for capsid binding early in infection. Interaction site mutants exhibited decreased viral growth kinetics; however, this defect was not a function of decreased particle production. Rather mutation of the cytoplasmic capsid-RNA interaction sites negatively affected the functional capacity of the incoming viral genomic RNAs leading to decreased infectivity. Furthermore, cytoplasmic capsid interaction site mutants are attenuated in a murine model of neurotropic alphavirus infection. Collectively, the findings of this study indicate that the identified cytoplasmic interactions of the viral capsid protein and genomic RNA, while not essential for particle formation, are necessary for genomic RNA function early during infection. This previously unappreciated role of capsid protein during the alphaviral replication cycle also constitutes a novel virulence determinant.


id: cord-002706-m3y35ozx
author: Guo, Fang
date: 2017-09-25
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5629035/
doi: 10.1371/journal.ppat.1006658
title:
  HBV core protein allosteric modulators differentially alter cccDNA biosynthesis from de novo infection and intracellular amplification pathways

abstract:
  Hepatitis B virus (HBV) core protein assembles viral pre-genomic (pg) RNA and DNA polymerase into nucleocapsids for reverse transcriptional DNA replication to take place. Several chemotypes of small molecules, including heteroaryldihydropyrimidines (HAPs) and sulfamoylbenzamides (SBAs), have been discovered to allosterically modulate core protein structure and consequentially alter the kinetics and pathway of core protein assembly, resulting in formation of irregularly-shaped core protein aggregates or “empty” capsids devoid of pre-genomic RNA and viral DNA polymerase. Interestingly, in addition to inhibiting nucleocapsid assembly and subsequent viral genome replication, we have now demonstrated that HAPs and SBAs differentially modulate the biosynthesis of covalently closed circular (ccc) DNA from de novo infection and intracellular amplification pathways by inducing disassembly of nucleocapsids derived from virions as well as double-stranded DNA-containing progeny nucleocapsids in the cytoplasm. Specifically, the mistimed cuing of nucleocapsid uncoating prevents cccDNA formation during de novo infection of hepatocytes, while transiently accelerating cccDNA synthesis from cytoplasmic progeny nucleocapsids. Our studies indicate that elongation of positive-stranded DNA induces structural changes of nucleocapsids, which confers ability of mature nucleocapsids to bind CpAMs and triggers its disassembly. Understanding the molecular mechanism underlying the dual effects of the core protein allosteric modulators on nucleocapsid assembly and disassembly will facilitate the discovery of novel core protein-targeting antiviral agents that can more efficiently suppress cccDNA synthesis and cure chronic hepatitis B.


id: cord-002835-qaogpxy9
title: Prohibitin plays a critical role in Enterovirus 71 neuropathogenesis
author: Too, Issac Horng Khit
date: 2018-01-11
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5764453/
doi: 10.1371/journal.ppat.1006778
abstract:
  A close relative of poliovirus, enterovirus 71 (EV71) is regarded as an important neurotropic virus of serious public health concern. EV71 causes Hand, Foot and Mouth Disease and has been associated with neurological complications in young children. Our limited understanding of the mechanisms involved in its neuropathogenesis has hampered the development of effective therapeutic options. Here, using a two-dimensional proteomics approach combined with mass spectrometry, we have identified a unique panel of host proteins that were differentially and dynamically modulated during EV71 infection of motor-neuron NSC-34 cells, which are found at the neuromuscular junctions where EV71 is believed to enter the central nervous system. Meta-analysis with previously published proteomics studies in neuroblastoma or muscle cell lines revealed minimal overlapping which suggests unique host-pathogen interactions in NSC-34 cells. Among the candidate proteins, we focused our attention on prohibitin (PHB), a protein that is involved in multiple cellular functions and the target of anti-cancer drug Rocaglamide (Roc-A). We demonstrated that cell surface-expressed PHB is involved in EV71 entry into neuronal cells specifically, while membrane-bound mitochondrial PHB associates with the virus replication complex and facilitates viral replication. Furthermore, Roc-A treatment of EV71-infected neuronal cells reduced significantly virus yields. However, the inhibitory effect of Roc-A on PHB in NSC-34 cells was not through blocking the CRAF/MEK/ERK pathway as previously reported. Instead, Roc-A treated NSC-34 cells had lower mitochondria-associated PHB and lower ATP levels that correlated with impaired mitochondria integrity. In vivo, EV71-infected mice treated with Roc-A survived longer than the vehicle-treated animals and had significantly lower virus loads in their spinal cord and brain, whereas virus titers in their limb muscles were comparable to controls. Together, this study uncovers PHB as the first host factor that is specifically involved in EV71 neuropathogenesis and a potential drug target to limit neurological complications.


id: cord-002893-d7hetoq0
author: Meng, Xiangzhi
date: 2018-02-15
txt: ./txt/cord-002893-d7hetoq0.txt
cache: ./cache/cord-002893-d7hetoq0.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5831749/
doi: 10.1371/journal.ppat.1006884
summary:
  Human SAMD9, a tumor suppressor and a restriction factor for poxviruses in cell lines, is antagonized by two classes of poxvirus proteins, represented by vaccinia virus (VACV) K1 and C7. Both paralogs are antagonized by VACV K1, C7 and C7 homologs from diverse mammalian poxviruses, but mouse SAMD9L is resistant to the C7 homolog encoded by a group of poxviruses with a narrow host range in ruminants, indicating that host species-specific difference in SAMD9/SAMD9L genes serves as a barrier for cross-species poxvirus transmission. VACV K1, C7 and C7 homologs from diverse poxviruses could overcome hSAMD9L restriction, but the sheeppox virus C7 homolog has reduced potency due to the variation of two residues To find out whether mammalian poxviruses could overcome the restriction of hSAMD9L, we induced hSAMD9L expression in ΔhSAMD9 HeLa cells with 200 U/ml of IFN-β and then infected the cells with our panel of vK1L -C7L --derived VACVs. Viruses that expressed VACV-K1, VACV-C7, MYXV-M62, SPPV-063, SWPV-064 and YLDV-67 (but not MYXV-M63 and MYXV-M64) grew in IFN-treated ΔhSAMD9 cells (Fig 5A) , indicating that all known SAMD9 antagonists could also antagonize hSAMD9L.

title:
  A paralogous pair of mammalian host restriction factors form a critical host barrier against poxvirus infection

abstract:
  Host restriction factors constitute a formidable barrier for viral replication to which many viruses have evolved counter-measures. Human SAMD9, a tumor suppressor and a restriction factor for poxviruses in cell lines, is antagonized by two classes of poxvirus proteins, represented by vaccinia virus (VACV) K1 and C7. A paralog of SAMD9, SAMD9L, is also encoded by some mammals, while only one of two paralogs is retained by others. Here, we show that SAMD9L functions similarly to SAMD9 as a restriction factor and that the two paralogs form a critical host barrier that poxviruses must overcome to establish infection. In mice, which naturally lack SAMD9, overcoming SAMD9L restriction with viral inhibitors is essential for poxvirus replication and pathogenesis. While a VACV deleted of both K1 and C7 (vK1L(-)C7L(-)) was restricted by mouse cells and highly attenuated in mice, its replication and virulence were completely restored in SAMD9L(-/-) mice. In humans, both SAMD9 and SAMD9L are poxvirus restriction factors, although the latter requires interferon induction in many cell types. While knockout of SAMD9 with Crispr-Cas9 was sufficient for abolishing the restriction for vK1L(-)C7L(-) in many human cells, knockout of both paralogs was required for abolishing the restriction in interferon-treated cells. Both paralogs are antagonized by VACV K1, C7 and C7 homologs from diverse mammalian poxviruses, but mouse SAMD9L is resistant to the C7 homolog encoded by a group of poxviruses with a narrow host range in ruminants, indicating that host species-specific difference in SAMD9/SAMD9L genes serves as a barrier for cross-species poxvirus transmission.


id: cord-003169-bdw5ke4i
author: Guo, Hongbo
date: 2018-08-13
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6107293/
doi: 10.1371/journal.ppat.1007233
title:
  Kinetic analysis of the influenza A virus HA/NA balance reveals contribution of NA to virus-receptor binding and NA-dependent rolling on receptor-containing surfaces

abstract:
  Interactions of influenza A virus (IAV) with sialic acid (SIA) receptors determine viral fitness and host tropism. Binding to mucus decoy receptors and receptors on epithelial host cells is determined by a receptor-binding hemagglutinin (HA), a receptor-destroying neuraminidase (NA) and a complex in vivo receptor-repertoire. The crucial but poorly understood dynamics of these multivalent virus-receptor interactions cannot be properly analyzed using equilibrium binding models and endpoint binding assays. In this study, the use of biolayer interferometric analysis revealed the virtually irreversible nature of IAV binding to surfaces coated with synthetic sialosides or engineered sialoglycoproteins in the absence of NA activity. In addition to HA, NA was shown to be able to contribute to the initial binding rate while catalytically active. Virus-receptor binding in turn contributed to receptor cleavage by NA. Multiple low-affinity HA-SIA interactions resulted in overall extremely high avidity but also permitted a dynamic binding mode, in which NA activity was driving rolling of virus particles over the receptor-surface. Virus dissociation only took place after receptor density of the complete receptor-surface was sufficiently decreased due to NA activity of rolling IAV particles. The results indicate that in vivo IAV particles, after landing on the mucus layer, reside continuously in a receptor-bound state while rolling through the mucus layer and over epithelial cell surfaces driven by the HA-NA-receptor balance. Quantitative BLI analysis enabled functional examination of this balance which governs this dynamic and motile interaction that is expected to be crucial for penetration of the mucus layer and subsequent infection of cells by IAV but likely also by other enveloped viruses carrying a receptor-destroying enzyme in addition to a receptor-binding protein.


id: cord-003441-810r5q03
author: Dzimianski, John V.
date: 2019-01-10
txt: ./txt/cord-003441-810r5q03.txt
cache: ./cache/cord-003441-810r5q03.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6343935/
doi: 10.1371/journal.ppat.1007515
summary:
  Specifically, that CCHFV vOTU DUB activity is not as promiscuous towards ubiquitinated host proteins as it first seemed based on the overexpression studies, but appears to be restricted to a targeted subset of cellular substrates associated with suppression of RIG-I-mediated early cellular responses to infection. Additionally, a structure of the FARV vOTU provides details into the structural nature of the additional residues in Hughes orthonairovirus vOTUs. Structureinformed mutagenesis of FARV vOTU identified residues involved specifically in di-Ub binding, representing the first report of the role of a second site involved in di-Ub binding in nairovirus vOTUs. This novel enzymatic and structural data not only provides insight into the nature of vOTU diversity, but also lays a foundation for understanding the impact of the vOTU interaction with the innate immune response and its connection to viral pathogenesis. Intriguingly, the vOTUs showed a diverse range of activity towards Ub. In general, vOTUs can be divided into groups possessing high (CCHFV, HAZV, NSDV/GANV, TAGV), moderate (DUGV, KUPEV, FARV, QYBV, ISKV), or low activity (ERVEV, DGKV, LPHV, HpTV-1) (Fig 2A) .

title:
  Probing the impact of nairovirus genomic diversity on viral ovarian tumor domain protease (vOTU) structure and deubiquitinase activity

abstract:
  Post-translational modification of host and viral proteins by ubiquitin (Ub) and Ub-like proteins, such as interferon stimulated gene product 15 (ISG15), plays a key role in response to infection. Viruses have been increasingly identified that contain proteases possessing deubiquitinase (DUB) and/or deISGylase functions. This includes viruses in the Nairoviridae family that encode a viral homologue of the ovarian tumor protease (vOTU). vOTU activity was recently demonstrated to be critical for replication of the often-fatal Crimean-Congo hemorrhagic fever virus, with DUB activity suppressing the type I interferon responses and deISGylase activity broadly removing ISG15 conjugated proteins. There are currently about 40 known nairoviruses classified into fourteen species. Recent genomic characterization has revealed a high degree of diversity, with vOTUs showing less than 25% amino acids identities within the family. Previous investigations have been limited to only a few closely related nairoviruses, leaving it unclear what impact this diversity has on vOTU function. To probe the effects of vOTU diversity on enzyme activity and specificity, we assessed representative vOTUs spanning the Nairoviridae family towards Ub and ISG15 fluorogenic substrates. This revealed great variation in enzymatic activity and specific substrate preferences. A subset of the vOTUs were further assayed against eight biologically relevant di-Ub substrates, uncovering both common trends and distinct preferences of poly-Ub linkages by vOTUs. Four novel X-ray crystal structures were obtained that provide a biochemical rationale for vOTU substrate preferences and elucidate structural features that distinguish the vOTUs, including a motif in the Hughes orthonairovirus species that has not been previously observed in OTU domains. Additionally, structure-informed mutagenesis provided the first direct evidence of a second site involved in di-Ub binding for vOTUs. These results provide new insight into nairovirus evolution and pathogenesis, and further enhances the development of tools for therapeutic purposes.


id: cord-003982-7v5xl1s3
author: Trus, Ivan
date: 2019-11-14
txt: ./txt/cord-003982-7v5xl1s3.txt
cache: ./cache/cord-003982-7v5xl1s3.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6855438/
doi: 10.1371/journal.ppat.1008038
summary:
  title: Subclinical in utero Zika virus infection is associated with interferon alpha sequelae and sex-specific molecular brain pathology in asymptomatic porcine offspring Collectively, our results provide strong evidence that two hallmarks of fetal ZIKV infection, altered type I IFN response and molecular brain pathology can persist after birth in offspring in the absence of congenital Zika syndrome. In our porcine model studies, subclinical persistent in utero infection in mid-gestation also increased interferon alpha (IFN-α) levels in fetal blood plasma and amniotic fluid, while IFN-α was below the detection limit in all control fetuses [18] . We performed a mixing test (S2 Video) on control and affected piglets at 35 days of age to identify whether subclinical in utero ZIKV infection and social stress have synergistic effects on cytokine responses in offspring.

title:
  Subclinical in utero Zika virus infection is associated with interferon alpha sequelae and sex-specific molecular brain pathology in asymptomatic porcine offspring

abstract:
  Zika virus (ZIKV) infection during human pregnancy may lead to severe fetal pathology and debilitating impairments in offspring. However, the majority of infections are subclinical and not associated with evident birth defects. Potentially detrimental life-long health outcomes in asymptomatic offspring evoke high concerns. Thus, animal models addressing sequelae in offspring may provide valuable information. To induce subclinical infection, we inoculated selected porcine fetuses at the mid-stage of development. Inoculation resulted in trans-fetal virus spread and persistent infection in the placenta and fetal membranes for two months. Offspring did not show congenital Zika syndrome (e.g., microcephaly, brain calcifications, congenital clubfoot, arthrogryposis, seizures) or other visible birth defects. However, a month after birth, a portion of offspring exhibited excessive interferon alpha (IFN-α) levels in blood plasma in a regular environment. Most affected offspring also showed dramatic IFN-α shutdown during social stress providing the first evidence for the cumulative impact of prenatal ZIKV exposure and postnatal environmental insult. Other eleven cytokines tested before and after stress were not altered suggesting the specific IFN-α pathology. While brains from offspring did not have histopathology, lesions, and ZIKV, the whole genome expression analysis of the prefrontal cortex revealed profound sex-specific transcriptional changes that most probably was the result of subclinical in utero infection. RNA-seq analysis in the placenta persistently infected with ZIKV provided independent support for the sex-specific pattern of in utero-acquired transcriptional responses. Collectively, our results provide strong evidence that two hallmarks of fetal ZIKV infection, altered type I IFN response and molecular brain pathology can persist after birth in offspring in the absence of congenital Zika syndrome.


id: cord-004428-ob5igsv5
author: Kizziah, James L.
date: 2020-02-18
txt: ./txt/cord-004428-ob5igsv5.txt
cache: ./cache/cord-004428-ob5igsv5.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7048315/
doi: 10.1371/journal.ppat.1008314
summary:
  Crystal structures have also been determined for Bacillus phage SPP1 Dit [25] and the baseplates of lactococcal phages p2 and TP901-1, which include Dit, Tal, BppU, and receptor binding proteins that are not similar to the ϕ11 RBP [23, 26] . Atomic models for MTP (gp53), Dit (gp58), RBP (gp61), and parts of Tal (gp59), FibL (gp62), FibU (gp68) and TMP (gp57) were built into either the C6 or C3 baseplate reconstructions, using I-TASSER models as starting points [29] , complemented with real-space refinement in PHENIX [30] . RBP is a trimeric protein consisting of four domains: an N-terminal α-helical coiled-coil "stem" domain (residues 1-142) that is folded by 155˚at a central hinge; a fivebladed β-propeller "platform" domain (residues 143-442) that was previously suggested to bind to the GlcNAc residues of WTA on the host cell surface [21, 24] ; and two highly similar C-terminal "tower" domains (residues 443-636; S1 Table) , each consisting of a bent β-sheet superposed by an α-helix (Fig 5A) .

title:
  Structure of the host cell recognition and penetration machinery of a Staphylococcus aureus bacteriophage

abstract:
  Staphylococcus aureus is a common cause of infections in humans. The emergence of virulent, antibiotic-resistant strains of S. aureus is a significant public health concern. Most virulence and resistance factors in S. aureus are encoded by mobile genetic elements, and transduction by bacteriophages represents the main mechanism for horizontal gene transfer. The baseplate is a specialized structure at the tip of bacteriophage tails that plays key roles in host recognition, cell wall penetration, and DNA ejection. We have used high-resolution cryo-electron microscopy to determine the structure of the S. aureus bacteriophage 80α baseplate at 3.75 Å resolution, allowing atomic models to be built for most of the major tail and baseplate proteins, including two tail fibers, the receptor binding protein, and part of the tape measure protein. Our structure provides a structural basis for understanding host recognition, cell wall penetration and DNA ejection in viruses infecting Gram-positive bacteria. Comparison to other phages demonstrates the modular design of baseplate proteins, and the adaptations to the host that take place during the evolution of staphylococci and other pathogens.


id: cord-010762-c01wgg4v
author: Wu, Jiqin
date: 2020-05-01
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7219791/
doi: 10.1371/journal.ppat.1008484
title:
  A conformation-based intra-molecular initiation factor identified in the flavivirus RNA-dependent RNA polymerase

abstract:
  The flaviviruses pose serious threats to human health. Being a natural fusion of a methyltransferase (MTase) and an RNA-dependent RNA polymerase (RdRP), NS5 is the most conserved flavivirus protein and an important antiviral target. Previously reported NS5 structures represented by those from the Japanese encephalitis virus (JEV) and Dengue virus serotype 3 (DENV3) exhibit two apparently different global conformations, defining two sets of intra-molecular MTase-RdRP interactions. However, whether these NS5 conformations are conserved in flaviviruses and their specific functions remain elusive. Here we report two forms of DENV serotype 2 (DENV2) NS5 crystal structures representing two conformational states with defined analogies to the JEV-mode and DENV3-mode conformations, respectively, demonstrating the conservation of both conformation modes and providing clues for how different conformational states may be interconnected. Data from in vitro polymerase assays further demonstrate that perturbing the JEV-mode but not the DENV3-mode intra-molecular interactions inhibits catalysis only at initiation, while the cell-based virological analysis suggests that both modes of interactions are important for virus proliferation. Our work highlights the role of MTase as a unique intra-molecular initiation factor specifically only through the JEV-mode conformation, providing an example of conformation-based crosstalk between naturally fused protein functional modules.


id: cord-011380-kfkp1zpn
title: Tuning antiviral CD8 T-cell response via proline-altered peptide ligand vaccination
author: Duru, Adil Doganay
date: 2020-05-04
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7224568/
doi: 10.1371/journal.ppat.1008244
abstract:
  Viral escape from CD8(+) cytotoxic T lymphocyte responses correlates with disease progression and represents a significant challenge for vaccination. Here, we demonstrate that CD8(+) T cell recognition of the naturally occurring MHC-I-restricted LCMV-associated immune escape variant Y4F is restored following vaccination with a proline-altered peptide ligand (APL). The APL increases MHC/peptide (pMHC) complex stability, rigidifies the peptide and facilitates T cell receptor (TCR) recognition through reduced entropy costs. Structural analyses of pMHC complexes before and after TCR binding, combined with biophysical analyses, revealed that although the TCR binds similarly to all complexes, the p3P modification alters the conformations of a very limited amount of specific MHC and peptide residues, facilitating efficient TCR recognition. This approach can be easily introduced in peptides restricted to other MHC alleles, and can be combined with currently available and future vaccination protocols in order to prevent viral immune escape.


id: cord-013481-3zwq67do
author: Guo, Kejun
date: 2020-10-16
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7592919/
doi: 10.1371/journal.ppat.1008986
title:
  Qualitative Differences Between the IFNα subtypes and IFNβ Influence Chronic Mucosal HIV-1 Pathogenesis

abstract:
  The Type I Interferons (IFN-Is) are innate antiviral cytokines that include 12 different IFNα subtypes and IFNβ that signal through the IFN-I receptor (IFNAR), inducing hundreds of IFN-stimulated genes (ISGs) that comprise the ‘interferome’. Quantitative differences in IFNAR binding correlate with antiviral activity, but whether IFN-Is exhibit qualitative differences remains controversial. Moreover, the IFN-I response is protective during acute HIV-1 infection, but likely pathogenic during the chronic stages. To gain a deeper understanding of the IFN-I response, we compared the interferomes of IFNα subtypes dominantly-expressed in HIV-1-exposed plasmacytoid dendritic cells (1, 2, 5, 8 and 14) and IFNβ in the earliest cellular targets of HIV-1 infection. Primary gut CD4 T cells from 3 donors were treated for 18 hours ex vivo with individual IFN-Is normalized for IFNAR signaling strength. Of 1,969 IFN-regulated genes, 246 ‘core ISGs’ were induced by all IFN-Is tested. However, many IFN-regulated genes were not shared between the IFNα subtypes despite similar induction of canonical antiviral ISGs such as ISG15, RSAD2 and MX1, formally demonstrating qualitative differences between the IFNα subtypes. Notably, IFNβ induced a broader interferome than the individual IFNα subtypes. Since IFNβ, and not IFNα, is upregulated during chronic HIV-1 infection in the gut, we compared core ISGs and IFNβ-specific ISGs from colon pinch biopsies of HIV-1-uninfected (n = 13) versus age- and gender-matched, antiretroviral-therapy naïve persons with HIV-1 (PWH; n = 19). Core ISGs linked to inflammation, T cell activation and immune exhaustion were elevated in PWH, positively correlated with plasma lipopolysaccharide (LPS) levels and gut IFNβ levels, and negatively correlated with gut CD4 T cell frequencies. In sharp contrast, IFNβ-specific ISGs linked to protein translation and anti-inflammatory responses were significantly downregulated in PWH, negatively correlated with gut IFNβ and LPS, and positively correlated with plasma IL6 and gut CD4 T cell frequencies. Our findings reveal qualitative differences in interferome induction by diverse IFN-Is and suggest potential mechanisms for how IFNβ may drive HIV-1 pathogenesis in the gut.


