key: cord-1022818-rtpsji0c
authors: Zhao, Zhuangzhuang; Qin, Pan; Huang, Yao-Wei
title: Lysosomal ion channels involved in cellular entry and uncoating of enveloped viruses: Implications for therapeutic strategies against SARS-CoV-2
date: 2021-01-23
journal: Cell Calcium
DOI: 10.1016/j.ceca.2021.102360
sha: e832c19305ef73e5c8440b8829e961b5f9b185fe
doc_id: 1022818
cord_uid: rtpsji0c

Ion channels are necessary for correct lysosomal function including degradation of cargoes originating from endocytosis. Almost all enveloped viruses, including coronaviruses (CoVs), enter host cells via endocytosis, and do not escape endosomal compartments into the cytoplasm (via fusion with the endolysosomal membrane) unless the virus-encoded envelope proteins are cleaved by lysosomal proteases. With the ongoing outbreak of severe acute respiratory syndrome (SARS)-CoV-2, endolysosomal two-pore channels represent an exciting and emerging target for antiviral therapies. This review focuses on the latest knowledge of the effects of lysosomal ion channels on the cellular entry and uncoating of enveloped viruses, which may aid in development of novel therapies against emerging infectious diseases such as SARS-CoV-2.

Viruses are made up of highly condensed nucleic acids (RNA or DNA) surrounded by a protective protein coat. The presence or absence of a host-derived lipid membrane envelope features strongly in the taxonomic classification of viruses, and many important enveloped viruses have been widely studied including the families Coronaviridae, Filoviridae (Ebola virus; EBOV) and Orthomyxoviridae (influenza virus). The envelope of such viruses contains virus-encoded proteins that are essential for binding and entry into host cells. The coronavirus (CoV) spike (S) glycoprotein, consisting of S1 and S2 subunits, exists on the virion surface as a trimer [1, 2] . The envelope of EBOV contains a metastable trimer of glycoprotein (GP) obtained during budding from cells, which is the primary determinant of viral entry [3] . Some viruses even have both "naked" and enveloped forms, such as hepatitis E virus (HEV; family Hepeviridae) and hepatitis A virus (HAV; family Picornaviridae) [4, 5] ; these "quasi-enveloped" viruses enter cells in a way similar to that of enveloped viruses. For example, HAV particles cloaked in host membranes can enter cells through endocytosis, where enzymatic degradation in the late endosomes leads to uncoating [6] . In certain conditions, infection of the jejunum and ileum may be facilitated by digestive enzymes in the intestinal lumen, wherein enveloped viruses can fuse with host cell membranes directly, releasing the genetic material into the cytoplasm [7, 8] . However, under normal conditions, almost all enveloped viruses enter host cells via endocytosis. Further study of the endolysosomal cues that trigger cellular entry and uncoating of enveloped viruses is essential for development of broad-spectrum antiviral strategies against such emerging pathogens as SARS-CoV-2.

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In order for an enveloped virus to establish a productive infection, it must overcome cellular barriers to deliver its genetic materials to the cytoplasm. The process of viral entry includes viral attachment to cells, intracellular trafficking, and delivery of the viral genome. Almost all enveloped viruses enter host cells via specific interactions with receptor proteins which trigger endocytosis pathways such as macro-or micropinocytosis, or by induction of clathrin coat formation [9] [10] [11] [12] . Once enveloped viruses are taken up, they are transported by the vesicular system via membrane trafficking and processed from early to late endosomes, with a gradual drop in pH, Rab-switching, transport to perinuclear regions, and eventually routed towards degradative organelles known as lysosomes (see Fig. 1 ). When a virus reaches an appropriate point of the pathway, viral envelope fusion with the endolysosomal membrane will create a fusion pore within the endosomal membrane, allowing its genome to be released into the cytosol for downstream viral replication processes. This entire process of viral uncoating also requires assistance from cellular factors [13] .