id: cord-013526-6fip93l2
author: Labadie, Thomas
date: 2020-10-19
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7595637/
doi: 10.1371/journal.ppat.1009015
title:
  A non-enveloped arbovirus released in lysosome-derived extracellular vesicles induces super-infection exclusion

abstract:
  Recent developments on extracellular vesicles (EVs) containing multiple virus particles challenge the rigid definition of non-enveloped viruses. However, how non-enveloped viruses hijack cell machinery to promote non-lytic release in EVs, and their functional roles, remain to be clarified. Here we used Bluetongue virus (BTV) as a model of a non-enveloped arthropod-borne virus and discovered that the majority of viruses are released in EVs. Based on the cellular proteins detected in these EVs, and use of inhibitors targeting the cellular degradation process, we demonstrated that these extracellular vesicles are derived from secretory lysosomes, in which the acidic pH is neutralized upon the infection. Moreover, we report that secreted EVs are more efficient than free-viruses for initiating infections, but that they trigger super-infection exclusion that only free-viruses can overcome.


id: cord-076082-4kpkhz0o
title: Evolutionary and Transmission Dynamics of Reassortant H5N1 Influenza Virus in Indonesia
author: Lam, Tommy Tsan-Yuk
date: 2008-08-22
url: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2515348/
doi: 10.1371/journal.ppat.1000130
abstract:
  H5N1 highly pathogenic avian influenza (HPAI) viruses have seriously affected the Asian poultry industry since their recurrence in 2003. The viruses pose a threat of emergence of a global pandemic influenza through point mutation or reassortment leading to a strain that can effectively transmit among humans. In this study, we present phylogenetic evidences for the interlineage reassortment among H5N1 HPAI viruses isolated from humans, cats, and birds in Indonesia, and identify the potential genetic parents of the reassorted genome segments. Parsimony analyses of viral phylogeography suggest that the reassortant viruses may have originated from greater Jakarta and surroundings, and subsequently spread to other regions in the West Java province. In addition, Bayesian methods were used to elucidate the genetic diversity dynamics of the reassortant strain and one of its genetic parents, which revealed a more rapid initial growth of genetic diversity in the reassortant viruses relative to their genetic parent. These results demonstrate that interlineage exchange of genetic information may play a pivotal role in determining viral genetic diversity in a focal population. Moreover, our study also revealed significantly stronger diversifying selection on the M1 and PB2 genes in the lineages preceding and subsequent to the emergence of the reassortant viruses, respectively. We discuss how the corresponding mutations might drive the adaptation and onward transmission of the newly formed reassortant viruses.


id: cord-253783-3a1qde41
title: Prions Adhere to Soil Minerals and Remain Infectious
author: Johnson, Christopher J
date: 2006-04-14
txt: ./txt/cord-253783-3a1qde41.txt
cache: ./cache/cord-253783-3a1qde41.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/16617377/
doi: 10.1371/journal.ppat.0020032
summary:
  We examined the potential for soil to serve as a TSE reservoir by studying the interaction of the disease-associated prion protein (PrP(Sc)) with common soil minerals. X-ray diffraction analysis provided no evidence that PrP Sc entered Mte interlayer spaces (Mte d 001 spacings were 1.22 nm and 1.47 nm before and after PrP Sc adsorption, respectively); prion protein appeared to adsorb to only external clay surfaces. Prion protein desorbed from Kte and quartz did not exhibit a change in molecular mass ( Figure 1A ), suggesting that surface properties specific to Mte were responsible for the cleavage. To control for any unbound prion protein that may have cosedimented with Mte particles, mineral-free PrP Sc suspensions were processed in the same manner as in sorption experiments. Our results demonstrate that all soil mineral surfaces examined bound PrP Sc and that Mte and quartz have larger specific binding capacities for PrP Sc than does Kte (Figure 1 ).

abstract:
  An unidentified environmental reservoir of infectivity contributes to the natural transmission of prion diseases (transmissible spongiform encephalopathies [TSEs]) in sheep, deer, and elk. Prion infectivity may enter soil environments via shedding from diseased animals and decomposition of infected carcasses. Burial of TSE-infected cattle, sheep, and deer as a means of disposal has resulted in unintentional introduction of prions into subsurface environments. We examined the potential for soil to serve as a TSE reservoir by studying the interaction of the disease-associated prion protein (PrP(Sc)) with common soil minerals. In this study, we demonstrated substantial PrP(Sc) adsorption to two clay minerals, quartz, and four whole soil samples. We quantified the PrP(Sc)-binding capacities of each mineral. Furthermore, we observed that PrP(Sc) desorbed from montmorillonite clay was cleaved at an N-terminal site and the interaction between PrP(Sc) and Mte was strong, making desorption of the protein difficult. Despite cleavage and avid binding, PrP(Sc) bound to Mte remained infectious. Results from our study suggest that PrP(Sc) released into soil environments may be preserved in a bioavailable form, perpetuating prion disease epizootics and exposing other species to the infectious agent.


id: cord-254735-8reu45yz
author: Reguera, Juan
date: 2012-08-02
url: https://doi.org/10.1371/journal.ppat.1002859
doi: 10.1371/journal.ppat.1002859
title:
  Structural Bases of Coronavirus Attachment to Host Aminopeptidase N and Its Inhibition by Neutralizing Antibodies

abstract:
  The coronaviruses (CoVs) are enveloped viruses of animals and humans associated mostly with enteric and respiratory diseases, such as the severe acute respiratory syndrome and 10–20% of all common colds. A subset of CoVs uses the cell surface aminopeptidase N (APN), a membrane-bound metalloprotease, as a cell entry receptor. In these viruses, the envelope spike glycoprotein (S) mediates the attachment of the virus particles to APN and subsequent cell entry, which can be blocked by neutralizing antibodies. Here we describe the crystal structures of the receptor-binding domains (RBDs) of two closely related CoV strains, transmissible gastroenteritis virus (TGEV) and porcine respiratory CoV (PRCV), in complex with their receptor, porcine APN (pAPN), or with a neutralizing antibody. The data provide detailed information on the architecture of the dimeric pAPN ectodomain and its interaction with the CoV S. We show that a protruding receptor-binding edge in the S determines virus-binding specificity for recessed glycan-containing surfaces in the membrane-distal region of the pAPN ectodomain. Comparison of the RBDs of TGEV and PRCV to those of other related CoVs, suggests that the conformation of the S receptor-binding region determines cell entry receptor specificity. Moreover, the receptor-binding edge is a major antigenic determinant in the TGEV envelope S that is targeted by neutralizing antibodies. Our results provide a compelling view on CoV cell entry and immune neutralization, and may aid the design of antivirals or CoV vaccines. APN is also considered a target for cancer therapy and its structure, reported here, could facilitate the development of anti-cancer drugs.


id: cord-258234-qn8xp4v9
title: Inhibitors of Peptidyl Proline Isomerases As Antivirals in Hepatitis C and Other Viruses
author: Striker, Rob
date: 2014-11-06
txt: ./txt/cord-258234-qn8xp4v9.txt
cache: ./cache/cord-258234-qn8xp4v9.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1004428
doi: 10.1371/journal.ppat.1004428
abstract: nan
summary:
  Cyclophilin inhibitors that reduce viral replication also block interactions between cyclophilin A and NS5A, suggesting that this association is important during the viral life cycle and might be the relevant target of the antiviral activity of cyclosporine [11] [12] [13] . HCV NS5A is a protein that is rich in both proline residues and disorder and that associates with cyclophilin A via domain 2. However, these regions share several common properties: the flexible cyclophilin-interacting loop in CA is also bracketed by prolines, just as the PAWARPDYNP motif in HCV; residues 86, 91, and 96 that surround the glycine-proline in CA influence viral susceptibility to cyclophillin inhibitors similar to mutations surrounding NS5A P319 [25] ; and the critical glycine-proline is amino-terminal to regions of CA that are actually more proline rich and predicted by bioinformatics analysis to be disordered, analogous to the positioning of the PAWARPDYNP motif in HCV NS5A.


id: cord-260729-b12v3c8c
author: de Lang, Anna
date: 2007-08-10
url: https://www.ncbi.nlm.nih.gov/pubmed/17696609/
doi: 10.1371/journal.ppat.0030112
title:
  Functional Genomics Highlights Differential Induction of Antiviral Pathways in the Lungs of SARS-CoV–Infected Macaques

abstract:
  The pathogenesis of severe acute respiratory syndrome coronavirus (SARS-CoV) is likely mediated by disproportional immune responses and the ability of the virus to circumvent innate immunity. Using functional genomics, we analyzed early host responses to SARS-CoV infection in the lungs of adolescent cynomolgus macaques (Macaca fascicularis) that show lung pathology similar to that observed in human adults with SARS. Analysis of gene signatures revealed induction of a strong innate immune response characterized by the stimulation of various cytokine and chemokine genes, including interleukin (IL)-6, IL-8, and IP-10, which corresponds to the host response seen in acute respiratory distress syndrome. As opposed to many in vitro experiments, SARS-CoV induced a wide range of type I interferons (IFNs) and nuclear translocation of phosphorylated signal transducer and activator of transcription 1 in the lungs of macaques. Using immunohistochemistry, we revealed that these antiviral signaling pathways were differentially regulated in distinctive subsets of cells. Our studies emphasize that the induction of early IFN signaling may be critical to confer protection against SARS-CoV infection and highlight the strength of combining functional genomics with immunohistochemistry to further unravel the pathogenesis of SARS.


id: cord-260949-w2xuf15h
title: Viral Control of Mitochondrial Apoptosis
author: Galluzzi, Lorenzo
date: 2008-05-30
txt: ./txt/cord-260949-w2xuf15h.txt
cache: ./cache/cord-260949-w2xuf15h.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1000018
doi: 10.1371/journal.ppat.1000018
summary:
  Bcl-2/Bcl-X L stabilizes mitochondrial membranes via multiple mechanisms, including (1) the sequestration into inactive complexes of its proapoptotic counterparts, Bax, Bak, and BH3only proteins (e.g., Bid) (for review: [S65,S78]), (2) inhibitory interactions with PTPC constituents, in particular with VDAC and the adenine nucleotide translocase (ANT) [12, 14] , (3) an enhancement of Cyt c oxidase activity and mitochondrial respiration [S79], and/or (4) indirect effects on intracellular Ca 2+ stores of the ER [S80,S81]. With regard to this, viral factors can be classified into one of the four following subgroups: proapoptotic proteins (1) that insert into mitochondrial membranes and hence trigger MMP through the action of amphipathic a-helical domains or (2) that promote MMP indirectly, through the activition of host-encoded factors (Table 1) , and antiapoptotic modulators (3) that exhibit sequence and/or structural similarity to multidomain BH1-4 members of the Bcl-2 family (so-called viral Bcl-2 proteins [vBcl-2s]) or (4) that inhibit apoptosis via other mechanisms (Table 2) .

abstract:
  Throughout the process of pathogen–host co-evolution, viruses have developed a battery of distinct strategies to overcome biochemical and immunological defenses of the host. Thus, viruses have acquired the capacity to subvert host cell apoptosis, control inflammatory responses, and evade immune reactions. Since the elimination of infected cells via programmed cell death is one of the most ancestral defense mechanisms against infection, disabling host cell apoptosis might represent an almost obligate step in the viral life cycle. Conversely, viruses may take advantage of stimulating apoptosis, either to kill uninfected cells from the immune system, or to induce the breakdown of infected cells, thereby favoring viral dissemination. Several viral polypeptides are homologs of host-derived apoptosis-regulatory proteins, such as members of the Bcl-2 family. Moreover, viral factors with no homology to host proteins specifically target key components of the apoptotic machinery. Here, we summarize the current knowledge on the viral modulation of mitochondrial apoptosis, by focusing in particular on the mechanisms by which viral proteins control the host cell death apparatus.


id: cord-262248-q5eijunp
title: The Ebola Epidemic Crystallizes the Potential of Passive Antibody Therapy for Infectious Diseases
author: Casadevall, Arturo
date: 2015-04-23
txt: ./txt/cord-262248-q5eijunp.txt
cache: ./cache/cord-262248-q5eijunp.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/25905897/
doi: 10.1371/journal.ppat.1004717
abstract: nan
summary:
  Currently, there are no drugs to treat Ebola, but, in the urgency and emergency triggered by the epidemic, two types of Ab-based therapies have been used: convalescent sera from patients who have recovered and a mAb cocktail known as ZMapp produced in plants [3] . For example, given that there is increasing evidence that antibiotic-induced disruptions of the microbiota are associated with deleterious effects in the treatment of bacterial diseases, the high specificity of Ab-based therapies offers a tremendous advantage. Regarding the limitation that highly specific Abs can fail to bind highly related variants, particularly for viruses, there are two strategies to overcome this problem: the development of Abs that target broadly neutralizing epitopes, as has now been demonstrated for HIV [8] , dengue [9] , and influenza viruses [10] , and the generation of Ab cocktails composed of Abs against multiple serotypes or variants, as has been done for rabies virus [11] and Clostridium difficile toxin [12] .


id: cord-262753-jld1ygxt
title: Global analysis of polysome-associated mRNA in vesicular stomatitis virus infected cells
author: Neidermyer, William J.
date: 2019-06-21
url: https://www.ncbi.nlm.nih.gov/pubmed/31226162/
doi: 10.1371/journal.ppat.1007875
abstract:
  Infection of mammalian cells with vesicular stomatitis virus (VSV) results in the inhibition of cellular translation while viral translation proceeds efficiently. VSV RNA synthesis occurs entirely within the cytoplasm, where during transcription the viral polymerase produces 5 mRNAs that are structurally indistinct to cellular mRNAs with respect to their 5′ cap-structure and 3′-polyadenylate tail. Using the global approach of massively parallel sequencing of total cytoplasmic, monosome- and polysome-associated mRNA, we interrogate the impact of VSV infection of HeLa cells on translation. Analysis of sequence reads in the different fractions shows >60% of total cytoplasmic and polysome-associated reads map to the 5 viral genes by 6 hours post-infection, a time point at which robust host cell translational shut-off is observed. Consistent with an overwhelming abundance of viral mRNA in the polysome fraction, the reads mapping to cellular genes were reduced. The cellular mRNAs that remain most polysome-associated following infection had longer half-lives, were typically larger, and were more AU rich, features that are shared with the viral mRNAs. Several of those mRNAs encode proteins known to positively affect viral replication, and using chemical inhibition and siRNA depletion we confirm that the host chaperone heat shock protein 90 (hsp90) and eukaryotic translation initiation factor 3A (eIF3A)—encoded by 2 such mRNAs—support viral replication. Correspondingly, regulated in development and DNA damage 1 (Redd1) encoded by a host mRNA with reduced polysome association inhibits viral infection. These data underscore the importance of viral mRNA abundance in the shut-off of host translation in VSV infected cells and link the differential translatability of some cellular mRNAs with pro- or antiviral function.


id: cord-263239-andje0wu
author: Dorobantu, Cristina M.
date: 2015-09-25
url: https://doi.org/10.1371/journal.ppat.1005185
doi: 10.1371/journal.ppat.1005185
title:
  Modulation of the Host Lipid Landscape to Promote RNA Virus Replication: The Picornavirus Encephalomyocarditis Virus Converges on the Pathway Used by Hepatitis C Virus

abstract:
  Cardioviruses, including encephalomyocarditis virus (EMCV) and the human Saffold virus, are small non-enveloped viruses belonging to the Picornaviridae, a large family of positive-sense RNA [(+)RNA] viruses. All (+)RNA viruses remodel intracellular membranes into unique structures for viral genome replication. Accumulating evidence suggests that picornaviruses from different genera use different strategies to generate viral replication organelles (ROs). For instance, enteroviruses (e.g. poliovirus, coxsackievirus, rhinovirus) rely on the Golgi-localized phosphatidylinositol 4-kinase III beta (PI4KB), while cardioviruses replicate independently of the kinase. By which mechanisms cardioviruses develop their ROs is currently unknown. Here we show that cardioviruses manipulate another PI4K, namely the ER-localized phosphatidylinositol 4-kinase III alpha (PI4KA), to generate PI4P-enriched ROs. By siRNA-mediated knockdown and pharmacological inhibition, we demonstrate that PI4KA is an essential host factor for EMCV genome replication. We reveal that the EMCV nonstructural protein 3A interacts with and is responsible for PI4KA recruitment to viral ROs. The ensuing phosphatidylinositol 4-phosphate (PI4P) proved important for the recruitment of oxysterol-binding protein (OSBP), which delivers cholesterol to EMCV ROs in a PI4P-dependent manner. PI4P lipids and cholesterol are shown to be required for the global organization of the ROs and for viral genome replication. Consistently, inhibition of OSBP expression or function efficiently blocked EMCV RNA replication. In conclusion, we describe for the first time a cellular pathway involved in the biogenesis of cardiovirus ROs. Remarkably, the same pathway was reported to promote formation of the replication sites of hepatitis C virus, a member of the Flaviviridae family, but not other picornaviruses or flaviviruses. Thus, our results highlight the convergent recruitment by distantly related (+)RNA viruses of a host lipid-modifying pathway underlying formation of viral replication sites.


id: cord-265947-7j9qqi8w
author: Du, Wenjuan
date: 2019-06-10
txt: ./txt/cord-265947-7j9qqi8w.txt
cache: ./cache/cord-265947-7j9qqi8w.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/31181126/
doi: 10.1371/journal.ppat.1007860
summary:
  NAs of avian, but not human viruses, contain a functional 2(nd) sialic acid (SIA)-binding site (2SBS) adjacent to the catalytic site, which contributes to sialidase activity against multivalent substrates. Using a recently established BLI-based kinetic binding assay [37] an enhanced initial binding rate to 3''SLNLN but not 6''SLNLN ( Fig 4A and 4B ), was observed for hH3aN2 virus containing Role for 2 nd SIA-binding site of IAV NA in HA-NA-receptor balance a functional 2SBS in comparison to hH3hN2. We therefore studied the effect of the 2SBS in NA when combined with an avian-type Role for 2 nd SIA-binding site of IAV NA in HA-NA-receptor balance HA preferring binding to α2,3-linked SIAs. We generated the corresponding recombinant A/ Hong Kong/1/68 (H3N2) viruses containing 7 amino acid substitutions in the HA (see S8A Fig) .

title:
  The 2(nd) sialic acid-binding site of influenza A virus neuraminidase is an important determinant of the hemagglutinin-neuraminidase-receptor balance

abstract:
  Influenza A virus (IAV) neuraminidase (NA) receptor-destroying activity and hemagglutinin (HA) receptor-binding affinity need to be balanced with the host receptor repertoire for optimal viral fitness. NAs of avian, but not human viruses, contain a functional 2(nd) sialic acid (SIA)-binding site (2SBS) adjacent to the catalytic site, which contributes to sialidase activity against multivalent substrates. The receptor-binding specificity and potentially crucial contribution of the 2SBS to the HA-NA balance of virus particles is, however, poorly characterized. Here, we elucidated the receptor-binding specificity of the 2SBS of N2 NA and established an important role for this site in the virion HA-NA-receptor balance. NAs of H2N2/1957 pandemic virus with or without a functional 2SBS and viruses containing this NA were analysed. Avian-like N2, with a restored 2SBS due to an amino acid substitution at position 367, was more active than human N2 on multivalent substrates containing α2,3-linked SIAs, corresponding with the pronounced binding-specificity of avian-like N2 for these receptors. When introduced into human viruses, avian-like N2 gave rise to altered plaque morphology and decreased replication compared to human N2. An opposite replication phenotype was observed when N2 was combined with avian-like HA. Specific bio-layer interferometry assays revealed a clear effect of the 2SBS on the dynamic interaction of virus particles with receptors. The absence or presence of a functional 2SBS affected virion-receptor binding and receptor cleavage required for particle movement on a receptor-coated surface and subsequent NA-dependent self-elution. The contribution of the 2SBS to virus-receptor interactions depended on the receptor-binding properties of HA and the identity of the receptors used. We conclude that the 2SBS is an important and underappreciated determinant of the HA-NA-receptor balance. The rapid loss of a functional 2SBS in pandemic viruses may have served to balance the novel host receptor-repertoire and altered receptor-binding properties of the corresponding HA protein.


id: cord-266078-h53zpjab
title: STING is required for host defense against neuropathological West Nile virus infection
author: McGuckin Wuertz, Kathryn
date: 2019-08-15
txt: ./txt/cord-266078-h53zpjab.txt
cache: ./cache/cord-266078-h53zpjab.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/31415679/
doi: 10.1371/journal.ppat.1007899
summary:
  Recent studies indicate that STimulator of INterferon Gene (STING), canonically known for initiating a type I IFN production and innate immune response to cytosolic DNA, is required for host defense against neurotropic RNA viruses. In vivo, STING-/mice developed an aberrant T cell response in both the spleen and brain during WNV infection that linked with increased and sustained CNS pathology compared to WT mice. In the CNS however, we observed a trend toward increased viral RNA and innate immune gene expression at 8 dpi in WNV-infected STING-/-mice, similar to that observed in BMDM (Fig 3A and 3D) . These observations suggest that STING plays a role in directing or maintaining the T cell response to specific loci within the CNS or that initial viral infection led to increased STING is required for host defense against neuropathological West Nile virus infection recruitment of a localized adaptive immune response that resulted in immunopathology.

abstract:
  West Nile Virus (WNV), an emerging and re-emerging RNA virus, is the leading source of arboviral encephalitic morbidity and mortality in the United States. WNV infections are acutely controlled by innate immunity in peripheral tissues outside of the central nervous system (CNS) but WNV can evade the actions of interferon (IFN) to facilitate CNS invasion, causing encephalitis, encephalomyelitis, and death. Recent studies indicate that STimulator of INterferon Gene (STING), canonically known for initiating a type I IFN production and innate immune response to cytosolic DNA, is required for host defense against neurotropic RNA viruses. We evaluated the role of STING in host defense to control WNV infection and pathology in a murine model of infection. When challenged with WNV, STING knock out (-/-) mice displayed increased morbidity and mortality compared to wild type (WT) mice. Virologic analysis and assessment of STING activation revealed that STING signaling was not required for control of WNV in the spleen nor was WNV sufficient to mediate canonical STING activation in vitro. However, STING-/- mice exhibited a clear trend of increased viral load and virus dissemination in the CNS. We found that STING-/- mice exhibited increased and prolonged neurological signs compared to WT mice. Pathological examination revealed increased lesions, mononuclear cellular infiltration and neuronal death in the CNS of STING-/- mice, with sustained pathology after viral clearance. We found that STING was required in bone marrow derived macrophages for early control of WNV replication and innate immune activation. In vivo, STING-/- mice developed an aberrant T cell response in both the spleen and brain during WNV infection that linked with increased and sustained CNS pathology compared to WT mice. Our findings demonstrate that STING plays a critical role in immune programming for the control of neurotropic WNV infection and CNS disease.


id: cord-267149-5twx9y5c
author: Abraham, Jonathan
date: 2009-04-03
url: https://doi.org/10.1371/journal.ppat.1000358
doi: 10.1371/journal.ppat.1000358
title:
  Host-Species Transferrin Receptor 1 Orthologs Are Cellular Receptors for Nonpathogenic New World Clade B Arenaviruses

abstract:
  The ability of a New World (NW) clade B arenavirus to enter cells using human transferrin receptor 1 (TfR1) strictly correlates with its ability to cause hemorrhagic fever. Amapari (AMAV) and Tacaribe (TCRV), two nonpathogenic NW clade B arenaviruses that do not use human TfR1, are closely related to the NW arenaviruses that cause hemorrhagic fevers. Here we show that pseudotyped viruses bearing the surface glycoprotein (GP) of AMAV or TCRV can infect cells using the TfR1 orthologs of several mammalian species, including those of their respective natural hosts, the small rodent Neacomys spinosus and the fruit bat Artibeus jamaicensis. Mutation of one residue in human TfR1 makes it a functional receptor for TCRV, and mutation of four residues makes it a functional receptor for AMAV. Our data support an in vivo role for TfR1 in the replication of most, if not all, NW clade B arenaviruses, and suggest that with modest changes in their GPs the nonpathogenic arenaviruses could use human TfR1 and emerge as human pathogens.


id: cord-268388-kkhuzf3p
title: Emergence of MERS-CoV in the Middle East: Origins, Transmission, Treatment, and Perspectives
author: Sharif-Yakan, Ahmad
date: 2014-12-04
txt: ./txt/cord-268388-kkhuzf3p.txt
cache: ./cache/cord-268388-kkhuzf3p.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/25474536/
doi: 10.1371/journal.ppat.1004457
abstract: nan
summary:
  Two years have passed since the initial description of the Middle East respiratory syndrome coronavirus (MERS-CoV), yet the epidemic is far from being controlled. First reported in 2012 [1] , Middle East respiratory syndrome coronavirus (MERS-CoV) is a novel coronavirus and the first lineage 2C Betacoronavirus known to infect humans [2] . Middle east respiratory syndrome coronavirus (MERS-CoV) RNA and neutralising antibodies in milk collected according to local customs from dromedary camels, qatar Middle east respiratory syndrome coronavirus (MERS-CoV) in dromedary camels Epidemiological, demographic, and clinical characteristics of 47 cases of middle east respiratory syndrome coronavirus disease from saudi arabia: A descriptive study Clinical features and viral diagnosis of two cases of infection with middle east respiratory syndrome coronavirus: A report of nosocomial transmission Clinical features and virological analysis of a case of middle east respiratory syndrome coronavirus infection Ribavirin and interferon therapy in patients infected with the middle east respiratory syndrome coronavirus: An observational study


id: cord-268677-ytxrrslz
author: Züst, Roland
date: 2013-08-01
txt: ./txt/cord-268677-ytxrrslz.txt
cache: ./cache/cord-268677-ytxrrslz.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1003521
doi: 10.1371/journal.ppat.1003521
summary:
  Here we demonstrate that DENV strains bearing a mutation in the catalytic site of the 29-O-MTase replicated to high titres in cell culture whereas these mutant viruses were highly attenuated in mice and rhesus monkeys. Mice and monkeys infected with the mutant virus developed an immune response that fully protected them from a challenge with wild-type virus. These data suggest that vaccination with the E216A/E217A mutants does not cause ADE during heterologous challenge even though lower neutralizing Ab titers are generated by the mutant strains compared to the wild-type virus. To assess the safety (viremia profile) and efficacy (neutralizing antibody response and protection against challenge) of the 29-O-MTase mutant DENV vaccine approach in an immunologically competent host, three groups of Rhesus monkeys (RM) were immunized with different doses of E217A. These data demonstrate that live, attenuated DENV MTase mutant virus, even when administrated at low dose (1610 3 PFU), can induce protective immunity in non-human primates.

title:
  Rational Design of a Live Attenuated Dengue Vaccine: 2′-O-Methyltransferase Mutants Are Highly Attenuated and Immunogenic in Mice and Macaques

abstract:
  Dengue virus is transmitted by Aedes mosquitoes and infects at least 100 million people every year. Progressive urbanization in Asia and South-Central America and the geographic expansion of Aedes mosquito habitats have accelerated the global spread of dengue, resulting in a continuously increasing number of cases. A cost-effective, safe vaccine conferring protection with ideally a single injection could stop dengue transmission. Current vaccine candidates require several booster injections or do not provide protection against all four serotypes. Here we demonstrate that dengue virus mutants lacking 2′-O-methyltransferase activity are highly sensitive to type I IFN inhibition. The mutant viruses are attenuated in mice and rhesus monkeys and elicit a strong adaptive immune response. Monkeys immunized with a single dose of 2′-O-methyltransferase mutant virus showed 100% sero-conversion even when a dose as low as 1,000 plaque forming units was administrated. Animals were fully protected against a homologous challenge. Furthermore, mosquitoes feeding on blood containing the mutant virus were not infected, whereas those feeding on blood containing wild-type virus were infected and thus able to transmit it. These results show the potential of 2′-O-methyltransferase mutant virus as a safe, rationally designed dengue vaccine that restrains itself due to the increased susceptibility to the host's innate immune response.


id: cord-269466-9hnal9ad
author: Agbeci, Maxime
date: 2013-10-03
txt: ./txt/cord-269466-9hnal9ad.txt
cache: ./cache/cord-269466-9hnal9ad.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/24098128/
doi: 10.1371/journal.ppat.1003683
summary:
  In this study, we used a novel dual gene cassette construct that differentiated primary infected cells from cells infected after virus intercellular movement to show that the early as well as the late secretory pathway, but not endocytosis, was important for TuMV transport. By using a dual cassette of genes encoding fluorescent proteins that can differentiate between primary infected cells and cells infected after intercellular transport, we provide evidence that turnip mosaic virus (TuMV) needs a functional secretory pathway where pre-and post-Golgi trafficking and the actomyosin network are important for its movement. Fig. 2E shows that that there was no significant difference in the ratio of red over green fluorescence during BFA and CMA treatments compared with the no inhibitor treatment (TuMV alone) at 4 dpinf, indicating that viral protein production in the primary infected cells was not affected by the drug treatments.

title:
  Contribution of Host Intracellular Transport Machineries to Intercellular Movement of Turnip Mosaic Virus

abstract:
  The contribution of different host cell transport systems in the intercellular movement of turnip mosaic virus (TuMV) was investigated. To discriminate between primary infections and secondary infections associated with the virus intercellular movement, a gene cassette expressing GFP-HDEL was inserted adjacent to a TuMV infectious cassette expressing 6K(2):mCherry, both within the T-DNA borders of the binary vector pCambia. In this system, both gene cassettes were delivered to the same cell by a single binary vector and primary infection foci emitted green and red fluorescence while secondarily infected cells emitted only red fluorescence. Intercellular movement was measured at 72 hours post infiltration and was estimated to proceed at an average rate of one cell being infected every three hours over an observation period of 17 hours. To determine if the secretory pathway were important for TuMV intercellular movement, chemical and protein inhibitors that blocked both early and late secretory pathways were used. Treatment with Brefeldin A or Concanamycin A or expression of ARF1 or RAB-E1d dominant negative mutants, all of which inhibit pre- or post-Golgi transport, reduced intercellular movement by the virus. These treatments, however, did not inhibit virus replication in primary infected cells. Pharmacological interference assays using Tyrphostin A23 or Wortmannin showed that endocytosis was not important for TuMV intercellular movement. Lack of co-localization by endocytosed FM4-64 and Ara7 (AtRabF2b) with TuMV-induced 6K(2)-tagged vesicles further supported this conclusion. Microfilament depolymerizing drugs and silencing expression of myosin XI-2 gene, but not myosin VIII genes, also inhibited TuMV intercellular movement. Expression of dominant negative myosin mutants confirmed the role played by myosin XI-2 as well as by myosin XI-K in TuMV intercellular movement. Using this dual gene cassette expression system and transport inhibitors, components of the secretory and actomyosin machinery were shown to be important for TuMV intercellular spread.


id: cord-271692-60nlid3c
title: Phylogeny and Origins of Hantaviruses Harbored by Bats, Insectivores, and Rodents
author: Guo, Wen-Ping
date: 2013-02-07
url: https://doi.org/10.1371/journal.ppat.1003159
doi: 10.1371/journal.ppat.1003159
abstract:
  Hantaviruses are among the most important zoonotic pathogens of humans and the subject of heightened global attention. Despite the importance of hantaviruses for public health, there is no consensus on their evolutionary history and especially the frequency of virus-host co-divergence versus cross-species virus transmission. Documenting the extent of hantavirus biodiversity, and particularly their range of mammalian hosts, is critical to resolving this issue. Here, we describe four novel hantaviruses (Huangpi virus, Lianghe virus, Longquan virus, and Yakeshi virus) sampled from bats and shrews in China, and which are distinct from other known hantaviruses. Huangpi virus was found in Pipistrellus abramus, Lianghe virus in Anourosorex squamipes, Longquan virus in Rhinolophus affinis, Rhinolophus sinicus, and Rhinolophus monoceros, and Yakeshi virus in Sorex isodon, respectively. A phylogenetic analysis of the available diversity of hantaviruses reveals the existence of four phylogroups that infect a range of mammalian hosts, as well as the occurrence of ancient reassortment events between the phylogroups. Notably, the phylogenetic histories of the viruses are not always congruent with those of their hosts, suggesting that cross-species transmission has played a major role during hantavirus evolution and at all taxonomic levels, although we also noted some evidence for virus-host co-divergence. Our phylogenetic analysis also suggests that hantaviruses might have first appeared in Chiroptera (bats) or Soricomorpha (moles and shrews), before emerging in rodent species. Overall, these data indicate that bats are likely to be important natural reservoir hosts of hantaviruses.


id: cord-275307-d7htyfcl
author: Gaglia, Marta Maria
date: 2015-12-08
url: https://www.ncbi.nlm.nih.gov/pubmed/26646420/
doi: 10.1371/journal.ppat.1005305
title:
  Transcriptome-Wide Cleavage Site Mapping on Cellular mRNAs Reveals Features Underlying Sequence-Specific Cleavage by the Viral Ribonuclease SOX

abstract:
  Many viruses express factors that reduce host gene expression through widespread degradation of cellular mRNA. An example of this class of proteins is the mRNA-targeting endoribonuclease SOX from the gamma-herpesvirus Kaposi’s sarcoma-associated herpesvirus (KSHV). Previous studies indicated that cleavage of messenger RNAs (mRNA) by SOX occurs at specific locations defined by the sequence of the target RNA, which is at odds with the down-regulation of a large portion of cellular transcripts. In this study, we address this paradox by using high-throughput sequencing of cleavage intermediates combined with a custom bioinformatics-based analysis pipeline to identify SOX cleavage sites across the mRNA transcriptome. These data, coupled with targeted mutagenesis, reveal that while cleavage sites are specific and reproducible, they are defined by a degenerate sequence motif containing a small number of conserved residues rather than a strong consensus sequence. This degenerate element is well represented in both human and KSHV mRNA, and its presence correlates with RNA destabilization by SOX. This represents a new endonuclease targeting strategy, in which use of a degenerate targeting element enables RNA cleavage at specific locations without restricting the range of targets. Furthermore, it shows that strong target selectivity can be achieved without a high degree of sequence specificity.


id: cord-277566-j3ehiwn9
title: Mouse Hepatitis Coronavirus RNA Replication Depends on GBF1-Mediated ARF1 Activation
author: Verheije, Monique H.
date: 2008-06-13
url: https://doi.org/10.1371/journal.ppat.1000088
doi: 10.1371/journal.ppat.1000088
abstract:
  Coronaviruses induce in infected cells the formation of double membrane vesicles, which are the sites of RNA replication. Not much is known about the formation of these vesicles, although recent observations indicate an important role for the endoplasmic reticulum in the formation of the mouse hepatitis coronavirus (MHV) replication complexes (RCs). We now show that MHV replication is sensitive to brefeldin A (BFA). Consistently, expression of a dominant-negative mutant of ARF1, known to mimic the action of the drug, inhibited MHV infection profoundly. Immunofluorescence analysis and quantitative electron microscopy demonstrated that BFA did not block the formation of RCs per se, but rather reduced their number. MHV RNA replication was not sensitive to BFA in MDCK cells, which are known to express the BFA-resistant guanine nucleotide exchange factor GBF1. Accordingly, individual knockdown of the Golgi-resident targets of BFA by transfection of small interfering RNAs (siRNAs) showed that GBF1, but not BIG1 or BIG2, was critically involved in MHV RNA replication. ARF1, the cellular effector of GBF1, also appeared to be involved in MHV replication, as siRNAs targeting this small GTPase inhibited MHV infection significantly. Collectively, our results demonstrate that GBF1-mediated ARF1 activation is required for efficient MHV RNA replication and reveal that the early secretory pathway and MHV replication complex formation are closely connected.


id: cord-278511-je1509ar
title: 5 challenges in understanding the role of the virome in health and disease
author: Wang, David
date: 2020-03-26
txt: ./txt/cord-278511-je1509ar.txt
cache: ./cache/cord-278511-je1509ar.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1008318
doi: 10.1371/journal.ppat.1008318
abstract: nan
summary:
  An even more significant problem is that in most virome studies, more than 50% of the sequences in virus-enriched preparations have no detectable sequence similarity to any known reference sequences; these unalignable sequences are referred to as viral "dark matter" [8] and may include novel, highly divergent viruses that are unrecognizable. To illustrate this point, the United States National Institutes of Health (NIH) virology study sections address only "non-bacteriophage viral genetics, infection and replication, cellular and host responses to viral infections, and mechanisms of viral disease pathogenesis." Thus, there is a great need to bring these disparate communities together in order to collectively attack questions associated with the virome, especially as more complex trans-kingdom interactions are identified linking phages, bacteria, eukaryotic viruses, and eukaryotic cells. With new cell-culture systems and animal models for novel viruses, there will ideally be studies that attribute causal roles for some of the associations.


id: cord-278569-yr06jwm1
title: CD200 Receptor Controls Sex-Specific TLR7 Responses to Viral Infection
author: Karnam, Guruswamy
date: 2012-05-17
url: https://doi.org/10.1371/journal.ppat.1002710
doi: 10.1371/journal.ppat.1002710
abstract:
  Immunological checkpoints, such as the inhibitory CD200 receptor (CD200R), play a dual role in balancing the immune system during microbial infection. On the one hand these inhibitory signals prevent excessive immune mediated pathology but on the other hand they may impair clearance of the pathogen. We studied the influence of the inhibitory CD200-CD200R axis on clearance and pathology in two different virus infection models. We find that lack of CD200R signaling strongly enhances type I interferon (IFN) production and viral clearance and improves the outcome of mouse hepatitis corona virus (MHV) infection, particularly in female mice. MHV clearance is known to be dependent on Toll like receptor 7 (TLR7)-mediated type I IFN production and sex differences in TLR7 responses previously have been reported for humans. We therefore hypothesize that CD200R ligation suppresses TLR7 responses and that release of this inhibition enlarges sex differences in TLR7 signaling. This hypothesis is supported by our findings that in vivo administration of synthetic TLR7 ligand leads to enhanced type I IFN production, particularly in female Cd200(−/−) mice and that CD200R ligation inhibits TLR7 signaling in vitro. In influenza A virus infection we show that viral clearance is determined by sex but not by CD200R signaling. However, absence of CD200R in influenza A virus infection results in enhanced lung neutrophil influx and pathology in females. Thus, CD200-CD200R and sex are host factors that together determine the outcome of viral infection. Our data predict a sex bias in both beneficial and pathological immune responses to virus infection upon therapeutic targeting of CD200-CD200R.


id: cord-280621-tph5n7ak
author: Kim, Yunjeong
date: 2016-03-30
txt: ./txt/cord-280621-tph5n7ak.txt
cache: ./cache/cord-280621-tph5n7ak.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/27027316/
doi: 10.1371/journal.ppat.1005531
summary:
  Shifts in tissue or cell tropism and resulting changes in virulence have also been reported for coronaviruses; porcine respiratory coronavirus causes mild respiratory infection in pigs and presumably arose from transmissible gastroenteritis virus (TGEV), the etiologic agent of gastroenteritis in young pigs with a high fatality, by spontaneous mutations and/or deletions in its genome [9] . Effective treatment intervention for coronavirus infections with an immunopathological component, such as SARS, MERS and FIP, is speculated to involve the judicious use of immunomodulatory agents to enhance protective host immunity and decrease pathological immune responses and antiviral drugs to directly inhibit viral replication. These results on viral titers show that FIPV 3CLpro is a valid target for FIPV antiviral drugs and GC376 can effectively reduce the virus load in the macrophages from the ascites and the omentum of cats with FIP.

title:
  Reversal of the Progression of Fatal Coronavirus Infection in Cats by a Broad-Spectrum Coronavirus Protease Inhibitor

abstract:
  Coronaviruses infect animals and humans causing a wide range of diseases. The diversity of coronaviruses in many mammalian species is contributed by relatively high mutation and recombination rates during replication. This dynamic nature of coronaviruses may facilitate cross-species transmission and shifts in tissue or cell tropism in a host, resulting in substantial change in virulence. Feline enteric coronavirus (FECV) causes inapparent or mild enteritis in cats, but a highly fatal disease, called feline infectious peritonitis (FIP), can arise through mutation of FECV to FIP virus (FIPV). The pathogenesis of FIP is intimately associated with immune responses and involves depletion of T cells, features shared by some other coronaviruses like Severe Acute Respiratory Syndrome Coronavirus. The increasing risks of highly virulent coronavirus infections in humans or animals call for effective antiviral drugs, but no such measures are yet available. Previously, we have reported the inhibitors that target 3C-like protease (3CLpro) with broad-spectrum activity against important human and animal coronaviruses. Here, we evaluated the therapeutic efficacy of our 3CLpro inhibitor in laboratory cats with FIP. Experimental FIP is 100% fatal once certain clinical and laboratory signs become apparent. We found that antiviral treatment led to full recovery of cats when treatment was started at a stage of disease that would be otherwise fatal if left untreated. Antiviral treatment was associated with a rapid improvement in fever, ascites, lymphopenia and gross signs of illness and cats returned to normal health within 20 days or less of treatment. Significant reduction in viral titers was also observed in cats. These results indicate that continuous virus replication is required for progression of immune-mediated inflammatory disease of FIP. These findings may provide important insights into devising therapeutic strategies and selection of antiviral compounds for further development for important coronaviruses in animals and humans.


id: cord-281404-5a8au32c
author: Gastaldello, Stefano
date: 2013-10-10
txt: ./txt/cord-281404-5a8au32c.txt
cache: ./cache/cord-281404-5a8au32c.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1003664
doi: 10.1371/journal.ppat.1003664
summary:
  The large tegument proteins of herpesviruses contain N-terminal cysteine proteases with potent ubiquitin and NEDD8-specific deconjugase activities, but the function of the enzymes during virus replication remains largely unknown. Here we report that induction of the productive virus cycle has no appreciable effects on the global levels of protein ubiquitination but is accompanied by a BPLF1-dependent decrease of cullin neddylation and stabilization of nuclear CRL substrates. The Akata-Bx1 cell line was used to study the contribution of the Ub-and NEDD8-specific deconjugase activities of the EBV large tegument protein BPLF1 to the productive virus cycle. In order to assess whether this regulatory interaction may operate during virus replication, the productive cycle was induced in Akata-Bx1 cells transiently transfected with plasmids expressing Myc-tagged CAND1 or the CAND1 Nterminus that compete for BPLF1 binding to cullins, or, as controls, the CAND1 C-terminus that binds to the opposite end of the cullin scaffold, and the empty vector ( Figure 5A ).

title:
  Caspase-1 Promotes Epstein-Barr Virus Replication by Targeting the Large Tegument Protein Deneddylase to the Nucleus of Productively Infected Cells

abstract:
  The large tegument proteins of herpesviruses contain N-terminal cysteine proteases with potent ubiquitin and NEDD8-specific deconjugase activities, but the function of the enzymes during virus replication remains largely unknown. Using as model BPLF1, the homologue encoded by Epstein-Barr virus (EBV), we found that induction of the productive virus cycle does not affect the total level of ubiquitin-conjugation but is accompanied by a BPLF1-dependent decrease of NEDD8-adducts and accumulation of free NEDD8. Expression of BPLF1 promotes cullin degradation and the stabilization of cullin-RING ligases (CRLs) substrates in the nucleus, while cytoplasmic CRLs and their substrates are not affected. The inactivation of nuclear CRLs is reversed by the N-terminus of CAND1, which inhibits the binding of BPLF1 to cullins and prevents efficient viral DNA replication. Targeting of the deneddylase activity to the nucleus is dependent on processing of the catalytic N-terminus by caspase-1. Inhibition of caspase-1 severely impairs viral DNA synthesis and the release of infectious virus, pointing a previously unrecognized role of the cellular response to danger signals triggered by EBV reactivation in promoting virus replication.