Uncoating is an obligatory second step for virus infection, which makes it an attractive antiviral target. Viral envelope fusion with the endolysosomal membrane depends on cellular factors, such as intracellular receptors. In the same way that viruses need cell surface receptors to bind to cells, inside the endosome, viruses also require an intracellular receptor to bind and trigger membrane fusion. All known filoviruses employ NPC1 (NPC intracellular cholesterol transporter 1) [14] [15] [16] , and Lassa virus uses lamp1 (lysosomal associated membrane protein 1) [17] as an internal receptor to trigger uncoating (see Fig. 1 ). Other cellular factors such as the lysosomal TPC2 (two-pore channel 2) induce the fusion J o u r n a l P r e -p r o o f process and release of viruses from the endolysosomal compartments to the cytoplasm [18] . TPC2 has also been shown to be required for release of the SARS-CoV-2 genome into target cells [19] , although the specific role of TPC2 in virus escape into the cytoplasm is not completely clear [20] [21] [22] [23] . Another lysosomal ion channel, TRPML2 (the second member of the mammalian mucolipin TRP channel subfamily), can affect viral entry by enhancing the efficiency of viral trafficking in the endosomal system [24] . The following sections will go into greater detail on the endolysosomal environment needed for cellular entry and uncoating of enveloped viruses (see the summary in Table 1 ).

Lysosomes are the cell's degradation and recycling center, where macromolecules are degraded by hydrolytic enzymes activated by low pH [25] . Lysosomes are known to be a dynamic hub for cellular metabolism, nutrient sensing, plasma membrane repair, secretion, and spatiotemporal intracellular signaling [26] [27] [28] . These vital cellular functions hinge on the precise control of ionic (Na + , K + , Ca 2+ , Cl − , H + ) gradients [29] . One can speculate that the coming years will bring many more examples of direct relationships between ion flux and endocytic function [30] . Lysosomes are the end-point of endocytosis, fed by the process of endosome maturation, and an understanding of the lysosomal ion environment will help to shed light on the entry process of enveloped viruses.

The ion environment within lysosomes is difficult to assess by classical techniques such as patch clamp analysis due to their internal J o u r n a l P r e -p r o o f localization and small size. A technique called lysosomal patch clamp was developed for direct recording from isolated endolysosomes which were pharmacologically enlarged using vacuolin-1 [31, 32] . This technique paved the way to study endolysosomal channels in their native membranes. A variety of lysosomal ion channels have been identified, such as two-pore channels (TPCs), TRPMLs and transmembrane protein 175 (TMEM175), though many channels have yet to be characterized, such as the unidentified H + 'leak' channel which may affect lysosomal pH via nutrient-sensitive cellular cues [33] .

Lysosomes are characterized by an acidic lumen pH, which can reach as low as 4.6 [34] . This acidic environment can be attributed to a proton pump V-ATPase complex, which pumps H + from the cytoplasm into the endolysosomal lumen in an energy-dependent manner. The V-ATPase complex has been identified as a critical host factor for viral entry [35, 36] . Inhibitors of V-ATPase such as bafilomycin A or other luminal pH-related lysosomotropic agents ammonium chloride (NH4Cl) and chloroquine (which accumulate in acidic organelles such as endosomes and lysosomes and neutralize their pH) [37] display a wide-spectrum of antiviral effects against influenza and CoVs [19, [38] [39] [40] .

The role of H + in viral infection is closely related to the activities of endolysosomal proteases, which are required for cleavage of the viral S glycoprotein [41] . In addition to protease activities, a low pH also induces conformational changes in the viral fusion loop [42, 43] , which inserts into the endosomal membrane, pulling EBOV and endolysosomal membranes together [44] .

Lysosomes are highly enriched in chloride ions (Cl -), and the luminal [Cl -] can reach 60-80 mM [45] , maintained by the voltage-gated J o u r n a l P r e -p r o o f chloride channel 7 (CLC7) [46, 47] . As Clis the antagonistic ion of H + and other positively charged ions, Clchannels help regulate lysosomal pH levels [46] , lysosomal enzyme activity [45] , Ca 2+ release from the lysosome, and cell volume. Tamoxifen and NPPB, two Clchannel regulators, have been found to block viral entry by inhibiting viral binding and penetration and by disrupting Ca 2+ homeostasis [48] . Chloride intracellular channel 1 (CLIC1) has been shown to be required for regulation of endolysosomal pH, and silencing of CLIC1 decreased infection by hepatitis C virus (HCV) [49] . This may be due to the fact that an acidic late endosome/lysosome pH is crucial for uncoating of HCV. Since the acidic endolysosomal pH is crucial for induction of viral membrane fusion to allow genome release, lysosomal Clchannels are a potential target for antiviral strategies against a broad-spectrum of endocytic pathogens including SARS-CoV-2.