id: cord-286072-kgpvdb42
title: Interplay between SARS-CoV-2 and the type I interferon response
author: Sa Ribero, Margarida
date: 2020-07-29
url: https://doi.org/10.1371/journal.ppat.1008737
doi: 10.1371/journal.ppat.1008737
abstract:
  The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is responsible for the current COVID-19 pandemic. An unbalanced immune response, characterized by a weak production of type I interferons (IFN-Is) and an exacerbated release of proinflammatory cytokines, contributes to the severe forms of the disease. SARS-CoV-2 is genetically related to SARS-CoV and Middle East respiratory syndrome-related coronavirus (MERS-CoV), which caused outbreaks in 2003 and 2013, respectively. Although IFN treatment gave some encouraging results against SARS-CoV and MERS-CoV in animal models, its potential as a therapeutic against COVID-19 awaits validation. Here, we describe our current knowledge of the complex interplay between SARS-CoV-2 infection and the IFN system, highlighting some of the gaps that need to be filled for a better understanding of the underlying molecular mechanisms. In addition to the conserved IFN evasion strategies that are likely shared with SARS-CoV and MERS-CoV, novel counteraction mechanisms are being discovered in SARS-CoV-2–infected cells. Since the last coronavirus epidemic, we have made considerable progress in understanding the IFN-I response, including its spatiotemporal regulation and the prominent role of plasmacytoid dendritic cells (pDCs), which are the main IFN-I–producing cells. While awaiting the results of the many clinical trials that are evaluating the efficacy of IFN-I alone or in combination with antiviral molecules, we discuss the potential benefits of a well-timed IFN-I treatment and propose strategies to boost pDC-mediated IFN responses during the early stages of viral infection.


id: cord-286137-4cbh3u3z
author: Honce, Rebekah
date: 2020-08-13
url: https://www.ncbi.nlm.nih.gov/pubmed/32790755/
doi: 10.1371/journal.ppat.1008711
abstract: nan
title:
  They are what you eat: Shaping of viral populations through nutrition and consequences for virulence


id: cord-287219-qxkwjkif
author: Geisbert, Thomas W.
date: 2008-11-28
txt: ./txt/cord-287219-qxkwjkif.txt
cache: ./cache/cord-287219-qxkwjkif.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1000225
doi: 10.1371/journal.ppat.1000225
summary:
  In order to address this concern, we evaluated the safety of the recombinant VSV vector expressing the Zaire ebolavirus glycoprotein (VSVΔG/ZEBOVGP) in six rhesus macaques infected with simian-human immunodeficiency virus (SHIV). Therefore, we conducted a study to assess the pathogenicity and protective efficacy of the recombinant VSVbased ZEBOV vaccine vector in rhesus macaques that were infected with simian-human immunodeficiency virus (SHIV) which is known to deplete the populations of naive CD4+ T cells, naive CD8+ T cells, and memory CD4+ T cells in these animals [22, 23] . Disease progressed in two of the VSVDG/ZEBOVGP-vaccinated SHIV-infected monkeys (Subject #5 and 6) and both of the placebo control animals with the development of additional evidence of clinical illness including increased levels of serum enzymes associated with liver function, depression, anorexia, and the appearance of macular rashes (Table 3 ).

title:
  Vesicular Stomatitis Virus-Based Ebola Vaccine Is Well-Tolerated and Protects Immunocompromised Nonhuman Primates

abstract:
  Ebola virus (EBOV) is a significant human pathogen that presents a public health concern as an emerging/re-emerging virus and as a potential biological weapon. Substantial progress has been made over the last decade in developing candidate preventive vaccines that can protect nonhuman primates against EBOV. Among these prospects, a vaccine based on recombinant vesicular stomatitis virus (VSV) is particularly robust, as it can also confer protection when administered as a postexposure treatment. A concern that has been raised regarding the replication-competent VSV vectors that express EBOV glycoproteins is how these vectors would be tolerated by individuals with altered or compromised immune systems such as patients infected with HIV. This is especially important as all EBOV outbreaks to date have occurred in areas of Central and Western Africa with high HIV incidence rates in the population. In order to address this concern, we evaluated the safety of the recombinant VSV vector expressing the Zaire ebolavirus glycoprotein (VSVΔG/ZEBOVGP) in six rhesus macaques infected with simian-human immunodeficiency virus (SHIV). All six animals showed no evidence of illness associated with the VSVΔG/ZEBOVGP vaccine, suggesting that this vaccine may be safe in immunocompromised populations. While one goal of the study was to evaluate the safety of the candidate vaccine platform, it was also of interest to determine if altered immune status would affect vaccine efficacy. The vaccine protected 4 of 6 SHIV-infected macaques from death following ZEBOV challenge. Evaluation of CD4+ T cells in all animals showed that the animals that succumbed to lethal ZEBOV challenge had the lowest CD4+ counts, suggesting that CD4+ T cells may play a role in mediating protection against ZEBOV.


id: cord-288231-vg8bwed9
title: The Application of Genomics to Emerging Zoonotic Viral Diseases
author: Haagmans, Bart L.
date: 2009-10-26
url: https://www.ncbi.nlm.nih.gov/pubmed/19855817/
doi: 10.1371/journal.ppat.1000557
abstract:
  Interspecies transmission of pathogens may result in the emergence of new infectious diseases in humans as well as in domestic and wild animals. Genomics tools such as high-throughput sequencing, mRNA expression profiling, and microarray-based analysis of single nucleotide polymorphisms are providing unprecedented ways to analyze the diversity of the genomes of emerging pathogens as well as the molecular basis of the host response to them. By comparing and contrasting the outcomes of an emerging infection with those of closely related pathogens in different but related host species, we can further delineate the various host pathways determining the outcome of zoonotic transmission and adaptation to the newly invaded species. The ultimate challenge is to link pathogen and host genomics data with biological outcomes of zoonotic transmission and to translate the integrated data into novel intervention strategies that eventually will allow the effective control of newly emerging infectious diseases.


id: cord-289879-fh7ic0kp
title: Tmprss2 Is Essential for Influenza H1N1 Virus Pathogenesis in Mice
author: Hatesuer, Bastian
date: 2013-12-05
url: https://doi.org/10.1371/journal.ppat.1003774
doi: 10.1371/journal.ppat.1003774
abstract:
  Annual influenza epidemics and occasional pandemics pose a severe threat to human health. Host cell factors required for viral spread but not for cellular survival are attractive targets for novel approaches to antiviral intervention. The cleavage activation of the influenza virus hemagglutinin (HA) by host cell proteases is essential for viral infectivity. However, it is unknown which proteases activate influenza viruses in mammals. Several candidates have been identified in cell culture studies, leading to the concept that influenza viruses can employ multiple enzymes to ensure their cleavage activation in the host. Here, we show that deletion of a single HA-activating protease gene, Tmprss2, in mice inhibits spread of mono-basic H1N1 influenza viruses, including the pandemic 2009 swine influenza virus. Lung pathology was strongly reduced and mutant mice were protected from weight loss, death and impairment of lung function. Also, after infection with mono-basic H3N2 influenza A virus body weight loss and survival was less severe in Tmprss2 mutant compared to wild type mice. As expected, Tmprss2-deficient mice were not protected from viral spread and pathology after infection with multi-basic H7N7 influenza A virus. In conclusion, these results identify TMPRSS2 as a host cell factor essential for viral spread and pathogenesis of mono-basic H1N1 and H3N2 influenza A viruses.


id: cord-290253-hxxizipk
title: Changes in temperature alter the potential outcomes of virus host shifts
author: Roberts, Katherine E.
date: 2018-10-19
url: https://doi.org/10.1371/journal.ppat.1007185
doi: 10.1371/journal.ppat.1007185
abstract:
  Host shifts–where a pathogen jumps between different host species–are an important source of emerging infectious disease. With on-going climate change there is an increasing need to understand the effect changes in temperature may have on emerging infectious disease. We investigated whether species’ susceptibilities change with temperature and ask if susceptibility is greatest at different temperatures in different species. We infected 45 species of Drosophilidae with an RNA virus and measured how viral load changes with temperature. We found the host phylogeny explained a large proportion of the variation in viral load at each temperature, with strong phylogenetic correlations between viral loads across temperature. The variance in viral load increased with temperature, while the mean viral load did not. This suggests that as temperature increases the most susceptible species become more susceptible, and the least susceptible less so. We found no significant relationship between a species’ susceptibility across temperatures, and proxies for thermal optima (critical thermal maximum and minimum or basal metabolic rate). These results suggest that whilst the rank order of species susceptibilities may remain the same with changes in temperature, some species may become more susceptible to a novel pathogen, and others less so.


id: cord-290617-45be6gxe
title: Footprint of the host restriction factors APOBEC3 on the genome of human viruses
author: Poulain, Florian
date: 2020-08-14
txt: ./txt/cord-290617-45be6gxe.txt
cache: ./cache/cord-290617-45be6gxe.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/32797103/
doi: 10.1371/journal.ppat.1008718
summary:
  Certain viruses actively encode viral proteins antagonizing the APOBEC3s, others passively face the APOBEC3 selection pressure thanks to a depleted genome for APOBEC3-targeted motifs. By breaking down the human viruses into their respective Baltimore''s group (S3 Fig), we observed that NTC depletion is not present in reverse transcribing nor in negative sense single strand RNA viruses. We also observed a mild general NCC depletion in single strand DNA and double strand RNA viruses, justifying further investigation for a possible A3G-induced footprint (S1 Fig). In order to identify A3-footprinted viruses, we detailed the NTC and NNGANN ratios for 870 human viral species (Fig 4A) . B. The observed/expected ratios of TC dinucleotide at various codon positions and on both strands (i.e. NNTCNN, TCN, NTC, GAN, NGA and NNGANN) were calculated for the putative A3-footprinted viral species and depicted by a heatmap.

abstract:
  APOBEC3 enzymes are innate immune effectors that introduce mutations into viral genomes. These enzymes are cytidine deaminases which transform cytosine into uracil. They preferentially mutate cytidine preceded by thymidine making the 5’TC motif their favored target. Viruses have evolved different strategies to evade APOBEC3 restriction. Certain viruses actively encode viral proteins antagonizing the APOBEC3s, others passively face the APOBEC3 selection pressure thanks to a depleted genome for APOBEC3-targeted motifs. Hence, the APOBEC3s left on the genome of certain viruses an evolutionary footprint. The aim of our study is the identification of these viruses having a genome shaped by the APOBEC3s. We analyzed the genome of 33,400 human viruses for the depletion of APOBEC3-favored motifs. We demonstrate that the APOBEC3 selection pressure impacts at least 22% of all currently annotated human viral species. The papillomaviridae and polyomaviridae are the most intensively footprinted families; evidencing a selection pressure acting genome-wide and on both strands. Members of the parvoviridae family are differentially targeted in term of both magnitude and localization of the footprint. Interestingly, a massive APOBEC3 footprint is present on both strands of the B19 erythroparvovirus; making this viral genome one of the most cleaned sequences for APOBEC3-favored motifs. We also identified the endemic coronaviridae as significantly footprinted. Interestingly, no such footprint has been detected on the zoonotic MERS-CoV, SARS-CoV-1 and SARS-CoV-2 coronaviruses. In addition to viruses that are footprinted genome-wide, certain viruses are footprinted only on very short sections of their genome. That is the case for the gamma-herpesviridae and adenoviridae where the footprint is localized on the lytic origins of replication. A mild footprint can also be detected on the negative strand of the reverse transcribing HIV-1, HIV-2, HTLV-1 and HBV viruses. Together, our data illustrate the extent of the APOBEC3 selection pressure on the human viruses and identify new putatively APOBEC3-targeted viruses.


id: cord-292424-daj4zcm1
author: Shang, Jian
date: 2020-03-09
url: https://www.ncbi.nlm.nih.gov/pubmed/32150576/
doi: 10.1371/journal.ppat.1008392
title:
  Structure of mouse coronavirus spike protein complexed with receptor reveals mechanism for viral entry

abstract:
  Coronaviruses recognize a variety of receptors using different domains of their envelope-anchored spike protein. How these diverse receptor recognition patterns affect viral entry is unknown. Mouse hepatitis coronavirus (MHV) is the only known coronavirus that uses the N-terminal domain (NTD) of its spike to recognize a protein receptor, CEACAM1a. Here we determined the cryo-EM structure of MHV spike complexed with mouse CEACAM1a. The trimeric spike contains three receptor-binding S1 heads sitting on top of a trimeric membrane-fusion S2 stalk. Three receptor molecules bind to the sides of the spike trimer, where three NTDs are located. Receptor binding induces structural changes in the spike, weakening the interactions between S1 and S2. Using protease sensitivity and negative-stain EM analyses, we further showed that after protease treatment of the spike, receptor binding facilitated the dissociation of S1 from S2, allowing S2 to transition from pre-fusion to post-fusion conformation. Together these results reveal a new role of receptor binding in MHV entry: in addition to its well-characterized role in viral attachment to host cells, receptor binding also induces the conformational change of the spike and hence the fusion of viral and host membranes. Our study provides new mechanistic insight into coronavirus entry and highlights the diverse entry mechanisms used by different viruses.


id: cord-293164-i50cgkvq
title: Characterization of Uncultivable Bat Influenza Virus Using a Replicative Synthetic Virus
author: Zhou, Bin
date: 2014-10-02
url: https://doi.org/10.1371/journal.ppat.1004420
doi: 10.1371/journal.ppat.1004420
abstract:
  Bats harbor many viruses, which are periodically transmitted to humans resulting in outbreaks of disease (e.g., Ebola, SARS-CoV). Recently, influenza virus-like sequences were identified in bats; however, the viruses could not be cultured. This discovery aroused great interest in understanding the evolutionary history and pandemic potential of bat-influenza. Using synthetic genomics, we were unable to rescue the wild type bat virus, but could rescue a modified bat-influenza virus that had the HA and NA coding regions replaced with those of A/PR/8/1934 (H1N1). This modified bat-influenza virus replicated efficiently in vitro and in mice, resulting in severe disease. Additional studies using a bat-influenza virus that had the HA and NA of A/swine/Texas/4199-2/1998 (H3N2) showed that the PR8 HA and NA contributed to the pathogenicity in mice. Unlike other influenza viruses, engineering truncations hypothesized to reduce interferon antagonism into the NS1 protein didn't attenuate bat-influenza. In contrast, substitution of a putative virulence mutation from the bat-influenza PB2 significantly attenuated the virus in mice and introduction of a putative virulence mutation increased its pathogenicity. Mini-genome replication studies and virus reassortment experiments demonstrated that bat-influenza has very limited genetic and protein compatibility with Type A or Type B influenza viruses, yet it readily reassorts with another divergent bat-influenza virus, suggesting that the bat-influenza lineage may represent a new Genus/Species within the Orthomyxoviridae family. Collectively, our data indicate that the bat-influenza viruses recently identified are authentic viruses that pose little, if any, pandemic threat to humans; however, they provide new insights into the evolution and basic biology of influenza viruses.


id: cord-294342-7ycgd0h7
title: Induction of Cell-Cell Fusion by Ebola Virus Glycoprotein: Low pH Is Not a Trigger
author: Markosyan, Ruben M.
date: 2016-01-05
url: https://doi.org/10.1371/journal.ppat.1005373
doi: 10.1371/journal.ppat.1005373
abstract:
  Ebola virus (EBOV) is a highly pathogenic filovirus that causes hemorrhagic fever in humans and animals. Currently, how EBOV fuses its envelope membrane within an endosomal membrane to cause infection is poorly understood. We successfully measure cell-cell fusion mediated by the EBOV fusion protein, GP, assayed by the transfer of both cytoplasmic and membrane dyes. A small molecule fusion inhibitor, a neutralizing antibody, as well as mutations in EBOV GP known to reduce viral infection, all greatly reduce fusion. By monitoring redistribution of small aqueous dyes between cells and by electrical capacitance measurements, we discovered that EBOV GP-mediated fusion pores do not readily enlarge—a marked difference from the behavior of other viral fusion proteins. EBOV GP must be cleaved by late endosome-resident cathepsins B or L in order to become fusion-competent. Cleavage of cell surface-expressed GP appears to occur in endosomes, as evidenced by the fusion block imposed by cathepsin inhibitors, agents that raise endosomal pH, or an inhibitor of anterograde trafficking. Treating effector cells with a recombinant soluble cathepsin B or thermolysin, which cleaves GP into an active form, increases the extent of fusion, suggesting that a fraction of surface-expressed GP is not cleaved. Whereas the rate of fusion is increased by a brief exposure to acidic pH, fusion does occur at neutral pH. Importantly, the extent of fusion is independent of external pH in experiments in which cathepsin activity is blocked and EBOV GP is cleaved by thermolysin. These results imply that low pH promotes fusion through the well-known pH-dependent activity of cathepsins; fusion induced by cleaved EBOV GP is a process that is fundamentally independent of pH. The cell-cell fusion system has revealed some previously unappreciated features of EBOV entry, which could not be readily elucidated in the context of endosomal entry.


id: cord-297702-vxcj25sn
author: Chen, Yuxin
date: 2020-09-10
txt: ./txt/cord-297702-vxcj25sn.txt
cache: ./cache/cord-297702-vxcj25sn.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1008796
doi: 10.1371/journal.ppat.1008796
summary:
  We continuously monitored the serum IgM and IgG responses specific to four SARS-CoV-2 related antigens, including the nucleoprotein (NP), receptor binding domain (RBD), S1 protein, and ectodomain (ECD) of the spike protein among non-severe and severe COVID-19 patients for seven weeks since disease onset. In this retrospective study, we successively monitored the serum IgM and IgG responses specific to four SARS-CoV-2 related antigens, including the NP protein, RBD protein, S1 protein, and ECD protein in 19 non-severe and 7 severe COVID-19 patients during the disease progression. The severe patients and non-severe patients had comparable reduced fold of IgM, IgG, and IgA binding titer specific to RBD, ECD, S1, and NP protein and neutralization activities. Furthermore, 80.7% of the convalescent sea from COVID-19 patients displayed varying levels of neutralization activities against SARS-CoV-2, which correlated with S1-specific and ECD-specific IgA responses in non-severe patients.

title:
  A comprehensive, longitudinal analysis of humoral responses specific to four recombinant antigens of SARS-CoV-2 in severe and non-severe COVID-19 patients

abstract:
  There is an urgent need for effective treatment and preventive vaccine to contain this devastating global pandemic, which requires a comprehensive understanding of humoral responses specific to SARS-CoV-2 during the disease progression and convalescent phase of COVID-19 patients. We continuously monitored the serum IgM and IgG responses specific to four SARS-CoV-2 related antigens, including the nucleoprotein (NP), receptor binding domain (RBD), S1 protein, and ectodomain (ECD) of the spike protein among non-severe and severe COVID-19 patients for seven weeks since disease onset. Most patients generated humoral responses against NP and spike protein-related antigens but with their distinct kinetics profiles. Combined detection of NP and ECD antigens as detecting antigen synergistically improved the sensitivity of the serological assay, compared to that of using NP or RBD as detection antigen. 80.7% of convalescent sera from COVID-19 patients revealed that the varying extents of neutralization activities against SARS-CoV-2. S1-specific and ECD-specific IgA responses were strongly correlated with the neutralization activities in non-severe patients, but not in severe patients. Moreover, the neutralizing activities of the convalescent sera were shown to significantly decline during the period between 21 days to 28 days after hospital discharge, accompanied by a substantial drop in RBD-specific IgA response. Our data provide evidence that are crucial for serological testing, antibody-based intervention, and vaccine design of COVID-19.


id: cord-302340-pw2xqhwa
title: IFITM3 Inhibits Influenza A Virus Infection by Preventing Cytosolic Entry
author: Feeley, Eric M.
date: 2011-10-27
url: https://www.ncbi.nlm.nih.gov/pubmed/22046135/
doi: 10.1371/journal.ppat.1002337
abstract:
  To replicate, viruses must gain access to the host cell's resources. Interferon (IFN) regulates the actions of a large complement of interferon effector genes (IEGs) that prevent viral replication. The interferon inducible transmembrane protein family members, IFITM1, 2 and 3, are IEGs required for inhibition of influenza A virus, dengue virus, and West Nile virus replication in vitro. Here we report that IFN prevents emergence of viral genomes from the endosomal pathway, and that IFITM3 is both necessary and sufficient for this function. Notably, viral pseudoparticles were inhibited from transferring their contents into the host cell cytosol by IFN, and IFITM3 was required and sufficient for this action. We further demonstrate that IFN expands Rab7 and LAMP1-containing structures, and that IFITM3 overexpression is sufficient for this phenotype. Moreover, IFITM3 partially resides in late endosomal and lysosomal structures, placing it in the path of invading viruses. Collectively our data are consistent with the prediction that viruses that fuse in the late endosomes or lysosomes are vulnerable to IFITM3's actions, while viruses that enter at the cell surface or in the early endosomes may avoid inhibition. Multiple viruses enter host cells through the late endocytic pathway, and many of these invaders are attenuated by IFN. Therefore these findings are likely to have significance for the intrinsic immune system's neutralization of a diverse array of threats.


id: cord-303403-9th2jiq6
author: Qing, Jie
date: 2014-10-02
url: https://www.ncbi.nlm.nih.gov/pubmed/25275585/
doi: 10.1371/journal.ppat.1004422
title:
  Cyclophilin A Associates with Enterovirus-71 Virus Capsid and Plays an Essential Role in Viral Infection as an Uncoating Regulator

abstract:
  Viruses utilize host factors for their efficient proliferation. By evaluating the inhibitory effects of compounds in our library, we identified inhibitors of cyclophilin A (CypA), a known immunosuppressor with peptidyl-prolyl cis-trans isomerase activity, can significantly attenuate EV71 proliferation. We demonstrated that CypA played an essential role in EV71 entry and that the RNA interference-mediated reduction of endogenous CypA expression led to decreased EV71 multiplication. We further revealed that CypA directly interacted with and modified the conformation of H-I loop of the VP1 protein in EV71 capsid, and thus regulated the uncoating process of EV71 entry step in a pH-dependent manner. Our results aid in the understanding of how host factors influence EV71 life cycle and provide new potential targets for developing antiviral agents against EV71 infection.


id: cord-304747-ojyxs3cp
title: Identification of a Novel Polyomavirus from Patients with Acute Respiratory Tract Infections
author: Gaynor, Anne M
date: 2007-05-04
url: https://www.ncbi.nlm.nih.gov/pubmed/17480120/
doi: 10.1371/journal.ppat.0030064
abstract:
  We report the identification of a novel polyomavirus present in respiratory secretions from human patients with symptoms of acute respiratory tract infection. The virus was initially detected in a nasopharyngeal aspirate from a 3-year-old child from Australia diagnosed with pneumonia. A random library was generated from nucleic acids extracted from the nasopharyngeal aspirate and analyzed by high throughput DNA sequencing. Multiple DNA fragments were cloned that possessed limited homology to known polyomaviruses. We subsequently sequenced the entire virus genome of 5,229 bp, henceforth referred to as WU virus, and found it to have genomic features characteristic of the family Polyomaviridae. The genome was predicted to encode small T antigen, large T antigen, and three capsid proteins: VP1, VP2, and VP3. Phylogenetic analysis clearly revealed that the WU virus was divergent from all known polyomaviruses. Screening of 2,135 patients with acute respiratory tract infections in Brisbane, Queensland, Australia, and St. Louis, Missouri, United States, using WU virus–specific PCR primers resulted in the detection of 43 additional specimens that contained WU virus. The presence of multiple instances of the virus in two continents suggests that this virus is geographically widespread in the human population and raises the possibility that the WU virus may be a human pathogen.


id: cord-304815-3datxv8j
title: The Scientific Response to a Pandemic
author: Gronvall, Gigi Kwik
date: 2006-02-24
txt: ./txt/cord-304815-3datxv8j.txt
cache: ./cache/cord-304815-3datxv8j.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/16738708/
doi: 10.1371/journal.ppat.0020009
abstract: nan
summary:
  More than 150 scientists and public health practitioners from 25 countries gathered in Lyon, France, to hear speakers from the WHO, the European Commission, scientific journals, Interpol, and public health networks-many of the institutions and individuals who will likely play key roles in the global response to the next pandemic. By discussing the biosafety and biosecurity challenges presented by past epidemics such as SARS, participants recognized the importance of scientific and public health collaboration in combating disease-and the need to plan. Researchers will need to share biological samples between laboratories, sometimes internationally; decision makers and journalists will want the latest information, which may not be peer reviewed; and researchers will risk contracting the disease they research, which could then spread outside of the laboratory. Scientists need access to samples from patients and laboratories in order to conduct research and public health surveillance, as well as to develop diagnostic tests.


id: cord-306624-1mjmttec
title: A Capsid-Encoded PPxY-Motif Facilitates Adenovirus Entry
author: Wodrich, Harald
date: 2010-03-19
url: https://www.ncbi.nlm.nih.gov/pubmed/20333243/
doi: 10.1371/journal.ppat.1000808
abstract:
  Viruses use cellular machinery to enter and infect cells. In this study we address the cell entry mechanisms of nonenveloped adenoviruses (Ads). We show that protein VI, an internal capsid protein, is rapidly exposed after cell surface attachment and internalization and remains partially associated with the capsid during intracellular transport. We found that a PPxY motif within protein VI recruits Nedd4 E3 ubiquitin ligases to bind and ubiquitylate protein VI. We further show that this PPxY motif is involved in rapid, microtubule-dependent intracellular movement of protein VI. Ads with a mutated PPxY motif can efficiently escape endosomes but are defective in microtubule-dependent trafficking toward the nucleus. Likewise, depletion of Nedd4 ligases attenuates nuclear accumulation of incoming Ad particles and infection. Our data provide the first evidence that virus-encoded PPxY motifs are required during virus entry, which may be of significance for several other pathogens.


id: cord-311383-1aqt65cc
title: The SARS-Unique Domain (SUD) of SARS Coronavirus Contains Two Macrodomains That Bind G-Quadruplexes
author: Tan, Jinzhi
date: 2009-05-15
url: https://www.ncbi.nlm.nih.gov/pubmed/19436709/
doi: 10.1371/journal.ppat.1000428
abstract:
  Since the outbreak of severe acute respiratory syndrome (SARS) in 2003, the three-dimensional structures of several of the replicase/transcriptase components of SARS coronavirus (SARS-CoV), the non-structural proteins (Nsps), have been determined. However, within the large Nsp3 (1922 amino-acid residues), the structure and function of the so-called SARS-unique domain (SUD) have remained elusive. SUD occurs only in SARS-CoV and the highly related viruses found in certain bats, but is absent from all other coronaviruses. Therefore, it has been speculated that it may be involved in the extreme pathogenicity of SARS-CoV, compared to other coronaviruses, most of which cause only mild infections in humans. In order to help elucidate the function of the SUD, we have determined crystal structures of fragment 389–652 (“SUD(core)”) of Nsp3, which comprises 264 of the 338 residues of the domain. Both the monoclinic and triclinic crystal forms (2.2 and 2.8 Å resolution, respectively) revealed that SUD(core) forms a homodimer. Each monomer consists of two subdomains, SUD-N and SUD-M, with a macrodomain fold similar to the SARS-CoV X-domain. However, in contrast to the latter, SUD fails to bind ADP-ribose, as determined by zone-interference gel electrophoresis. Instead, the entire SUD(core) as well as its individual subdomains interact with oligonucleotides known to form G-quadruplexes. This includes oligodeoxy- as well as oligoribonucleotides. Mutations of selected lysine residues on the surface of the SUD-N subdomain lead to reduction of G-quadruplex binding, whereas mutations in the SUD-M subdomain abolish it. As there is no evidence for Nsp3 entering the nucleus of the host cell, the SARS-CoV genomic RNA or host-cell mRNA containing long G-stretches may be targets of SUD. The SARS-CoV genome is devoid of G-stretches longer than 5–6 nucleotides, but more extended G-stretches are found in the 3′-nontranslated regions of mRNAs coding for certain host-cell proteins involved in apoptosis or signal transduction, and have been shown to bind to SUD in vitro. Therefore, SUD may be involved in controlling the host cell's response to the viral infection. Possible interference with poly(ADP-ribose) polymerase-like domains is also discussed.


id: cord-312743-9e4yufo5
author: Breiman, Adrien
date: 2020-05-21
txt: ./txt/cord-312743-9e4yufo5.txt
cache: ./cache/cord-312743-9e4yufo5.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/32437478/
doi: 10.1371/journal.ppat.1008556
abstract: nan
summary:
  These observations suggested that, when produced in cells that express the A or B blood group enzymes, infectious SARS virions are decorated by the corresponding glycan antigens and that the presence of anti-A and anti-B antibodies in blood group O individuals could prevent infection by blocking virus attachment and entry. We therefore hypothesize that as they are produced in cells coexpressing the ACE2 receptor and either the αGal, NeuGc, or A/B blood group antigens, both SARS-CoV and SARS-CoV2 harbor the corresponding glycan epitopes. Likewise, impairment of transmission by the anti-blood group antibodies may not work to its full potential because of their variable titers in the population and of the high affinity of the SARS-CoV2 for ACE2 [18] , rendering its neutralization more difficult.

title:
  Harnessing the natural anti-glycan immune response to limit the transmission of enveloped viruses such as SARS-CoV-2


id: cord-314451-mqnqjn0c
title: A Mouse-Adapted SARS-Coronavirus Causes Disease and Mortality in BALB/c Mice
author: Roberts, Anjeanette
date: 2007-01-12
txt: ./txt/cord-314451-mqnqjn0c.txt
cache: ./cache/cord-314451-mqnqjn0c.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/17222058/
doi: 10.1371/journal.ppat.0030005
summary:
  To generate a model that satisfies these criteria, we have serially passaged SARS-CoV in the respiratory tract of young BALB/c mice, resulting in a lethal virus that causes dosedependent weight loss and mortality associated with higher viral titers in the respiratory tract than are seen with the wildtype virus and with histopathologic findings of severe pulmonary disease. Northern blot analysis of RNA from infected Vero E6 cells indicated that genomic vRNA and viral mRNA and all eight sub-genomic mRNAs were present in similar ratios for the recombinant viruses and MA15 virus as for SARS-CoV (Urbani) ( Figure 2A ). In order to evaluate whether changes in tissue tropism or levels of viral replication could contribute to the lethal phenotype of the MA15 virus, viral titers in lungs, spleen, liver, and brain of BALB/c mice were determined at various time points following intranasal inoculation with SARS-CoV (Urbani) or MA15.

abstract:
  No single animal model for severe acute respiratory syndrome (SARS) reproduces all aspects of the human disease. Young inbred mice support SARS-coronavirus (SARS-CoV) replication in the respiratory tract and are available in sufficient numbers for statistical evaluation. They are relatively inexpensive and easily accessible, but their use in SARS research is limited because they do not develop illness following infection. Older (12- to 14-mo-old) BALB/c mice develop clinical illness and pneumonitis, but they can be hard to procure, and immune senescence complicates pathogenesis studies. We adapted the SARS-CoV (Urbani strain) by serial passage in the respiratory tract of young BALB/c mice. Fifteen passages resulted in a virus (MA15) that is lethal for mice following intranasal inoculation. Lethality is preceded by rapid and high titer viral replication in lungs, viremia, and dissemination of virus to extrapulmonary sites accompanied by lymphopenia, neutrophilia, and pathological changes in the lungs. Abundant viral antigen is extensively distributed in bronchial epithelial cells and alveolar pneumocytes, and necrotic cellular debris is present in airways and alveoli, with only mild and focal pneumonitis. These observations suggest that mice infected with MA15 die from an overwhelming viral infection with extensive, virally mediated destruction of pneumocytes and ciliated epithelial cells. The MA15 virus has six coding mutations associated with adaptation and increased virulence; when introduced into a recombinant SARS-CoV, these mutations result in a highly virulent and lethal virus (rMA15), duplicating the phenotype of the biologically derived MA15 virus. Intranasal inoculation with MA15 reproduces many aspects of disease seen in severe human cases of SARS. The availability of the MA15 virus will enhance the use of the mouse model for SARS because infection with MA15 causes morbidity, mortality, and pulmonary pathology. This virus will be of value as a stringent challenge in evaluation of the efficacy of vaccines and antivirals.


id: cord-315316-w7cn9iqp
author: Earnest, James T.
date: 2017-07-31
url: https://www.ncbi.nlm.nih.gov/pubmed/28759649/
doi: 10.1371/journal.ppat.1006546
title:
  The tetraspanin CD9 facilitates MERS-coronavirus entry by scaffolding host cell receptors and proteases

abstract:
  Infection by enveloped coronaviruses (CoVs) initiates with viral spike (S) proteins binding to cellular receptors, and is followed by proteolytic cleavage of receptor-bound S proteins, which prompts S protein-mediated virus-cell membrane fusion. Infection therefore requires close proximity of receptors and proteases. We considered whether tetraspanins, scaffolding proteins known to facilitate CoV infections, hold receptors and proteases together on cell membranes. Using knockout cell lines, we found that the tetraspanin CD9, but not the tetraspanin CD81, formed cell-surface complexes of dipeptidyl peptidase 4 (DPP4), the MERS-CoV receptor, and the type II transmembrane serine protease (TTSP) member TMPRSS2, a CoV-activating protease. This CD9-facilitated condensation of receptors and proteases allowed MERS-CoV pseudoviruses to enter cells rapidly and efficiently. Without CD9, MERS-CoV viruses were not activated by TTSPs, and they trafficked into endosomes to be cleaved much later and less efficiently by cathepsins. Thus, we identified DPP4:CD9:TTSP as the protein complexes necessary for early, efficient MERS-CoV entry. To evaluate the importance of these complexes in an in vivo CoV infection model, we used recombinant Adenovirus 5 (rAd5) vectors to express human DPP4 in mouse lungs, thereby sensitizing the animals to MERS-CoV infection. When the rAd5-hDPP4 vectors co-expressed small RNAs silencing Cd9 or Tmprss2, the animals were significantly less susceptible, indicating that CD9 and TMPRSS2 facilitated robust in vivo MERS-CoV infection of mouse lungs. Furthermore, the S proteins of virulent mouse-adapted MERS-CoVs acquired a CD9-dependent cell entry character, suggesting that CD9 is a selective agent in the evolution of CoV virulence.


id: cord-316584-b1aa1lri
author: Seiradake, Elena
date: 2009-01-02
url: https://www.ncbi.nlm.nih.gov/pubmed/19119424/
doi: 10.1371/journal.ppat.1000277
title:
  The Cell Adhesion Molecule “CAR” and Sialic Acid on Human Erythrocytes Influence Adenovirus In Vivo Biodistribution

abstract:
  Although it has been known for 50 years that adenoviruses (Ads) interact with erythrocytes ex vivo, the molecular and structural basis for this interaction, which has been serendipitously exploited for diagnostic tests, is unknown. In this study, we characterized the interaction between erythrocytes and unrelated Ad serotypes, human 5 (HAd5) and 37 (HAd37), and canine 2 (CAV-2). While these serotypes agglutinate human erythrocytes, they use different receptors, have different tropisms and/or infect different species. Using molecular, biochemical, structural and transgenic animal-based analyses, we found that the primary erythrocyte interaction domain for HAd37 is its sialic acid binding site, while CAV-2 binding depends on at least three factors: electrostatic interactions, sialic acid binding and, unexpectedly, binding to the coxsackievirus and adenovirus receptor (CAR) on human erythrocytes. We show that the presence of CAR on erythrocytes leads to prolonged in vivo blood half-life and significantly reduced liver infection when a CAR-tropic Ad is injected intravenously. This study provides i) a molecular and structural rationale for Ad–erythrocyte interactions, ii) a basis to improve vector-mediated gene transfer and iii) a mechanism that may explain the biodistribution and pathogenic inconsistencies found between human and animal models.


id: cord-321874-eenaixp0
title: Adaptive Gene Amplification As an Intermediate Step in the Expansion of Virus Host Range
author: Brennan, Greg
date: 2014-03-13
url: https://www.ncbi.nlm.nih.gov/pubmed/24626510/
doi: 10.1371/journal.ppat.1004002
abstract:
  The majority of recently emerging infectious diseases in humans is due to cross-species pathogen transmissions from animals. To establish a productive infection in new host species, viruses must overcome barriers to replication mediated by diverse and rapidly evolving host restriction factors such as protein kinase R (PKR). Many viral antagonists of these restriction factors are species specific. For example, the rhesus cytomegalovirus PKR antagonist, RhTRS1, inhibits PKR in some African green monkey (AGM) cells, but does not inhibit human or rhesus macaque PKR. To model the evolutionary changes necessary for cross-species transmission, we generated a recombinant vaccinia virus that expresses RhTRS1 in a strain that lacks PKR inhibitors E3L and K3L (VVΔEΔK+RhTRS1). Serially passaging VVΔEΔK+RhTRS1 in minimally-permissive AGM cells increased viral replication 10- to 100-fold. Notably, adaptation in these AGM cells also improved virus replication 1000- to 10,000-fold in human and rhesus cells. Genetic analyses including deep sequencing revealed amplification of the rhtrs1 locus in the adapted viruses. Supplying additional rhtrs1 in trans confirmed that amplification alone was sufficient to improve VVΔEΔK+RhTRS1 replication. Viruses with amplified rhtrs1 completely blocked AGM PKR, but only partially blocked human PKR, consistent with the replication properties of these viruses in AGM and human cells. Finally, in contrast to AGM-adapted viruses, which could be serially propagated in human cells, VVΔEΔK+RhTRS1 yielded no progeny virus after only three passages in human cells. Thus, rhtrs1 amplification in a minimally permissive intermediate host was a necessary step, enabling expansion of the virus range to previously nonpermissive hosts. These data support the hypothesis that amplification of a weak viral antagonist may be a general evolutionary mechanism to permit replication in otherwise resistant host species, providing a molecular foothold that could enable further adaptations necessary for efficient replication in the new host.


id: cord-321957-ybtk9cp1
author: Carey, Brian D.
date: 2020-03-09
url: https://www.ncbi.nlm.nih.gov/pubmed/32150585/
doi: 10.1371/journal.ppat.1008282
title:
  Protein Kinase C subtype δ interacts with Venezuelan equine encephalitis virus capsid protein and regulates viral RNA binding through modulation of capsid phosphorylation

abstract:
  Protein phosphorylation plays an important role during the life cycle of many viruses. Venezuelan equine encephalitis virus (VEEV) capsid protein has recently been shown to be phosphorylated at four residues. Here those studies are extended to determine the kinase responsible for phosphorylation and the importance of capsid phosphorylation during the viral life cycle. Phosphorylation site prediction software suggests that Protein Kinase C (PKC) is responsible for phosphorylation of VEEV capsid. VEEV capsid co-immunoprecipitated with PKCδ, but not other PKC isoforms and siRNA knockdown of PKCδ caused a decrease in viral replication. Furthermore, knockdown of PKCδ by siRNA decreased capsid phosphorylation. A virus with capsid phosphorylation sites mutated to alanine (VEEV CPD) displayed a lower genomic copy to pfu ratio than the parental virus; suggesting more efficient viral assembly and more infectious particles being released. RNA:capsid binding was significantly increased in the mutant virus, confirming these results. Finally, VEEV CPD is attenuated in a mouse model of infection, with mice showing increased survival and decreased clinical signs as compared to mice infected with the parental virus. Collectively our data support a model in which PKCδ mediated capsid phosphorylation regulates viral RNA binding and assembly, significantly impacting viral pathogenesis.


id: cord-323568-s0wmll4q
author: Shang, Jian
date: 2018-04-23
url: https://doi.org/10.1371/journal.ppat.1007009
doi: 10.1371/journal.ppat.1007009
title:
  Cryo-EM structure of infectious bronchitis coronavirus spike protein reveals structural and functional evolution of coronavirus spike proteins

abstract:
  As cell-invading molecular machinery, coronavirus spike proteins pose an evolutionary conundrum due to their high divergence. In this study, we determined the cryo-EM structure of avian infectious bronchitis coronavirus (IBV) spike protein from the γ-genus. The trimeric IBV spike ectodomain contains three receptor-binding S1 heads and a trimeric membrane-fusion S2 stalk. While IBV S2 is structurally similar to those from the other genera, IBV S1 possesses structural features that are unique to different other genera, thereby bridging these diverse spikes into an evolutionary spectrum. Specifically, among different genera, the two domains of S1, the N-terminal domain (S1-NTD) and C-terminal domain (S1-CTD), diverge from simpler tertiary structures and quaternary packing to more complex ones, leading to different functions of the spikes in receptor usage and membrane fusion. Based on the above structural and functional comparisons, we propose that the evolutionary spectrum of coronavirus spikes follows the order of α-, δ-, γ-, and β-genus. This study has provided insight into the evolutionary relationships among coronavirus spikes and deepened our understanding of their structural and functional diversity.


id: cord-324928-cpryxa6p
title: Cross-utilisation of template RNAs by alphavirus replicases
author: Lello, Laura Sandra
date: 2020-09-04
txt: ./txt/cord-324928-cpryxa6p.txt
cache: ./cache/cord-324928-cpryxa6p.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/32886709/
doi: 10.1371/journal.ppat.1008825
summary:
  The differences between outgroup viruses and those belonging to the SFV complex had an impact on the design of the template RNAs. The 5'' region of the CHIKV genome contains seven SL structures that affect genome replication in mammalian and/or mosquito cells [33] . Cross-utilisation of template RNAs by alphavirus replicases structures can be predicted for RNAs of other members of the SFV complex (S1 Fig), the 5'' ends of the template RNAs of SFV, MAYV, RRV and ONNV had an identical design to that of the previously constructed CHIKV template, i.e. the 5'' UTR was followed by 231nt encoding the N-terminus of nsP1 [43] . The levels of Fluc and Gluc expression in human cells, transfected with plasmids expressing template RNA and corresponding polymerase negative control replicase (P1234 GAA ), were similar for trans-replicases derived from different viruses and the same was observed for Ae albopictus cells.

abstract:
  Most alphaviruses (family Togaviridae) including Sindbis virus (SINV) and other human pathogens, are transmitted by arthropods. The first open reading frame in their positive strand RNA genome encodes for the non-structural polyprotein, a precursor to four separate subunits of the replicase. The replicase interacts with cis-acting elements located near the intergenic region and at the ends of the viral RNA genome. A trans-replication assay was developed and used to analyse the template requirements for nine alphavirus replicases. Replicases of alphaviruses of the Semliki Forest virus complex were able to cross-utilize each other’s templates as well as those of outgroup alphaviruses. Templates of outgroup alphaviruses, including SINV and the mosquito-specific Eilat virus, were promiscuous; in contrast, their replicases displayed a limited capacity to use heterologous templates, especially in mosquito cells. The determinants important for efficient replication of template RNA were mapped to the 5' region of the genome. For SINV these include the extreme 5'- end of the genome and sequences corresponding to the first stem-loop structure in the 5' untranslated region. Mutations introduced in these elements drastically reduced infectivity of recombinant SINV genomes. The trans-replicase tools and approaches developed here can be instrumental in studying alphavirus recombination and evolution, but can also be applied to study other viruses such as picornaviruses, flaviviruses and coronaviruses.


id: cord-325192-italbsed
author: Desai, Tanay M.
date: 2014-04-03
txt: ./txt/cord-325192-italbsed.txt
cache: ./cache/cord-325192-italbsed.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/24699674/
doi: 10.1371/journal.ppat.1004048
summary:
  Interferon-induced transmembrane proteins (IFITMs) inhibit infection of diverse enveloped viruses, including the influenza A virus (IAV) which is thought to enter from late endosomes. Although late endosomes of IFITM3-expressing cells accumulated cholesterol, other interventions leading to aberrantly high levels of this lipid did not inhibit virus fusion. Our results show that IFITM3 does not inhibit the lipid mixing stage of IAV fusion but blocks the release of viral contents into the cytosol, and that this phenotype does not correlate with cholesterol accumulation in intracellular compartments. Our results thus demonstrate that IFITM3 restricts the IAV fusion at a post-hemifusion step, most likely at the point of fusion pore opening, as evidenced by the dramatic decrease of the BlaM signal in A549 and MDCK cells expressing this protein (Fig. 1A ). Neither IAV lipid mixing (vDiD dequenching) nor fusion (BlaM signal) was inhibited by silencing NPC1 in A549 cells (Fig. 5E, F) .

title:
  IFITM3 Restricts Influenza A Virus Entry by Blocking the Formation of Fusion Pores following Virus-Endosome Hemifusion

abstract:
  Interferon-induced transmembrane proteins (IFITMs) inhibit infection of diverse enveloped viruses, including the influenza A virus (IAV) which is thought to enter from late endosomes. Recent evidence suggests that IFITMs block virus hemifusion (lipid mixing in the absence of viral content release) by altering the properties of cell membranes. Consistent with this mechanism, excess cholesterol in late endosomes of IFITM-expressing cells has been reported to inhibit IAV entry. Here, we examined IAV restriction by IFITM3 protein using direct virus-cell fusion assay and single virus imaging in live cells. IFITM3 over-expression did not inhibit lipid mixing, but abrogated the release of viral content into the cytoplasm. Although late endosomes of IFITM3-expressing cells accumulated cholesterol, other interventions leading to aberrantly high levels of this lipid did not inhibit virus fusion. These results imply that excess cholesterol in late endosomes is not the mechanism by which IFITM3 inhibits the transition from hemifusion to full fusion. The IFITM3's ability to block fusion pore formation at a post-hemifusion stage shows that this protein stabilizes the cytoplasmic leaflet of endosomal membranes without adversely affecting the lumenal leaflet. We propose that IFITM3 interferes with pore formation either directly, through partitioning into the cytoplasmic leaflet of a hemifusion intermediate, or indirectly, by modulating the lipid/protein composition of this leaflet. Alternatively, IFITM3 may redirect IAV fusion to a non-productive pathway, perhaps by promoting fusion with intralumenal vesicles within multivesicular bodies/late endosomes.