The lysosome lumen is generally thought to be high in K + and low in Na + [50, 51] , suggesting a lack of a Na + or K + concentration gradient across the lysosomal membrane. However, a recent study has challenged this view by showing the isolated lysosome fractions were low in [K + ] and high in [Na + ] in the lysosomal lumen [52] . Recent whole-endolysosome patch-clamp studies showing the presence of multiple Na + -and K +selective channels in the lysosome [52] [53] [54] [55] . The lysosomal Na + and K + gradients control various lysosomal functions, including lysosomal acidification and catabolite export [53, 56] . K + was identified as a biochemical cue to activate the viral entry process, and K + channel inhibition can alter the distribution of K + across the endosomal system and arrest virus trafficking in endosomes [57] .

Recent studies have identified two lysosomal K + channels: TMEM175 and large conductance Ca 2+ -activated K + (BK) channel [53, 55, 58] .

J o u r n a l P r e -p r o o f TMEM175 is responsible for K + conductance in endosomes and lysosomes [54] , which plays important roles in setting the lysosomal membrane potential (Δψ, defined as Vcytosol-Vlumen; Vlumen is set to 0 mV [52, 54] and maintaining pH stability. BK channels regulate Δψ of endolysosomes in both excitable and non-excitable cells [53, 55] . TMEM175 and BK likely affect viral entry. A recent study showed that modulators targeting BK channels had no effect on Bunyavirus [59] , but the modulator (Tram 34) used in the study is actually a potent blocker of the intermediateconductance Ca 2+ -activated K + (IKCa) channel rather than the BK channel. Thus, the conclusion that BK channels are not involved in Bunyavirus infection is not supported, and whether the BK channel in lysosomes is a potential antiviral target for SARS-CoV-2 or other enveloped viruses still lacks experimental evidence. Quinine and tetraethylammonium, which inhibit Bunyavirus, could not inhibit TMEM175 [57] , although whether TMEM175 affects SARS-CoV-2 infection is still lacking experimental evidence.

Other lysosomal potassium channels: weakly inwardly rectifying K + channel 2 (TWIK2) is a K2P channel expressed in lysosomes. TWIK2 generates functional background K + currents in the endolysosomes, and its expression affects the number and mean size of the lysosomes [60] .

Kcnk6 (encoding TWIK2) and other K2P channels are involved in Bunyavirus intrusion [59] . K2P channels in viral infection are thought to influence virus endosomal trafficking, as their inhibition can alter the distribution of K + across the endosomal system and arrest virus trafficking in endosomes [57] . In addition to influencing virus endosomal trafficking, K2P channels also effect conformational changes in the viral fusion loop. As mentioned above, K2P channels are involved in endosomal K + accumulation, and some studies have shown that exposure of Hazara virus to elevated K + causes dramatic structural changes in Hazara virus glycoprotein spikes, promotes spike-membrane interactions, and expedites infectivity [61] , although the mechanism of this effect needs further study.

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The lysosome lumen is high in [Na + ], although the ion transporters that establish the lysosomal Na + gradient are not yet known [62] . High [Na + ] pretreatment can increase the infectivity of Bunyavirus but not influenza A virus [57, 63] . TPCs, which have been shown to be Na +selective cation channels in lysosomal electrophysiological analyses [52, 64, 65] , were identified as essential for the release of EBOV's genome into the cytoplasm. Inhibition of TPCs prevents the fusion of EBOV with lysosomes [18] , though their activation mechanisms and ion selectivities are areas of active investigation [66, 67] . Although lysosomal patch clamp studies have suggested that TPCs are sodium channels, many studies have shown them to be required for NAADP (nicotinic acid adenine dinucleotide phosphate)-mediated Ca 2+ signaling [52, 68, 69] .

A recent study showed that the ion selectivity of TPC2 in endo-lysosomes is instead a mutable property that depends on the nature of the activating agonist [70] , which may help resolve conflicts in its cation selectivity. Whether Na + selectivity of TPC2 is involved in virus invasion still needs further study.