id: cord-325326-2bbqz4o7
author: Beitzel, Brett F.
date: 2010-10-14
txt: ./txt/cord-325326-2bbqz4o7.txt
cache: ./cache/cord-325326-2bbqz4o7.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1001146
doi: 10.1371/journal.ppat.1001146
summary:
  We have developed a high-resolution genomic mapping technique that combines transposon-mediated insertional mutagenesis with either capillary electrophoresis or massively parallel sequencing to identify functionally important regions of the Venezuelan equine encephalitis virus (VEEV) genome. Toward this goal, we used transposon mutagenesis, reverse genetics, and fragment analysis by capillary electrophoresis to identify regions of the nsP3 gene that are important for replication and that result in temperature sensitive (ts) mutations. We used transposon mutagenesis to construct a cDNA library with small DNA fragments randomly inserted throughout the VEEV genome and then produced replication-competent virus through reverse genetics. Comparing transposon insertion sites in the resultant viruses to those in the starting library, we were able to produce a functional map of the entire genome of VEEV, and to identify several hundred potential ts mutations, including those we originally identified with the capillary electrophoresis method.

title:
  High-Resolution Functional Mapping of the Venezuelan Equine Encephalitis Virus Genome by Insertional Mutagenesis and Massively Parallel Sequencing

abstract:
  We have developed a high-resolution genomic mapping technique that combines transposon-mediated insertional mutagenesis with either capillary electrophoresis or massively parallel sequencing to identify functionally important regions of the Venezuelan equine encephalitis virus (VEEV) genome. We initially used a capillary electrophoresis method to gain insight into the role of the VEEV nonstructural protein 3 (nsP3) in viral replication. We identified several regions in nsP3 that are intolerant to small (15 bp) insertions, and thus are presumably functionally important. We also identified nine separate regions in nsP3 that will tolerate small insertions at low temperatures (30°C), but not at higher temperatures (37°C, and 40°C). Because we found this method to be extremely effective at identifying temperature sensitive (ts) mutations, but limited by capillary electrophoresis capacity, we replaced the capillary electrophoresis with massively parallel sequencing and used the improved method to generate a functional map of the entire VEEV genome. We identified several hundred potential ts mutations throughout the genome and we validated several of the mutations in nsP2, nsP3, E3, E2, E1 and capsid using single-cycle growth curve experiments with virus generated through reverse genetics. We further demonstrated that two of the nsP3 ts mutants were attenuated for virulence in mice but could elicit protective immunity against challenge with wild-type VEEV. The recombinant ts mutants will be valuable tools for further studies of VEEV replication and virulence. Moreover, the method that we developed is applicable for generating such tools for any virus with a robust reverse genetics system.


id: cord-325624-6anybxnk
title: RNase L Mediated Protection from Virus Induced Demyelination
author: Ireland, Derek D. C.
date: 2009-10-02
txt: ./txt/cord-325624-6anybxnk.txt
cache: ./cache/cord-325624-6anybxnk.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/19798426/
doi: 10.1371/journal.ppat.1000602
summary:
  The unique phenotype of infected RL(−/−) mice was rather manifested in earlier onset and increased severity of demyelination and axonal damage in brain stem and spinal cord without evidence for enhanced neuronal infection. RNase L specifically protected the brain stem from sustained infection and prevented spread of virus to microglia/macrophages located in spinal cord grey matter. The increased pathological changes in both brain stem and spinal cord thus correlated with the high mortality rates of RL 2/2 mice starting at day 9 p.i. The inability to directly link increased axonal damage with enhanced neuronal infection remains puzzling and supports dysregulation of oligodendrocyte function and/or neuroprotective functions of microglia as contributing factors to the severe pathological phenotype and clinical outcome. Numerous functions of RNase L, not directly associated with anti-viral activity, may contribute to the increased susceptibility of RL 2/2 mice to MHV-JHM infection.

abstract:
  IFN-α/β plays a critical role in limiting viral spread, restricting viral tropism and protecting mice from neurotropic coronavirus infection. However, the IFN-α/β dependent mechanisms underlying innate anti-viral functions within the CNS are poorly understood. The role of RNase L in viral encephalomyelitis was explored based on its functions in inhibiting translation, inducing apoptosis, and propagating the IFN-α/β pathway through RNA degradation intermediates. Infection of RNase L deficient (RL(−/−)) mice with a sub-lethal, demyelinating mouse hepatitis virus variant revealed that the majority of mice succumbed to infection by day 12 p.i. However, RNase L deficiency did not affect overall control of infectious virus, or diminish IFN-α/β expression in the CNS. Furthermore, increased morbidity and mortality could not be attributed to altered proinflammatory signals or composition of cells infiltrating the CNS. The unique phenotype of infected RL(−/−) mice was rather manifested in earlier onset and increased severity of demyelination and axonal damage in brain stem and spinal cord without evidence for enhanced neuronal infection. Increased tissue damage coincided with sustained brain stem infection, foci of microglia infection in grey matter, and increased apoptotic cells. These data demonstrate a novel protective role for RNase L in viral induced CNS encephalomyelitis, which is not reflected in overall viral control or propagation of IFN-α/β mediated signals. Protective function is rather associated with cell type specific and regional restriction of viral replication in grey matter and ameliorated neurodegeneration and demyelination.


id: cord-325825-0lyt8gfq
author: Griffiths, Samantha J.
date: 2013-08-08
txt: ./txt/cord-325825-0lyt8gfq.txt
cache: ./cache/cord-325825-0lyt8gfq.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1003514
doi: 10.1371/journal.ppat.1003514
summary:
  Host factors (HFs) which positively or negatively regulate HSV-1 replication were identified by screening a druggable genome siRNA library (4 siRNAs per gene) targeting 7,237 human genes against a HSV-1 reporter virus expressing the enhanced green fluorescent protein (eGFP; HSV-1 strain C12) in the epithelial Hela cell line, due to their ease of transfection and susceptibility to HSV-1 infection [34] . Combined bioinformatic analyses of protein interaction and siRNA depletion screens found a significant functional enrichment for proteins involved in transcription, and identified multi-protein complexes enriched for pro-viral HFs which strongly inhibited HSV-1 upon depletion, including the RNA-polymerase II, eIF3 and Mediator complexes ( Figure 3a) . Furthermore, qPCR analysis found depletion of Med23 inhibited the induction of IFN-l expression following HSV-1 infection of A549 cells in comparison to cells transfected with the RSCF siRNA control ( Figure 4e ).

title:
  A Systematic Analysis of Host Factors Reveals a Med23-Interferon-λ Regulatory Axis against Herpes Simplex Virus Type 1 Replication

abstract:
  Herpes simplex virus type 1 (HSV-1) is a neurotropic virus causing vesicular oral or genital skin lesions, meningitis and other diseases particularly harmful in immunocompromised individuals. To comprehensively investigate the complex interaction between HSV-1 and its host we combined two genome-scale screens for host factors (HFs) involved in virus replication. A yeast two-hybrid screen for protein interactions and a RNA interference (RNAi) screen with a druggable genome small interfering RNA (siRNA) library confirmed existing and identified novel HFs which functionally influence HSV-1 infection. Bioinformatic analyses found the 358 HFs were enriched for several pathways and multi-protein complexes. Of particular interest was the identification of Med23 as a strongly anti-viral component of the largely pro-viral Mediator complex, which links specific transcription factors to RNA polymerase II. The anti-viral effect of Med23 on HSV-1 replication was confirmed in gain-of-function gene overexpression experiments, and this inhibitory effect was specific to HSV-1, as a range of other viruses including Vaccinia virus and Semliki Forest virus were unaffected by Med23 depletion. We found Med23 significantly upregulated expression of the type III interferon family (IFN-λ) at the mRNA and protein level by directly interacting with the transcription factor IRF7. The synergistic effect of Med23 and IRF7 on IFN-λ induction suggests this is the major transcription factor for IFN-λ expression. Genotypic analysis of patients suffering recurrent orofacial HSV-1 outbreaks, previously shown to be deficient in IFN-λ secretion, found a significant correlation with a single nucleotide polymorphism in the IFN-λ3 (IL28b) promoter strongly linked to Hepatitis C disease and treatment outcome. This paper describes a link between Med23 and IFN-λ, provides evidence for the crucial role of IFN-λ in HSV-1 immune control, and highlights the power of integrative genome-scale approaches to identify HFs critical for disease progression and outcome.


id: cord-328947-3l9ydspz
title: Chikungunya virus antagonizes cGAS-STING mediated type-I interferon responses by degrading cGAS
author: Webb, L. G.
date: 2020-10-15
txt: ./txt/cord-328947-3l9ydspz.txt
cache: ./cache/cord-328947-3l9ydspz.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1008999
doi: 10.1371/journal.ppat.1008999
summary:
  CHIKV is no exception, the type-I interferon (IFN) response has been demonstrated to be key in controlling CHIKV infection [21] [22] [23] [24] [25] [26] and primary sensing of the virus is via the pattern recognition receptor (PRR) retinoic acid-inducible gene I (RIG-I)-mitochondrial antiviral signaling protein (MAVS) axis [24, 26] . The lack of cGAS degradation seen in the exogenous expression system when compared to the infected cells shows that although there is a clear interaction of cGAS with nsP4, the degradation of cGAS observed is not mediated by the non-structural proteins of CHIKV, but must require other viral or host factors. Chikungunya virus inhibits DNA sensing by degrading cGAS both mock and infected cells by 4 hpi suggesting that the stress produced during the process of infection (mock or CHIKV) in HFF-1s stimulates increased translation at early timepoints ( Fig 4D, lanes 2 and 5) .

abstract:
  Chikungunya virus (CHIKV) is a mosquito-borne alphavirus known to cause epidemics resulting in predominantly symptomatic infections, which in rare cases cause long term debilitating arthritis and arthralgia. Significant progress has been made in understanding the roles of canonical RNA sensing pathways in the host recognition of CHIKV; however, less is known regarding antagonism of CHIKV by cytosolic DNA sensing pathways like that of cyclic GMP-AMP synthase (cGAS) and Stimulator of Interferon Genes (STING). With the use of cGAS or STING null cells we demonstrate that the pathway restricts CHIKV replication in fibroblasts and immune cells. We show that DNA accumulates in the cytoplasm of infected cells and that CHIKV blocks DNA dependent IFN-β transcription. This antagonism of DNA sensing is via an early autophagy-mediated degradation of cGAS and expression of the CHIKV capsid protein is sufficient to induce cGAS degradation. Furthermore, we identify an interaction of CHIKV nsP1 with STING and map the interaction to 23 residues in the cytosolic loop of the adaptor protein. This interaction stabilizes the viral protein and increases the level of palmitoylated nsP1 in cells. Together, this work supports previous publications highlighting the relevance of the cGAS-STING pathway in the early detection of (+)ssRNA viruses and provides direct evidence that CHIKV interacts with and antagonizes cGAS-STING signaling.


id: cord-331721-l5kocy4f
author: Liang, Jingjing
date: 2017-01-11
txt: ./txt/cord-331721-l5kocy4f.txt
cache: ./cache/cord-331721-l5kocy4f.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1006132
doi: 10.1371/journal.ppat.1006132
summary:
  Here, we demonstrate that the WW-domain of BAG3 interacts with the PPxY motif of both EBOV and MARV VP40 and, unexpectedly, inhibits budding of both eVP40 and mVP40 virus-like particles (VLPs), as well as infectious VSV-EBOV recombinants. As BAG3 is the first WW-domain interactor identified that negatively regulates budding of VP40 VLPs and infectious virus, we propose that the chaperone-mediated autophagy function of BAG3 represents a specific host defense strategy to counteract the function of VP40 in promoting efficient egress and spread of virus particles. Indeed, we confirmed that hypothesis by first using co-IP to validate the specificity of the PPxY/WW-domain physical interaction between VP40 (both eVP40 and mVP40) and BAG3, and functionally demonstrated that expression of BAG3 inhibited VP40 VLP production, as well as budding of a VSV recombinant virus containing the EBOV VP40 PPxY L-domain motif.

title:
  Chaperone-Mediated Autophagy Protein BAG3 Negatively Regulates Ebola and Marburg VP40-Mediated Egress

abstract:
  Ebola (EBOV) and Marburg (MARV) viruses are members of the Filoviridae family which cause outbreaks of hemorrhagic fever. The filovirus VP40 matrix protein is essential for virus assembly and budding, and its PPxY L-domain motif interacts with WW-domains of specific host proteins, such as Nedd4 and ITCH, to facilitate the late stage of virus-cell separation. To identify additional WW-domain-bearing host proteins that interact with VP40, we used an EBOV PPxY-containing peptide to screen an array of 115 mammalian WW-domain-bearing proteins. Using this unbiased approach, we identified BCL2 Associated Athanogene 3 (BAG3), a member of the BAG family of molecular chaperone proteins, as a specific VP40 PPxY interactor. Here, we demonstrate that the WW-domain of BAG3 interacts with the PPxY motif of both EBOV and MARV VP40 and, unexpectedly, inhibits budding of both eVP40 and mVP40 virus-like particles (VLPs), as well as infectious VSV-EBOV recombinants. BAG3 is a stress induced protein that regulates cellular protein homeostasis and cell survival through chaperone-mediated autophagy (CMA). Interestingly, our results show that BAG3 alters the intracellular localization of VP40 by sequestering VP40 away from the plasma membrane. As BAG3 is the first WW-domain interactor identified that negatively regulates budding of VP40 VLPs and infectious virus, we propose that the chaperone-mediated autophagy function of BAG3 represents a specific host defense strategy to counteract the function of VP40 in promoting efficient egress and spread of virus particles.


id: cord-331802-wo462anq
author: Xia, Hongjie
date: 2015-07-28
txt: ./txt/cord-331802-wo462anq.txt
cache: ./cache/cord-331802-wo462anq.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/26218680/
doi: 10.1371/journal.ppat.1005067
summary:
  Herein, we report that the nonstructural protein 2C(ATPase) of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3′-to-5′ unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. Herein, we report that although bacterially expressed EV71 2C ATPase did not exhibit any RNA remodeling activity, eukaryotically expressed EV71 2C ATPase functions not only as an RNA helicase that unwinds RNA helices from 3 0 to 5 0 in an ATP-dependent manner but also as an RNA chaperone that destabilizes helices from either direction and facilitates strand annealing and complex RNA structure formation independently of ATP.

title:
  Human Enterovirus Nonstructural Protein 2C(ATPase) Functions as Both an RNA Helicase and ATP-Independent RNA Chaperone

abstract:
  RNA helicases and chaperones are the two major classes of RNA remodeling proteins, which function to remodel RNA structures and/or RNA-protein interactions, and are required for all aspects of RNA metabolism. Although some virus-encoded RNA helicases/chaperones have been predicted or identified, their RNA remodeling activities in vitro and functions in the viral life cycle remain largely elusive. Enteroviruses are a large group of positive-stranded RNA viruses in the Picornaviridae family, which includes numerous important human pathogens. Herein, we report that the nonstructural protein 2C(ATPase) of enterovirus 71 (EV71), which is the major causative pathogen of hand-foot-and-mouth disease and has been regarded as the most important neurotropic enterovirus after poliovirus eradication, functions not only as an RNA helicase that 3′-to-5′ unwinds RNA helices in an adenosine triphosphate (ATP)-dependent manner, but also as an RNA chaperone that destabilizes helices bidirectionally and facilitates strand annealing and complex RNA structure formation independently of ATP. We also determined that the helicase activity is based on the EV71 2C(ATPase) middle domain, whereas the C-terminus is indispensable for its RNA chaperoning activity. By promoting RNA template recycling, 2C(ATPase) facilitated EV71 RNA synthesis in vitro; when 2C(ATPase) helicase activity was impaired, EV71 RNA replication and virion production were mostly abolished in cells, indicating that 2C(ATPase)-mediated RNA remodeling plays a critical role in the enteroviral life cycle. Furthermore, the RNA helicase and chaperoning activities of 2C(ATPase) are also conserved in coxsackie A virus 16 (CAV16), another important enterovirus. Altogether, our findings are the first to demonstrate the RNA helicase and chaperoning activities associated with enterovirus 2C(ATPase), and our study provides both in vitro and cellular evidence for their potential roles during viral RNA replication. These findings increase our understanding of enteroviruses and the two types of RNA remodeling activities.


id: cord-332205-ydijp66b
title: Virologists—Heroes need weapons
author: Hufsky, Franziska
date: 2018-02-08
txt: ./txt/cord-332205-ydijp66b.txt
cache: ./cache/cord-332205-ydijp66b.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1006771
doi: 10.1371/journal.ppat.1006771
abstract: nan
summary:
  Nowadays, nearly everyone in the life sciences has used BLAST [8] at least once, or made an alignment, or asked a bioinformatician to analyze high-throughput sequencing data. Bioinformaticians routinely have to develop tailored, study-specific algorithms and tools used by a wide variety of scientists, including biochemists, biologists, geneticists, and molecular life scientists; but we rarely find virus-specific tools used by virologists. But astonishingly, we now know that the human genome consists of 8%-60% virus-derived sequences (depending on how this is measured: 8% can be directly traced back to viruses, whereas a figure of 60% includes LINEs and SINEs that are thought to be of viral origin [12] ). The EVBC aims to develop bioinformatical tools for nearly all areas: (1) for detection of viruses, e.g., from high-throughput sequencing data; (2) virus assembly; (3) quasispecies reconstruction; (4) intraviral interactions; (5) virus entry, i.e., protein-protein interaction; (6) virus -host interactions; (7) phylogeny/cophylogeny; and (8) therapy.


id: cord-332747-u46xryoo
author: Mingorance, Lidia
date: 2018-09-18
url: https://www.ncbi.nlm.nih.gov/pubmed/30226904/
doi: 10.1371/journal.ppat.1007284
title:
  Host phosphatidic acid phosphatase lipin1 is rate limiting for functional hepatitis C virus replicase complex formation

abstract:
  Hepatitis C virus (HCV) infection constitutes a significant health burden worldwide, because it is a major etiologic agent of chronic liver disease, cirrhosis and hepatocellular carcinoma. HCV replication cycle is closely tied to lipid metabolism and infection by this virus causes profound changes in host lipid homeostasis. We focused our attention on a phosphatidate phosphate (PAP) enzyme family (the lipin family), which mediate the conversion of phosphatidate to diacylglycerol in the cytoplasm, playing a key role in triglyceride biosynthesis and in phospholipid homeostasis. Lipins may also translocate to the nucleus to act as transcriptional regulators of genes involved in lipid metabolism. The best-characterized member of this family is lipin1, which cooperates with lipin2 to maintain glycerophospholipid homeostasis in the liver. Lipin1-deficient cell lines were generated by RNAi to study the role of this protein in different steps of HCV replication cycle. Using surrogate models that recapitulate different aspects of HCV infection, we concluded that lipin1 is rate limiting for the generation of functional replicase complexes, in a step downstream primary translation that leads to early HCV RNA replication. Infection studies in lipin1-deficient cells overexpressing wild type or phosphatase-defective lipin1 proteins suggest that lipin1 phosphatase activity is required to support HCV infection. Finally, ultrastructural and biochemical analyses in replication-independent models suggest that lipin1 may facilitate the generation of the membranous compartment that contains functional HCV replicase complexes.


id: cord-333473-c1lykari
title: High-Resolution Analysis of Coronavirus Gene Expression by RNA Sequencing and Ribosome Profiling
author: Irigoyen, Nerea
date: 2016-02-26
txt: ./txt/cord-333473-c1lykari.txt
cache: ./cache/cord-333473-c1lykari.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/26919232/
doi: 10.1371/journal.ppat.1005473
summary:
  Also, it has been employed in the study of the replication of large DNA viruses; namely, human cytomegalovirus [17] [18] , Kaposi''s sarcoma-associated herpesvirus [19] , herpes simplex virus 1 [20] , vaccinia virus [21] , and bacteriophage lambda [22] , providing insights into the temporal regulation of gene expression in these viruses and identifying numerous previously unrecognized translated ORFs, including novel protein-coding ORFs and short regulatory uORFs. In this paper, we describe the first analysis of RNA virus replication and gene expression by ribosome profiling (and parallel RNASeq), using MHV as a model system. The latter are calculated on a per gene (rather than per transcript) basis, using RNASeq and RiboSeq reads contained entirely within annotated CDS regions (i.e. excluding 5 0 and 3 0 UTRs and also RPFs accumulating at or near to initiation or termination sites), and, like the virus values, are expressed relative to the mean levels for the cell (due to normalization by library size).

abstract:
  Members of the family Coronaviridae have the largest genomes of all RNA viruses, typically in the region of 30 kilobases. Several coronaviruses, such as Severe acute respiratory syndrome-related coronavirus (SARS-CoV) and Middle East respiratory syndrome-related coronavirus (MERS-CoV), are of medical importance, with high mortality rates and, in the case of SARS-CoV, significant pandemic potential. Other coronaviruses, such as Porcine epidemic diarrhea virus and Avian coronavirus, are important livestock pathogens. Ribosome profiling is a technique which exploits the capacity of the translating ribosome to protect around 30 nucleotides of mRNA from ribonuclease digestion. Ribosome-protected mRNA fragments are purified, subjected to deep sequencing and mapped back to the transcriptome to give a global “snap-shot” of translation. Parallel RNA sequencing allows normalization by transcript abundance. Here we apply ribosome profiling to cells infected with Murine coronavirus, mouse hepatitis virus, strain A59 (MHV-A59), a model coronavirus in the same genus as SARS-CoV and MERS-CoV. The data obtained allowed us to study the kinetics of virus transcription and translation with exquisite precision. We studied the timecourse of positive and negative-sense genomic and subgenomic viral RNA production and the relative translation efficiencies of the different virus ORFs. Virus mRNAs were not found to be translated more efficiently than host mRNAs; rather, virus translation dominates host translation at later time points due to high levels of virus transcripts. Triplet phasing of the profiling data allowed precise determination of translated reading frames and revealed several translated short open reading frames upstream of, or embedded within, known virus protein-coding regions. Ribosome pause sites were identified in the virus replicase polyprotein pp1a ORF and investigated experimentally. Contrary to expectations, ribosomes were not found to pause at the ribosomal frameshift site. To our knowledge this is the first application of ribosome profiling to an RNA virus.