As the lysosome is an intracellular Ca 2+ store, with a lumenal [Ca 2+ ] that can reach 0.5 mM, maintained by a putative Ca 2+ /H + exchanger [71] .

Ca 2+ efflux from endosomes and lysosomes is thought to be important for intracellular signaling, organelle acidification and control of many cellular mechanisms including vesicle transport and fusion of lysosomes with late endosomes [72] [73] [74] [75] . Calcium sensing proteins including calmodulin and Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) have been identified as important for cell entry of EBOV [76] , regulated by lysosomal Ca 2+ flux [77] . NAADP is the most potent of the established Ca 2+ -mobilizing messengers, and it stimulates intracellular calcium J o u r n a l P r e -p r o o f channels to release Ca 2+ from endosomes and lysosomes [78] . NAADP-induced lysosomal Ca 2+ flux is required for EBOV [18] and Middle East respiratory syndrome (MERS)-CoV [79] . Thus, compounds that inhibit NAADP-evoked Ca 2+ release should be studied as potential inhibitors of SARS-CoV-2.

TPCs have been shown to be required for NAADP-mediated Ca 2+ signaling in many studies [69] . TPCs are dimeric ion channels composed of a duplicated domain architecture, and are likely an evolutionary bridge to four-domain voltage-gated Ca 2+ and Na + channels [80] [81] [82] . NAADP appears to interact with TPCs indirectly through putative binding proteins [83] [84] [85] [86] . In contrast, the endo-lysosomal lipid phosphatidylinositol-3,5-bisphosphate [PI (3, 5 )P2] has emerged as a direct channel activator that binds within the first domain of TPCs [52, 87, 88] . Filoviruses require TPC function for release of the viral genome into the host cell [18] , and a pharmacological block of TPCs by trans-Ned-19 (NED19) or other calcium channel blockers inhibits viral infectivity [18] .

The exact role of TPCs in viral infection is not clear. Knockdown of TPC1 (biased endosomally) or TPC2 (biased lysosomally) decreased the activity of furin, a protease which facilitates MERS-CoV fusion with cellular membranes [79] . In addition to protease activities, the TPCspecific blocker tetrandrine could prevent the capsid disassembly and nuclear transport required for successful virus entry [89] . The efficacy of tetrandrine is related to the inhibition of viral infection, possibly by preventing viral-endosome membrane fusion from within TPC2-positive structures [18] , or by interfering with a TPC2-driven late endosome-lysosome maturation process [20] . Finally, membrane fusion can occur not only between different intracellular compartments, but also between lipid-bound structures such as viral particles and cellular membranes, and J o u r n a l P r e -p r o o f they may share common principles [90] . Research has shown that TPC1 is required for formation of contact sites between the ER and endosome, and this was associated with disruptions in late endosome and lysosome morphology [91] , which may help explain why TPCs affect virusendosomal fusion.

A recent study showed that blocking TPC channels by inhibiting PI (3, 5) P2 formation led to a pronounced depletion of plasmalemmal Mac-1, which was instead trapped in endomembrane vacuoles [92] . Angiotensin-converting enzyme 2 (ACE2), the primary receptor for SARS-CoV-2, is also a plasmalemmal protein. It is foreseeable that blocking TPC channels could deplete plasmalemmal ACE2 and reduce viral binding, thus affecting the endocytosis of SARS-CoV-2, although this does not seem to be the main reason for TPC's effect on viral infection. Blockage of TPC channels can block virus-endosome membrane fusion and viral gene release to the cytoplasm as detected by a virus contents release assay [18] .

Apart from TPCs, the mucolipin subfamily of transient receptor potential (TRP) cation channels (TRPMLs), which consist of TRPML1, TRPML2 and TRPML3 (a.k.a. MCOLN1-3) are Ca 2+ -permeable cation channels expressed in the membranes of endosomes and lysosomes [65] .

Among these, TRPML2 can increase the infectivity of endocytosed viruses including influenza A virus, yellow fever virus, and Zika by promoting viral vesicular trafficking, resulting in increased endosomal escape [24] . Whether TRPML2 also increases SARS-CoV-2 infection is yet unknown; according to the NextBio Research Disease Atlas (http://www.nextbio.com), a strong downregulation of TRPML3 is associated with cytomegalovirus infection in humans. For TRPML1, although its endogenous agonist PI (3, 5) P2 is directly related to the entry of enveloped J o u r n a l P r e -p r o o f viruses [93] [94] [95] [96] , TRPML1 itself does not affect their entry, including SARS-CoV-2 [19, 79] . MLSA1, a TRPML1 agonist, also did not affect EBOV infection or rescue EBOV infection in PI(3,5)P2-defective cells [94] . Some studies have shown that rapamycin can increase autophagic flux in a TRPML1-dependent manner [97] , and rapamycin's ability to increase the autophagic flux is closely related to viral infectivity [98] [99] [100] , suggesting that TRPML1 might play an indirect role in antiviral processes.

PI(3,5)P2 is a late endosome/lysosome-specific phosphoinositide, produced by the lipid kinase PIKfyve in mammalian cells [91] . PI(3,5)P2 regulates membrane trafficking and activity of ion channels [91] , and its effectors are fewer compared to those of Phosphatidylinositol 3phosphate (PtdIns3P). The scarce effectors of PI(3,5)P2 thus far identified include TRPMLs and TPCs [52, 101] , all of which are directly or indirectly related to the entry of enveloped viruses.

Viruses that require deeper trafficking in the endocytic pathway to endo-lysosomes rely on PI(3,5)P2. Inhibition of PIKfyve and PI(3,5)P2 production by apilimod or YM201636 has been shown to inhibit viral fusion in late endosomes and lysosomes, for example in vaccinia virus [96] , African swine fever virus [102] , SARS-CoV-2 [19, 103] , and some filoviruses [93, 94] . The mechanism of inhibition is likely due to a defect in the maturation of endo-lysosomes, impairing traffic of incoming virus to sites where NPC1 resides and membrane fusion takes place [93] .

J o u r n a l P r e -p r o o f

Viruses have developed strategies to exploit host cell machinery and organelles to promote viral infection. Experiments have shown that entry of SARS-CoV-2 into host cells is mainly mediated by endocytosis, and is closely related to lysosomes [19, 22, 104, 105] . More specifically, TPC2 is critical for SARS-CoV-2 infection, which supports the notion that lysosomal ion channels may be targets for therapeutic strategies against SARS-CoV-2. Lysosomes are equipped with various ion channels, some of which have been proven to affect entry of specific enveloped viruses (see the summary in Table 1 ). There are many licensed pharmaceutical preparations that target ion channels, improving our understanding of the role of ion channels in viral pathogenesis may reveal their potential as targets of new, safe drugs with broad antiviral activity.

Although several lysosomal ion channels have been confirmed as potential novel targets for the treatment of viruses, several basic questions remain:

 What is the role of TPCs in viral uncoating?  How does NAADP induce calcium release from lysosomes, and what is the role for the NAADP-dependent Ca 2+ signaling in viral translocation?

 How do lysosomal ∆ and Ca 2+ regulate the process of membrane fusion between enveloped viruses and lysosomes?  Are there any other (known or unknown) lysosomal ion channel that may affect viral entry?

J o u r n a l P r e -p r o o f Further investigation is required to address these questions, which will help us gain a much better understanding of the functional relevance of these lysosomal ion channels. In addition, these investigations may discover potential new therapeutic lines in the fight against SARS-CoV-2.

The authors declare no conflict of interests. [57, 59, 60] J o u r n a l P r e -p r o o f

Structure, Function, and Evolution of Coronavirus Spike Proteins

Coronavirus Spike Protein and Tropism Changes

Ebolavirus glycoprotein structure and mechanism of entry

Characterization of the Quasi-Enveloped Hepatitis E Virus Particles Released by the Cellular Exosomal Pathway

A pathogenic picornavirus acquires an envelope by hijacking cellular membranes

Cellular entry and uncoating of naked and quasi-enveloped human hepatoviruses

Porcine deltacoronavirus enters cells via two pathways: A protease-mediated one at the cell surface and another facilitated by cathepsins in the endosome

Trypsin promotes porcine deltacoronavirus mediating cell-to-cell fusion in a cell type-dependent manner