id: cord-334315-ymkrgj0h
title: Cytoplasmic Viruses: Rage against the (Cellular RNA Decay) Machine
author: Moon, Stephanie L.
date: 2013-12-05
txt: ./txt/cord-334315-ymkrgj0h.txt
cache: ./cache/cord-334315-ymkrgj0h.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/24339774/
doi: 10.1371/journal.ppat.1003762
abstract: nan
summary:
  This review describes the myriad ways that viruses deal with the general host RNA decay machinery that is active in the cell immediately upon viral infection-turning what, at first, appears to be very hostile territory for a foreign transcript into a sort of ''''promised land'''' for viral gene expression. Disparate RNA viruses have, therefore, evolved unique mechanisms by which they disarm host RNA decay pathways by inactivating or proteolytically degrading important nucleases to promote productive viral infections. In addition, to make a cell more amenable to virus production, these virally encoded nucleases may also create a new ''''sandbox'''' in the cytoplasm for viral RNAs by initiating the large-scale decay of cellular mRNAs and dramatically altering the landscape of host gene expression. Considering the importance of RNA stability in regulating transcript abundance, the inactivation or commandeering of cellular RNA decay factors by viruses is likely to significantly alter host gene expression.


id: cord-334947-pa0p5dif
author: Rozen-Gagnon, Kathryn
date: 2014-01-16
txt: ./txt/cord-334947-pa0p5dif.txt
cache: ./cache/cord-334947-pa0p5dif.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/24453971/
doi: 10.1371/journal.ppat.1003877
summary:
  Since residue 483 is conserved among alphaviruses, we examined the analogous mutations in Sindbis virus (SINV), which also reduced polymerase fidelity and generated replication defects in mosquito cells. To estimate the population diversity of variants by deep sequencing, cDNA libraries were prepared by Superscript III from RNA extracted from virus generated in BHK-21 or C6/36 cells, and the viral genome was amplified using a high fidelity polymerase (Phusion) to generate 5 overlapping amplicons 2-3 kb in length. To address the possibility that the fitness cost of defective replication, observed in mosquito cell culture, would favor the reversion of these mutant polymerases to wildtype, we deep sequenced virus from the body of an individual mosquito that presented the median titer from each group.

title:
  Alphavirus Mutator Variants Present Host-Specific Defects and Attenuation in Mammalian and Insect Models

abstract:
  Arboviruses cycle through both vertebrates and invertebrates, which requires them to adapt to disparate hosts while maintaining genetic integrity during genome replication. To study the genetic mechanisms and determinants of these processes, we use chikungunya virus (CHIKV), a re-emerging human pathogen transmitted by the Aedes mosquito. We previously isolated a high fidelity (or antimutator) polymerase variant, C483Y, which had decreased fitness in both mammalian and mosquito hosts, suggesting this residue may be a key molecular determinant. To further investigate effects of position 483 on RNA-dependent RNA-polymerase (RdRp) fidelity, we substituted every amino acid at this position. We isolated novel mutators with decreased replication fidelity and higher mutation frequencies, allowing us to examine the fitness of error-prone arbovirus variants. Although CHIKV mutators displayed no major replication defects in mammalian cell culture, they had reduced specific infectivity and were attenuated in vivo. Unexpectedly, mutator phenotypes were suppressed in mosquito cells and the variants exhibited significant defects in RNA synthesis. Consequently, these replication defects resulted in strong selection for reversion during infection of mosquitoes. Since residue 483 is conserved among alphaviruses, we examined the analogous mutations in Sindbis virus (SINV), which also reduced polymerase fidelity and generated replication defects in mosquito cells. However, replication defects were mosquito cell-specific and were not observed in Drosophila S2 cells, allowing us to evaluate the potential attenuation of mutators in insect models where pressure for reversion was absent. Indeed, the SINV mutator variant was attenuated in fruit flies. These findings confirm that residue 483 is a determinant regulating alphavirus polymerase fidelity and demonstrate proof of principle that arboviruses can be attenuated in mammalian and insect hosts by reducing fidelity.


id: cord-338053-8zlxsf1z
author: Foo, Suan-Sin
date: 2015-02-19
txt: ./txt/cord-338053-8zlxsf1z.txt
cache: ./cache/cord-338053-8zlxsf1z.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/25695775/
doi: 10.1371/journal.ppat.1004649
summary:
  In summary, our data demonstrated the crucial role of PTX3 in modulating alphavirus-induced immune responses and disease manifestation through its N-terminal interaction with the virus particles leading to enhanced viral entry and replication. To confirm that the results of enhanced virus production was due to PTX3 enhancing RRV replication, HEK 293T cells transiently transfected with vector or hPTX3 plasmids were harvested at 20 hour post transfection (hpt) ( To further characterize the effect of PTX3 during alphaviral infection, we examined the potential of PTX3 to directly interact with the virus and enhance viral entry. To demonstrate that the effect of PTX3on enhancing RRV entry and replication contributed to the increased level of virus detected in the in vivo studies, we performed RRV infection on primary fibroblasts isolated from tails of PTX3 -/and WT C57BL/6 mice.

title:
  Role of Pentraxin 3 in Shaping Arthritogenic Alphaviral Disease: From Enhanced Viral Replication to Immunomodulation

abstract:
  The rising prevalence of arthritogenic alphavirus infections, including chikungunya virus (CHIKV) and Ross River virus (RRV), and the lack of antiviral treatments highlight the potential threat of a global alphavirus pandemic. The immune responses underlying alphavirus virulence remain enigmatic. We found that pentraxin 3 (PTX3) was highly expressed in CHIKV and RRV patients during acute disease. Overt expression of PTX3 in CHIKV patients was associated with increased viral load and disease severity. PTX3-deficient (PTX3(-/-)) mice acutely infected with RRV exhibited delayed disease progression and rapid recovery through diminished inflammatory responses and viral replication. Furthermore, binding of the N-terminal domain of PTX3 to RRV facilitated viral entry and replication. Thus, our study demonstrates the pivotal role of PTX3 in shaping alphavirus-triggered immunity and disease and provides new insights into alphavirus pathogenesis.


id: cord-338400-30vl2hks
author: Epstein, Jonathan H.
date: 2010-07-01
txt: ./txt/cord-338400-30vl2hks.txt
cache: ./cache/cord-338400-30vl2hks.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1000972
doi: 10.1371/journal.ppat.1000972
summary:
  Phylogenetic analysis indicates that this first GBV-like flavivirus reported in bats constitutes a distinct species within the Flaviviridae family and is ancestral to the GBV-A and -C virus clades. GBV-A viruses have been described in New World primates and are not known to infect humans [17] [18] [19] , while GBV-C (also known as Hepatitis G virus (HGV)) have frequently been isolated from humans in many regions of the World, including India and Bangladesh [19] [20] [21] [22] [23] , and from wild chimpanzees (Pan troglodytes) in Africa [24, 25] . Our findings provide new insight into the range of known hosts for GB-like viruses and demonstrate the power of unbiased sequencing to characterize the diversity of potentially zoonotic pathogens carried by bats and other reservoirs. Molecular analyses of sera from Pteropus giganteus bats from Faridpur, Bangladesh led to the identification of a 9,633 nt sequence consistent in genomic organization with known GBV and other species within the family Flaviviridae [16] .

title:
  Identification of GBV-D, a Novel GB-like Flavivirus from Old World Frugivorous Bats (Pteropus giganteus) in Bangladesh

abstract:
  Bats are reservoirs for a wide range of zoonotic agents including lyssa-, henipah-, SARS-like corona-, Marburg-, Ebola-, and astroviruses. In an effort to survey for the presence of other infectious agents, known and unknown, we screened sera from 16 Pteropus giganteus bats from Faridpur, Bangladesh, using high-throughput pyrosequencing. Sequence analyses indicated the presence of a previously undescribed virus that has approximately 50% identity at the amino acid level to GB virus A and C (GBV-A and -C). Viral nucleic acid was present in 5 of 98 sera (5%) from a single colony of free-ranging bats. Infection was not associated with evidence of hepatitis or hepatic dysfunction. Phylogenetic analysis indicates that this first GBV-like flavivirus reported in bats constitutes a distinct species within the Flaviviridae family and is ancestral to the GBV-A and -C virus clades.


id: cord-341034-2oigu75k
title: AMP-Activated Kinase Restricts Rift Valley Fever Virus Infection by Inhibiting Fatty Acid Synthesis
author: Moser, Theresa S.
date: 2012-04-19
url: https://doi.org/10.1371/journal.ppat.1002661
doi: 10.1371/journal.ppat.1002661
abstract:
  The cell intrinsic innate immune responses provide a first line of defense against viral infection, and often function by targeting cellular pathways usurped by the virus during infection. In particular, many viruses manipulate cellular lipids to form complex structures required for viral replication, many of which are dependent on de novo fatty acid synthesis. We found that the energy regulator AMPK, which potently inhibits fatty acid synthesis, restricts infection of the Bunyavirus, Rift Valley Fever Virus (RVFV), an important re-emerging arthropod-borne human pathogen for which there are no effective vaccines or therapeutics. We show restriction of RVFV both by AMPK and its upstream activator LKB1, indicating an antiviral role for this signaling pathway. Furthermore, we found that AMPK is activated during RVFV infection, leading to the phosphorylation and inhibition of acetyl-CoA carboxylase, the first rate-limiting enzyme in fatty acid synthesis. Activating AMPK pharmacologically both restricted infection and reduced lipid levels. This restriction could be bypassed by treatment with the fatty acid palmitate, demonstrating that AMPK restricts RVFV infection through its inhibition of fatty acid biosynthesis. Lastly, we found that this pathway plays a broad role in antiviral defense since additional viruses from disparate families were also restricted by AMPK and LKB1. Therefore, AMPK is an important component of the cell intrinsic immune response that restricts infection through a novel mechanism involving the inhibition of fatty acid metabolism.


id: cord-342291-imn7g084
title: Bats reveal the true power of influenza A virus adaptability
author: Ciminski, Kevin
date: 2020-04-16
txt: ./txt/cord-342291-imn7g084.txt
cache: ./cache/cord-342291-imn7g084.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/32298389/
doi: 10.1371/journal.ppat.1008384
abstract: nan
summary:
  Indeed, the HA of the newly discovered Old World bat H9N2 virus binds to α2,3-sialic In contrast, the internal gene segments of the bat-derived IAVs form two outgroups that are located at a more basal position. Conventional IAVs and the bat H9N2 virus possess the surface glycoprotein neuraminidase (NA), which removes sialic acid residues from infected cells to facilitate the release of newly formed viral particles (Fig 2A) . The amazing ability of H18 to rapidly overcome the absence of a functional N11 suggests that the structure of H18 (and possibly H17) may provide a broader scope of evolutionary flexibility than that of conventional sialic acid-dependent IAVs. It is therefore tempting to speculate that bat IAV HA proteins, and in particular H18, might have the potential to adapt to novel and so far unknown entry receptors different from MHC-II (Fig 2D) .


id: cord-342419-liaihahj
title: Antibody recognition of the Pneumovirus fusion protein trimer interface
author: Huang, Jiachen
date: 2020-10-09
url: https://www.ncbi.nlm.nih.gov/pubmed/33035266/
doi: 10.1371/journal.ppat.1008942
abstract:
  Human metapneumovirus (hMPV) is a leading cause of viral respiratory infection in children, and can cause severe lower respiratory tract infection in infants, the elderly, and immunocompromised patients. However, there remain no licensed vaccines or specific treatments for hMPV infection. Although the hMPV fusion (F) protein is the sole target of neutralizing antibodies, the immunological properties of hMPV F remain poorly understood. To further define the humoral immune response to the hMPV F protein, we isolated two new human monoclonal antibodies (mAbs), MPV458 and MPV465. Both mAbs are neutralizing in vitro and were determined to target a unique antigenic site using competitive biolayer interferometry. We determined both MPV458 and MPV465 have higher affinity for monomeric hMPV F than trimeric hMPV F. MPV458 was co-crystallized with hMPV F, and the mAb primarily interacts with an alpha helix on the F2 region of the hMPV F protein. Surprisingly, the major epitope for MPV458 lies within the trimeric interface of the hMPV F protein, suggesting significant breathing of the hMPV F protein must occur for host immune recognition of the novel epitope. In addition, significant glycan interactions were observed with a somatically mutated light chain framework residue. The data presented identifies a novel epitope on the hMPV F protein for epitope-based vaccine design, and illustrates a new mechanism for human antibody neutralization of viral glycoproteins.


id: cord-342825-s8ddek2f
title: Consecutive Inhibition of ISG15 Expression and ISGylation by Cytomegalovirus Regulators
author: Kim, Ye Ji
date: 2016-08-26
txt: ./txt/cord-342825-s8ddek2f.txt
cache: ./cache/cord-342825-s8ddek2f.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1005850
doi: 10.1371/journal.ppat.1005850
summary:
  RNA interference of UBE1L (E1), UbcH8 (E2), Herc5 (E3), and UBP43 (ISG15 protease) revealed that ISGylation inhibits HCMV growth by downregulating viral gene expression and virion release in a manner that is more prominent at low multiplicity of infection. This result suggests that an indirect effect of virus infection on neighboring uninfected cells is responsible for the greater induction of ISG15 and protein ISGylation by HCMV than by UV-HCMV at low MOIs. Since HCMV IE1 inhibits the activation of ISRE-containing promoters by sequestering STAT2 [58] [59] [60] and PML [61] , IE1 expression may be responsible for the suppression of free ISG15 and ISG15 conjugate levels during HCMV infection. In addition to IE1, the presence of additional viral functions that downregulate protein ISGylation was prompted by our observation that UV-HCMV infection resulted in a higher level of ISG15 conjugates than IE1-deleted mutant virus at late times (Fig 2B) .

abstract:
  Interferon-stimulated gene 15 (ISG15) encodes an ubiquitin-like protein that covalently conjugates protein. Protein modification by ISG15 (ISGylation) is known to inhibit the replication of many viruses. However, studies on the viral targets and viral strategies to regulate ISGylation-mediated antiviral responses are limited. In this study, we show that human cytomegalovirus (HCMV) replication is inhibited by ISGylation, but the virus has evolved multiple countermeasures. HCMV-induced ISG15 expression was mitigated by IE1, a viral inhibitor of interferon signaling, however, ISGylation was still strongly upregulated during virus infection. RNA interference of UBE1L (E1), UbcH8 (E2), Herc5 (E3), and UBP43 (ISG15 protease) revealed that ISGylation inhibits HCMV growth by downregulating viral gene expression and virion release in a manner that is more prominent at low multiplicity of infection. A viral regulator pUL26 was found to interact with ISG15, UBE1L, and Herc5, and be ISGylated. ISGylation of pUL26 regulated its stability and inhibited its activities to suppress NF-κB signaling and complement the growth of UL26-null mutant virus. Moreover, pUL26 reciprocally suppressed virus-induced ISGylation independent of its own ISGylation. Consistently, ISGylation was more pronounced in infections with the UL26-deleted mutant virus, whose growth was more sensitive to IFNβ treatment than that of the wild-type virus. Therefore, pUL26 is a viral ISG15 target that also counteracts ISGylation. Our results demonstrate that ISGylation inhibits HCMV growth at multiple steps and that HCMV has evolved countermeasures to suppress ISG15 transcription and protein ISGylation, highlighting the importance of the interplay between virus and ISGylation in productive viral infection.


id: cord-343221-e29of29o
title: Early endonuclease-mediated evasion of RNA sensing ensures efficient coronavirus replication
author: Kindler, Eveline
date: 2017-02-03
txt: ./txt/cord-343221-e29of29o.txt
cache: ./cache/cord-343221-e29of29o.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/28158275/
doi: 10.1371/journal.ppat.1006195
summary:
  Specifically, we show that genetically engineered mutants of mouse hepatitis virus (MHV) and human coronavirus 229E (HCoV-229E), respectively, that encode an EndoU active-site substitution known to abolish nucleolytic activity, were severely attenuated and elicited an immediate and simultaneous activation of host cell dsRNA sensors. The severe attenuation of MHV H277A and HCoV-229E H250A replication in wild-type murine and human macrophages, respectively, precluded the use of primary macrophages to assess the sensitivity of EndoU-deficient coronaviruses to IFN-I pre-treatment. Our data suggest that direct restriction of replication of EndoU-deficient coronaviruses is mediated by RNase L-mediated RNA degradation and inhibition of host cell translation through activation of PKR. Notably, also in this case Xrn1 is required to further process the de-capped mRNAs. Our data show that early during coronavirus infection the EndoU activity conceptually fulfils the same task, namely the removal of dsRNA that would otherwise trigger host cell dsRNA responses, such as IFN-β expression and activation of PKR and OAS/RNase L.

abstract:
  Coronaviruses are of veterinary and medical importance and include highly pathogenic zoonotic viruses, such as SARS-CoV and MERS-CoV. They are known to efficiently evade early innate immune responses, manifesting in almost negligible expression of type-I interferons (IFN-I). This evasion strategy suggests an evolutionary conserved viral function that has evolved to prevent RNA-based sensing of infection in vertebrate hosts. Here we show that the coronavirus endonuclease (EndoU) activity is key to prevent early induction of double-stranded RNA (dsRNA) host cell responses. Replication of EndoU-deficient coronaviruses is greatly attenuated in vivo and severely restricted in primary cells even during the early phase of the infection. In macrophages we found immediate induction of IFN-I expression and RNase L-mediated breakdown of ribosomal RNA. Accordingly, EndoU-deficient viruses can retain replication only in cells that are deficient in IFN-I expression or sensing, and in cells lacking both RNase L and PKR. Collectively our results demonstrate that the coronavirus EndoU efficiently prevents simultaneous activation of host cell dsRNA sensors, such as Mda5, OAS and PKR. The localization of the EndoU activity at the site of viral RNA synthesis–within the replicase complex—suggests that coronaviruses have evolved a viral RNA decay pathway to evade early innate and intrinsic antiviral host cell responses.


id: cord-346053-mk1mzc5z
author: Morris, Cindy E.
date: 2009-12-24
txt: ./txt/cord-346053-mk1mzc5z.txt
cache: ./cache/cord-346053-mk1mzc5z.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/20041212/
doi: 10.1371/journal.ppat.1000693
abstract: nan
summary:
  We present numerous examples of virulence traits in plant pathogenic microorganisms that also have a function in their survival and growth in nonagricultural and nonplant habitats. Adaptation to biotic and abiotic stresses, within or outside of agricultural habitats, likely plays as important a role in the evolution of parasitic fitness of plant pathogens as it does for human pathogens. As illustrated above, traits that confer fitness in response to biotic and abiotic environmental stress can have dual-use as virulence factors in human pathogens. In plant pathogens, the transport systems for toxins and antimicrobials can have broad spectrum activity, leading to resistance to agricultural fungicides and also contributing to virulence [12] . The examples listed above that describe traits that play roles in both environmental fitness and virulence to plants provide a compelling incentive to expand our paradigms concerning the forces that drive evolution of plant pathogenicity.

title:
  Expanding the Paradigms of Plant Pathogen Life History and Evolution of Parasitic Fitness beyond Agricultural Boundaries


id: cord-348799-qu4zin3o
author: Wu, Nannan
date: 2019-10-10
txt: ./txt/cord-348799-qu4zin3o.txt
cache: ./cache/cord-348799-qu4zin3o.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1008093
doi: 10.1371/journal.ppat.1008093
summary:
  This lack of specificity was not present in cells however, given that the ectopic expression of ISG20 did not lead to the indiscriminate degradation of cellular RNAs (ribosomal as well as small nucleolar RNAs previously shown to be associated to ISG20 [2] ), nor of an Hepatitis C virus (HCV)-derived Luciferase reporter mRNA produced in vitro and then transfected into cells (S4C and S4D Fig) , suggesting that in cells the RNase activity of ISG20 is tightly controlled. Self mimicry allows the escape of target genes'' mRNAs from the effects of ISG20 VSV infection and ectopic DNA or RNA transfection do not bear much in common apart from the fact that both represent artificial injections into the cell of non-self genetic material from without. To further determine whether ISG20 acted on translation initiation and more specifically through specific initiation factors (eIFs), capped and polyadenylated mRNAs coding luciferase under the control of several 5'' untranslated regions (5'' UTRs) were generated in vitro and directly transfected in ISG20-expressing cells (Fig 3D) .

title:
  The interferon stimulated gene 20 protein (ISG20) is an innate defense antiviral factor that discriminates self versus non-self translation

abstract:
  ISG20 is a broad spectrum antiviral protein thought to directly degrade viral RNA. However, this mechanism of inhibition remains controversial. Using the Vesicular Stomatitis Virus (VSV) as a model RNA virus, we show here that ISG20 interferes with viral replication by decreasing protein synthesis in the absence of RNA degradation. Importantly, we demonstrate that ISG20 exerts a translational control over a large panel of non-self RNA substrates including those originating from transfected DNA, while sparing endogenous transcripts. This activity correlates with the protein’s ability to localize in cytoplasmic processing bodies. Finally, these functions are conserved in the ISG20 murine ortholog, whose genetic ablation results in mice with increased susceptibility to viral infection. Overall, our results posit ISG20 as an important defense factor able to discriminate the self/non-self origins of the RNA through translation modulation.


id: cord-350640-sz6xj5o3
author: Menzel, Nicolas
date: 2012-07-26
url: https://www.ncbi.nlm.nih.gov/pubmed/22911431/
doi: 10.1371/journal.ppat.1002829
title:
  MAP-Kinase Regulated Cytosolic Phospholipase A2 Activity Is Essential for Production of Infectious Hepatitis C Virus Particles

abstract:
  Hepatitis C virus (HCV) has infected around 160 million individuals. Current therapies have limited efficacy and are fraught with side effects. To identify cellular HCV dependency factors, possible therapeutic targets, we manipulated signaling cascades with pathway-specific inhibitors. Using this approach we identified the MAPK/ERK regulated, cytosolic, calcium-dependent, group IVA phospholipase A2 (PLA2G4A) as a novel HCV dependency factor. Inhibition of PLA2G4A activity reduced core protein abundance at lipid droplets, core envelopment and secretion of particles. Moreover, released particles displayed aberrant protein composition and were 100-fold less infectious. Exogenous addition of arachidonic acid, the cleavage product of PLA2G4A-catalyzed lipolysis, but not other related poly-unsaturated fatty acids restored infectivity. Strikingly, production of infectious Dengue virus, a relative of HCV, was also dependent on PLA2G4A. These results highlight previously unrecognized parallels in the assembly pathways of these human pathogens, and define PLA2G4A-dependent lipolysis as crucial prerequisite for production of highly infectious viral progeny.