Live cell imaging of viral entry

Endocytosis by random initiation and stabilization of clathrin-coated pits

Ebolavirus is internalized into host cells via macropinocytosis in a viral glycoprotein-dependent manner

Cellular entry of ebola virus involves uptake by a macropinocytosis-like mechanism and subsequent trafficking through early and late endosomes

Virus entry by endocytosis

Small molecule inhibitors reveal Niemann-Pick C1 is essential for Ebola virus infection

Ebola virus entry requires the cholesterol transporter Niemann-Pick C1

Ebola virus entry requires the host-programmed recognition of an intracellular receptor

Virus entry. Lassa virus entry requires a trigger-induced receptor switch

Ebola virus. Two-pore channels control Ebola virus host cell entry and are drug targets for disease treatment

Characterization of spike glycoprotein of SARS-CoV-2 on virus entry and its immune cross-reactivity with SARS-CoV

Ebolavirus Glycoprotein Directs Fusion through NPC1+ Endolysosomes

Could an endo-lysosomal ion channel be the Achilles heel of SARS-CoV2?

Could the Inhibition of Endo-Lysosomal Two-Pore Channels (TPCs) by the Natural Flavonoid Naringenin Represent an Option to Fight SARS-CoV-2 Infection?

Mining of Ebola virus entry inhibitors identifies approved drugs as two-pore channel pore blockers

Mucolipin-2 Cation Channel Increases Trafficking Efficiency of Endocytosed Viruses

Principles of lysosomal membrane digestion: stimulation of sphingolipid degradation by sphingolipid activator proteins and anionic lysosomal lipids

The Lysosome as a Regulatory Hub

The lysosome as a command-and-control center for cellular metabolism

Amino acids and mTORC1: from lysosomes to disease

Ion flux and the function of endosomes and lysosomes: pH is just the start: the flux of ions across endosomal membranes influences endosome function not only through regulation of the luminal pH

The type IV mucolipidosis-associated protein TRPML1 is an endolysosomal iron release channel

Activating mutations of the TRPML1 channel revealed by proline-scanning mutagenesis

The voltage-gated sodium channel TPC1 confers endolysosomal excitability

The position of lysosomes within the cell determines their luminal pH

Human host factors required for influenza virus replication

Requirement for vacuolar proton-ATPase activity during entry of influenza virus into cells

Akt inhibition promotes autophagy and sensitizes PTEN-null tumors to lysosomotropic agents

New insights into the antiviral effects of chloroquine

Chloroquine is a potent inhibitor of SARS coronavirus infection and spread

SARS coronavirus entry into host cells through a novel clathrin-and caveolae-independent endocytic pathway

Characterization of severe acute respiratory syndromeassociated coronavirus (SARS-CoV) spike glycoprotein-mediated viral entry

Structure and function of the complete internal fusion loop from Ebolavirus glycoprotein 2

Ebolavirus entry requires a compact hydrophobic fist at the tip of the fusion loop

Late-penetrating viruses

High lumenal chloride in the lysosome is critical for lysosome function

The Cl-/H+ antiporter ClC-7 is the primary chloride permeation pathway in lysosomes

CLC Chloride Channels and Transporters: Structure, Function, Physiology, and Disease

Inhibition of herpes simplex virus type 1 entry by chloride channel inhibitors tamoxifen and NPPB

Requirement for chloride channel function during the hepatitis C virus life cycle

Molecular mechanisms of endolysosomal Ca2+ signalling in health and disease

A cation counterflux supports lysosomal acidification

TPC Proteins Are Phosphoinositide-Activated Sodium-Selective Ion Channels in Endosomes and Lysosomes

A voltagedependent K(+) channel in the lysosome is required for refilling lysosomal Ca(2+) stores

TMEM175 Is an Organelle K(+) Channel Regulating Lysosomal Function

BK Channels Alleviate Lysosomal Storage Diseases by Providing Positive Feedback Regulation of Lysosomal Ca2+ Release

Activator SLC38A9 Is Required to Efflux Essential Amino Acids from Lysosomes and Use Protein as a Nutrient

Bunyavirus requirement for endosomal K+ reveals new roles of cellular ion channels during infection

Lysosomal Potassium Channels: Potential Roles in Lysosomal Function and Neurodegenerative Diseases

Modulation of Potassium Channels Inhibits Bunyavirus Infection

Recombinant tandem of pore-domains in a Weakly Inward rectifying K(+) channel 2 (TWIK2) forms active lysosomal channels

Potassium is a trigger for conformational change in the fusion spike of an enveloped RNA virus

Lysosomal Ion Channels as Decoders of Cellular Signals

Stepwise priming by acidic pH and a high K+ concentration is required for efficient uncoating of influenza A virus cores after penetration

From mucolipidosis type IV to Ebola: TRPML and two-pore channels at the crossroads of endo-lysosomal trafficking and disease

Questioning regulation of two-pore channels by NAADP, Messenger (Los Angel

Two-pore channels (TPCs): current controversies

Ren, mTOR regulates lysosomal ATP-sensitive two-pore Na(+) channels to adapt to metabolic state

NAADP mobilizes calcium from acidic organelles through two-pore channels

Agonist-mediated switching of ion selectivity in TPC2 differentially promotes lysosomal function

Release and uptake mechanisms of vesicular Ca(2+) stores

Calcium: a fundamental regulator of intracellular membrane fusion?

The role of calcium and other ions in sorting and delivery in the late endocytic pathway

Regulation of membrane trafficking by signalling on endosomal and lysosomal membranes

Lysosomal acidification mechanisms

Identification of novel cellular targets for therapeutic intervention against Ebola virus infection by siRNA screening

Lysosomal calcium regulates autophagy

NAADP-dependent Ca(2+) signaling regulates Middle East respiratory syndrome-coronavirus pseudovirus translocation through the endolysosomal system

Domain assembly of NAADP-gated two-pore channels

Two-pore channels provide insight into the evolution of voltage-gated Ca2+ and Na+ channels

Isolated pores dissected from human two-pore channel 2 are functional

Photoaffinity labeling of nicotinic acid adenine dinucleotide phosphate (NAADP) targets in mammalian cells

Photoaffinity labeling of high affinity nicotinic acid adenine dinucleotide phosphate (NAADP)-binding proteins in sea urchin egg

Nicotinic Acid Adenine Dinucleotide 2'-Phosphate (NAADP) Binding Proteins in T-Lymphocytes

The Molecular Basis for Ca(2+) Signalling by NAADP: Two-Pore Channels in a Complex?

Structural insights into the voltage and phospholipid activation of the mammalian TPC1 channel

Two-pore channels open up

Identification of Potassium and Calcium Channel Inhibitors as Modulators of Polyomavirus Endosomal Trafficking

Mechanisms of membrane fusion: disparate players and common principles

An Endosomal NAADP-Sensitive Two-Pore Ca(2+) Channel Regulates ER-Endosome Membrane Contact Sites to Control Growth Factor Signaling

Lipid-gated monovalent ion fluxes regulate endocytic traffic and support immune surveillance

The phosphatidylinositol-3-phosphate 5-kinase inhibitor apilimod blocks filoviral entry and infection

Ebola virus requires phosphatidylinositol (3,5) bisphosphate production for efficient viral entry

Redistribution of Endosomal Membranes to the African Swine Fever Virus Replication Site

Vaccinia Virus Infection Requires Maturation of Macropinosomes

Rapamycin directly activates lysosomal mucolipin TRP channels independent of mTOR

Rapamycin-induced autophagy restricts porcine epidemic diarrhea virus infectivity in porcine intestinal epithelial cells

Autophagy during viral infection -a double-edged sword

Autophagy and Viral Infection

PI(3,5)P(2) controls membrane trafficking by direct activation of mucolipin Ca(2+) release channels in the endolysosome

Endosomal maturation, Rab7 GTPase and phosphoinositides in African swine fever virus entry

Remdesivir and chloroquine effectively inhibit the recently emerged novel coronavirus (2019-nCoV) in vitro

The lysosome: A potential juncture between SARS-CoV-2 infectivity and Niemann-Pick disease type C, with therapeutic implications

This study was supported by the National Natural Science Foundation of China (32041003 and 31872488). The professional editing service NB Revisions was used for technical preparation of the text prior to submission.J o u r n a l P r e -p r o o f