id: cord-351389-48tszqh5
author: Xu, Kai
date: 2013-10-10
txt: ./txt/cord-351389-48tszqh5.txt
cache: ./cache/cord-351389-48tszqh5.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/24130486/
doi: 10.1371/journal.ppat.1003684
summary:
  title: Crystal Structure of the Hendra Virus Attachment G Glycoprotein Bound to a Potent Cross-Reactive Neutralizing Human Monoclonal Antibody One cross-reactive and receptor-blocking hmAb (m102.4) was recently demonstrated to be an effective post-exposure therapy in two animal models of NiV and HeV infection, has been used in several people on a compassionate use basis, and is currently in development for use in humans. We used X-ray crystallography to determine the high-resolution structures of the Hendra virus G glycoprotein in complex with a cross-reactive neutralizing human monoclonal antibody. Taken together, the success of hmAb m102.4 in vivo as an effective post-exposure treatment against henipavirus disease in two different well-characterized animal models (the ferret and nonhuman primate), along with the new detailed structural findings on its viral G glycoprotein binding features that help explain its superior cross-reactive neutralizing activity, will facilitate efforts aimed at obtaining approved human use application to treat accidental exposure to HeV or NiV infection.

title:
  Crystal Structure of the Hendra Virus Attachment G Glycoprotein Bound to a Potent Cross-Reactive Neutralizing Human Monoclonal Antibody

abstract:
  The henipaviruses, represented by Hendra (HeV) and Nipah (NiV) viruses are highly pathogenic zoonotic paramyxoviruses with uniquely broad host tropisms responsible for repeated outbreaks in Australia, Southeast Asia, India and Bangladesh. The high morbidity and mortality rates associated with infection and lack of licensed antiviral therapies make the henipaviruses a potential biological threat to humans and livestock. Henipavirus entry is initiated by the attachment of the G envelope glycoprotein to host cell membrane receptors. Previously, henipavirus-neutralizing human monoclonal antibodies (hmAb) have been isolated using the HeV-G glycoprotein and a human naïve antibody library. One cross-reactive and receptor-blocking hmAb (m102.4) was recently demonstrated to be an effective post-exposure therapy in two animal models of NiV and HeV infection, has been used in several people on a compassionate use basis, and is currently in development for use in humans. Here, we report the crystal structure of the complex of HeV-G with m102.3, an m102.4 derivative, and describe NiV and HeV escape mutants. This structure provides detailed insight into the mechanism of HeV and NiV neutralization by m102.4, and serves as a blueprint for further optimization of m102.4 as a therapeutic agent and for the development of entry inhibitors and vaccines.


id: cord-351548-jvl63652
author: Juranic Lisnic, Vanda
date: 2013-09-26
txt: ./txt/cord-351548-jvl63652.txt
cache: ./cache/cord-351548-jvl63652.txt
extension: .txt
mime: text/plain
url: https://doi.org/10.1371/journal.ppat.1003611
doi: 10.1371/journal.ppat.1003611
summary:
  Finally, a recent transcriptomic analysis of newly synthesized RNA in MCMV infected fibroblasts [15] applied RNA-Seq technology to study regulation of viral gene expression and identified a very early peak of viral gene transcriptional activity at 1-2 hours post infection followed by rapid cellular countermeasures but did not attempt to re-annotate MCMV genome. The MCMV transcripts identified through our classical cDNA cloning and sequencing (green arrows) and the RNA-Seq expression profiles (gray histograms), showed complementary results to each other but diverged dramatically from current annotations. Because cDNA cloning and RNA-Seq identified significant differences between the MCMV transcriptome and current annotations, we performed an in depth analysis of several genomic regions by northern analyses ( Figure 5 , Figures S2, S3 , S4, S5) using our cDNA clones to generate strand specific riboprobes ( Table 1) .

title:
  Dual Analysis of the Murine Cytomegalovirus and Host Cell Transcriptomes Reveal New Aspects of the Virus-Host Cell Interface

abstract:
  Major gaps in our knowledge of pathogen genes and how these gene products interact with host gene products to cause disease represent a major obstacle to progress in vaccine and antiviral drug development for the herpesviruses. To begin to bridge these gaps, we conducted a dual analysis of Murine Cytomegalovirus (MCMV) and host cell transcriptomes during lytic infection. We analyzed the MCMV transcriptome during lytic infection using both classical cDNA cloning and sequencing of viral transcripts and next generation sequencing of transcripts (RNA-Seq). We also investigated the host transcriptome using RNA-Seq combined with differential gene expression analysis, biological pathway analysis, and gene ontology analysis. We identify numerous novel spliced and unspliced transcripts of MCMV. Unexpectedly, the most abundantly transcribed viral genes are of unknown function. We found that the most abundant viral transcript, recently identified as a noncoding RNA regulating cellular microRNAs, also codes for a novel protein. To our knowledge, this is the first viral transcript that functions both as a noncoding RNA and an mRNA. We also report that lytic infection elicits a profound cellular response in fibroblasts. Highly upregulated and induced host genes included those involved in inflammation and immunity, but also many unexpected transcription factors and host genes related to development and differentiation. Many top downregulated and repressed genes are associated with functions whose roles in infection are obscure, including host long intergenic noncoding RNAs, antisense RNAs or small nucleolar RNAs. Correspondingly, many differentially expressed genes cluster in biological pathways that may shed new light on cytomegalovirus pathogenesis. Together, these findings provide new insights into the molecular warfare at the virus-host interface and suggest new areas of research to advance the understanding and treatment of cytomegalovirus-associated diseases.


id: cord-351952-lhhjax3s
author: Pickering, Suzanne
date: 2020-09-24
url: https://doi.org/10.1371/journal.ppat.1008817
doi: 10.1371/journal.ppat.1008817
title:
  Comparative assessment of multiple COVID-19 serological technologies supports continued evaluation of point-of-care lateral flow assays in hospital and community healthcare settings

abstract:
  There is a clear requirement for an accurate SARS-CoV-2 antibody test, both as a complement to existing diagnostic capabilities and for determining community seroprevalence. We therefore evaluated the performance of a variety of antibody testing technologies and their potential use as diagnostic tools. Highly specific in-house ELISAs were developed for the detection of anti-spike (S), -receptor binding domain (RBD) and -nucleocapsid (N) antibodies and used for the cross-comparison of ten commercial serological assays—a chemiluminescence-based platform, two ELISAs and seven colloidal gold lateral flow immunoassays (LFIAs)—on an identical panel of 110 SARS-CoV-2-positive samples and 50 pre-pandemic negatives. There was a wide variation in the performance of the different platforms, with specificity ranging from 82% to 100%, and overall sensitivity from 60.9% to 87.3%. However, the head-to-head comparison of multiple sero-diagnostic assays on identical sample sets revealed that performance is highly dependent on the time of sampling, with sensitivities of over 95% seen in several tests when assessing samples from more than 20 days post onset of symptoms. Furthermore, these analyses identified clear outlying samples that were negative in all tests, but were later shown to be from individuals with mildest disease presentation. Rigorous comparison of antibody testing platforms will inform the deployment of point-of-care technologies in healthcare settings and their use in the monitoring of SARS-CoV-2 infections.


id: cord-353337-o302vxqm
author: Visser, Linda J.
date: 2020-07-15
txt: ./txt/cord-353337-o302vxqm.txt
cache: ./cache/cord-353337-o302vxqm.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/32667958/
doi: 10.1371/journal.ppat.1008702
summary:
  To study the effects of L pro on the induction of type I IFN in picornavirus-infected cells, we used two previously generated recombinant viruses; EMCV-L Zn , which contains inactivating mutations in the zinc-finger domain of the Leader (i.e. EMCV''s RLR signaling antagonist) [1, 43, 44] , and EMCV-L pro , which was derived from EMCV-L Zn and additionally encodes FMDV L pro at the N-terminus of its polyprotein ( Fig 1A) [45]. Cleavage of RLR signaling proteins by FMDV L pro correlates with type I IFN suppression degradation of NF-κB p65 and DUB activity as well as to impair L pro ''s ability to reduce IFN-β mRNA expression [36, 39] , also affected L pro ''s ability to cleave MAVS and TBK1. Notably, infection with EMCV L pro L143A, which displayed wt deISGylase/DUB activity but is strongly impaired in its ability to cleave/degrade RLR signaling proteins MAVS, TBK1 and NFκB p65, failed to suppress the induction of IFN-β mRNA.

title:
  Dissecting distinct proteolytic activities of FMDV L(pro) implicates cleavage and degradation of RLR signaling proteins, not its deISGylase/DUB activity, in type I interferon suppression

abstract:
  The type I interferon response is an important innate antiviral pathway. Recognition of viral RNA by RIG-I-like receptors (RLRs) activates a signaling cascade that leads to type I interferon (IFN-α/β) gene transcription. Multiple proteins in this signaling pathway (e.g. RIG-I, MDA5, MAVS, TBK1, IRF3) are regulated by (de)ubiquitination events. Most viruses have evolved mechanisms to counter this antiviral response. The leader protease (L(pro)) of foot-and-mouth-disease virus (FMDV) has been recognized to reduce IFN-α/β gene transcription; however, the exact mechanism is unknown. The proteolytic activity of L(pro) is vital for releasing itself from the viral polyprotein and for cleaving and degrading specific host cell proteins, such as eIF4G and NF-κB. In addition, L(pro) has been demonstrated to have deubiquitination/deISGylation activity. L(pro)’s deubiquitination/deISGylation activity and the cleavage/degradation of signaling proteins have both been postulated to be important for reduced IFN-α/β gene transcription. Here, we demonstrate that TBK1, the kinase that phosphorylates and activates the transcription factor IRF3, is cleaved by L(pro) in FMDV-infected cells as well as in cells infected with a recombinant EMCV expressing L(pro). In vitro cleavage experiments revealed that L(pro) cleaves TBK1 at residues 692–694. We also observed cleavage of MAVS in HeLa cells infected with EMCV-L(pro), but only observed decreasing levels of MAVS in FMDV-infected porcine LFPK αVβ6 cells. We set out to dissect L(pro)’s ability to cleave RLR signaling proteins from its deubiquitination/deISGylation activity to determine their relative contributions to the reduction of IFN-α/β gene transcription. The introduction of specific mutations, of which several were based on the recently published structure of L(pro) in complex with ISG15, allowed us to identify specific amino acid substitutions that separate the different proteolytic activities of L(pro). Characterization of the effects of these mutations revealed that L(pro)’s ability to cleave RLR signaling proteins but not its deubiquitination/deISGylation activity correlates with the reduced IFN-β gene transcription.


id: cord-353353-njvalb44
author: Lau, Susanna K. P.
date: 2016-10-13
txt: ./txt/cord-353353-njvalb44.txt
cache: ./cache/cord-353353-njvalb44.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/27737017/
doi: 10.1371/journal.ppat.1005911
summary:
  Analysis of 13 complete genome sequences showed that "Rosavirus B" and "Rosavirus C" represent two potentially novel picornavirus species infecting different rodents. Rosavirus C isolated from 3T3 cells causes multisystemic diseases in a mouse model, with high viral loads and positive viral antigen expression in organs of infected mice after oral or intracerebral inoculation. Rosavirus C isolated from cell culture causes multisystemic diseases in a mouse model, with histopathological changes and positive viral antigen expression in lungs and liver of infected mice. A total of 13 complete genomes from samples of four different wild rodent species (chestnut spiny rat, Coxing''s white-bellied rat, roof rat and Indochinese forest rat) positive for "Rosavirus C" and one street rodent species (Norway rat) positive for "Rosavirus B" were sequenced directly from the positive respiratory or alimentary samples and characterized. Infected cell lines and tissues from challenged mice that were tested positive for rosavirus C RASM14A by RT-PCR were subject to viral load studies and immunohistochemical staining for viral VP1 protein as described previously [28, 69] .

title:
  Identification of Novel Rosavirus Species That Infects Diverse Rodent Species and Causes Multisystemic Dissemination in Mouse Model

abstract:
  While novel picornaviruses are being discovered in rodents, their host range and pathogenicity are largely unknown. We identified two novel picornaviruses, rosavirus B from the street rat, Norway rat, and rosavirus C from five different wild rat species (chestnut spiny rat, greater bandicoot rat, Indochinese forest rat, roof rat and Coxing's white-bellied rat) in China. Analysis of 13 complete genome sequences showed that “Rosavirus B” and “Rosavirus C” represent two potentially novel picornavirus species infecting different rodents. Though being most closely related to rosavirus A, rosavirus B and C possessed distinct protease cleavage sites and variations in Yn-Xm-AUG sequence in 5’UTR and myristylation site in VP4. Anti-rosavirus B VP1 antibodies were detected in Norway rats, whereas anti-rosavirus C VP1 and neutralizing antibodies were detected in Indochinese forest rats and Coxing's white-bellied rats. While the highest prevalence was observed in Coxing's white-bellied rats by RT-PCR, the detection of rosavirus C from different rat species suggests potential interspecies transmission. Rosavirus C isolated from 3T3 cells causes multisystemic diseases in a mouse model, with high viral loads and positive viral antigen expression in organs of infected mice after oral or intracerebral inoculation. Histological examination revealed alveolar fluid exudation, interstitial infiltration, alveolar fluid exudate and wall thickening in lungs, and hepatocyte degeneration and lymphocytic/monocytic inflammatory infiltrates with giant cell formation in liver sections of sacrificed mice. Since rosavirus A2 has been detected in fecal samples of children, further studies should elucidate the pathogenicity and emergence potential of different rosaviruses.


id: cord-353703-u86ggw11
author: Gao, Peng
date: 2019-11-18
url: https://www.ncbi.nlm.nih.gov/pubmed/31738790/
doi: 10.1371/journal.ppat.1008169
title:
  Reprogramming the unfolded protein response for replication by porcine reproductive and respiratory syndrome virus

abstract:
  The unfolded protein response (UPR) in the endoplasmic reticulum (ER) constitutes a critical component of host innate immunity against microbial infections. In this report, we show that porcine reproductive and respiratory syndrome virus (PRRSV) utilizes the UPR machinery for its own benefit. We provide evidence that the virus targets the UPR central regulator GRP78 for proteasomal degradation via a mechanism that requires viral glycoprotein GP2a, while both IRE1-XBP1s and PERK-eIF2α-ATF4 signaling branches of the UPR are turned on at early stage of infection. The activated effector XBP1s was found to enter the nucleus, but ATF4 was unexpectedly diverted to cytoplasmic viral replication complexes by means of nonstructural proteins nsp2/3 to promote viral RNA synthesis. RNAi knockdown of either ATF4 or XBP1s dramatically attenuated virus titers, while overexpression caused increases. These observations reveal attractive host targets (e.g., ATF4 and XBP1s) for antiviral drugs and have implications in vaccine development.


id: cord-355610-7xy4s483
author: Hu, Dan
date: 2019-06-26
txt: ./txt/cord-355610-7xy4s483.txt
cache: ./cache/cord-355610-7xy4s483.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/31242272/
doi: 10.1371/journal.ppat.1007836
summary:
  Here, by using sequential antigen panning of a yeast antibody library derived from healthy donors against the DENV envelop protein domain III (DIII) combined with depletion by an entry defective DIII mutant, we identified a cross-reactive human monoclonal antibody (mAb), m366.6, which bound with high affinity to DENV DIII from all four DENV serotypes. Furthermore, as the first germline-like mAb derived from a naïve antibody library that could neutralize all four DENV serotypes, the m366.6 can be a tool for exploring mechanisms of DENV infection, and is a promising therapeutic candidate. Its fully human origin, the germline-like nature, combined with high-affinity and broad neutralizing activity toward all DENV serotypes, suggest that m366.6 is a promising candidate antiviral agent and may also provide a unique template for designing effective dengue vaccine immunogens. Two antibodies, designated as m360 and m366, bound potently to DENV DIIIs. Their scFv gene were fused with human IgG1 Fc for protein expression, and surface plasmon resonance (SPR) experiments were used to evaluate the antigens binding.

title:
  A broadly neutralizing germline-like human monoclonal antibody against dengue virus envelope domain III

abstract:
  Dengue is the most widespread vector-borne viral disease caused by dengue virus (DENV) for which there are no safe, effective drugs approved for clinical use. Here, by using sequential antigen panning of a yeast antibody library derived from healthy donors against the DENV envelop protein domain III (DIII) combined with depletion by an entry defective DIII mutant, we identified a cross-reactive human monoclonal antibody (mAb), m366.6, which bound with high affinity to DENV DIII from all four DENV serotypes. Immunogenetic analysis indicated that m366.6 is a germline-like mAb with very few somatic mutations from the closest VH and Vλ germline genes. Importantly, we demonstrated that it potently neutralized DENV both in vitro and in the mouse models of DENV infection without detectable antibody-dependent enhancement (ADE) effect. The epitope of m366.6 was mapped to the highly conserved regions on DIII, which may guide the design of effective dengue vaccine immunogens. Furthermore, as the first germline-like mAb derived from a naïve antibody library that could neutralize all four DENV serotypes, the m366.6 can be a tool for exploring mechanisms of DENV infection, and is a promising therapeutic candidate.


id: cord-355839-o0m71kvw
author: Sedeyn, Koen
date: 2019-10-17
txt: ./txt/cord-355839-o0m71kvw.txt
cache: ./cache/cord-355839-o0m71kvw.txt
extension: .txt
mime: text/plain
url: https://www.ncbi.nlm.nih.gov/pubmed/31622448/
doi: 10.1371/journal.ppat.1007984
summary:
  ATF2, activating transcription factor 2; CDS, cytoplasmic DNA sensor; ER, endoplasmic reticulum; IFN, interferon; IKK, inhibitor of nuclear factor kappa-B kinase; IKKε, inhibitor of nuclear factor kappa-B kinase subunit epsilon; IRF3, interferon regulatory factor 3; IRF7, interferon regulatory factor 7; JNK, c-Jun N-terminal kinase; MAPK, mitogen-activated protein kinase; MAVS, mitochondrial antiviral-signaling protein; MDA5, melanoma differentiation-associated protein 5; MyD88, myeloid differentiation primary response protein MyD88; NF-kB, nuclear factor-kappa B; NOD2, nucleotide-binding oligomerization domain-containing protein 2; PAMP, pathogenassociated molecular pattern; PRR, pattern recognition receptor; RIG, retinoic-acid-inducible gene-I; RLR, RIG-I-like receptor; RSV, respiratory syncytial virus; STING, stimulator of interferon protein; TBK1, tank binding kinase 1; TICAM1, toll/interleukin-1 receptor domain-containing adapter molecule 1; TIRAP, toll/ interleukin-1 receptor domain-containing adapter protein; TLR, toll-like receptor; TRAF3, tumor necrosis factor receptor-associated factor 3; TRAF6, tumor necrosis factor receptor-associated factor 6; TRAM, toll-like receptor adaptor molecule.

title:
  Respiratory syncytial virus nonstructural proteins 1 and 2: Exceptional disrupters of innate immune responses

abstract:
  Human respiratory syncytial virus (RSV) is the most important cause of acute lower respiratory tract disease in infants worldwide. As a first line of defense against respiratory infections, innate immune responses, including the production of type I and III interferons (IFNs), play an important role. Upon infection with RSV, multiple pattern recognition receptors (PRRs) can recognize RSV-derived pathogen-associated molecular patterns (PAMPs) and mount innate immune responses. Retinoic-acid-inducible gene-I (RIG-I) and nucleotide-binding oligomerization domain-containing protein 2 (NOD2) have been identified as important innate receptors to mount type I IFNs during RSV infection. However, type I IFN levels remain surprisingly low during RSV infection despite strong viral replication. The poor induction of type I IFNs can be attributed to the cooperative activity of 2 unique, nonstructural (NS) proteins of RSV, i.e., NS1 and NS2. These viral proteins have been shown to suppress both the production and signaling of type I and III IFNs by counteracting a plethora of key host innate signaling proteins. Moreover, increasing numbers of IFN-stimulated genes (ISGs) are being identified as targets of the NS proteins in recent years, highlighting an underexplored protein family in the identification of NS target proteins. To understand the diverse effector functions of NS1 and NS2, Goswami and colleagues proposed the hypothesis of the NS degradasome (NSD) complex, a multiprotein complex made up of, at least, NS1 and NS2. Furthermore, the crystal structure of NS1 was resolved recently and, remarkably, identified NS1 as a structural paralogue of the RSV matrix protein. Unfortunately, no structural data on NS2 have been published so far. In this review, we briefly describe the PRRs that mount innate immune responses upon RSV infection and provide an overview of the various effector functions of NS1 and NS2. Furthermore, we discuss the ubiquitination effector functions of NS1 and NS2, which are in line with the hypothesis that the NSD shares features with the canonical 26S proteasome.


id: cord-356115-vblgotjn
title: Functional and Genetic Analysis of Coronavirus Replicase-Transcriptase Proteins
author: Sawicki, Stanley G
date: 2005-12-09
url: https://www.ncbi.nlm.nih.gov/pubmed/16341254/
doi: 10.1371/journal.ppat.0010039
abstract:
  The coronavirus replicase-transcriptase complex is an assembly of viral and cellular proteins that mediate the synthesis of genome and subgenome-sized mRNAs in the virus-infected cell. Here, we report a genetic and functional analysis of 19 temperature-sensitive (ts) mutants of Murine hepatitis virus MHV-A59 that are unable to synthesize viral RNA when the infection is initiated and maintained at the non-permissive temperature. Both classical and biochemical complementation analysis leads us to predict that the majority of MHV-A59 ORF1a replicase gene products (non-structural proteins nsp1–nsp11) form a single complementation group (cistron1) while the replicase gene products encoded in ORF1b (non-structural proteins nsp12–nsp16) are able to function in trans and comprise at least three, and possibly five, further complementation groups (cistrons II–VI). Also, we have identified mutations in the non-structural proteins nsp 4, nsp5, nsp10, nsp12, nsp14, and nsp16 that are responsible for the ts phenotype of eight MHV-A59 mutants, which allows us to conclude that these proteins are essential for the assembly of a functional replicase-transcriptase complex. Finally, our analysis of viral RNA synthesis in ts mutant virus-infected cells allows us to discriminate three phenotypes with regard to the inability of specific mutants to synthesize viral RNA at the non-permissive temperature. Mutant LA ts6 appeared to be defective in continuing negative-strand synthesis, mutant Alb ts16 appeared to form negative strands but these were not utilized for positive-strand RNA synthesis, and mutant Alb ts22 was defective in the elongation of both positive- and negative-strand RNA. On the basis of these results, we propose a model that describes a pathway for viral RNA synthesis in MHV-A59-infected cells. Further biochemical analysis of these mutants should allow us to identify intermediates in this pathway and elucidate the precise function(s) of the viral replicase proteins involved.


