pp900196 803..804 On the Inside Trichomes: The Source of Heavy Metal in Tobacco Smoke? Smoking of tobacco (Nicotiana taba- cum) leaves is one of the principal routes of exposure to heavy metals (Fig. 1). Little is known about the mechanisms of heavy-metal accumula- tion and detoxification in tobacco. Recently, it was shown that the tri- chomes of tobacco exposed to Cd21 and Ca21 produced calcium (Ca)/cadmium (Cd)-containing grains. Other effects of Cd exposure were a retardation of plant growth and a 2-fold increase of the number of trichomes in comparison with untreated plants. An increased concentration of Ca in the nutrient medium was found to have a protective effect toward Cd toxicity and enhanced the production of the grains. The Ca/ Cd-containing grains were 20 to 150 mm in diameter, and formed on the head cells of both the short and long tri- chomes of tobacco leaves. Thus, these studies revealed a new function of tobacco trichomes, the excretion of Cd in the form of particles. Sarret et al. (pp. 1021–1034) bring to bear an arsenal of cutting-edge techniques to examine the question of whether the trichomes of tobacco leaves also play a role in the responses of tobacco plants to toxic levels of zinc (Zn). Zn exposure re- sulted in toxicity signs in plants, and these damages were partly reduced by a Ca supplement. Confocal imaging of intracellular Zn using a fluorescent indicator showed that Zn was prefer- entially accumulated in trichomes. Exposure to Zn alone and Zn plus Ca increased the trichome density and induced the production of Ca/Zn min- eral grains on the head cells of tri- chomes. These grains were aggregates of submicrometer-sized crystals and poorly crystalline material, and con- tained Ca as major element. Micro x-ray diffraction revealed that the large majority of the grains were composed essentially of metal-substituted calcite (calcium carbonate). Thus, the produc- tion of Zn-containing biogenic calcite and other Zn compounds through the trichomes is a novel mechanism in- volved in Zn detoxification. This study also illustrates the potential of laterally resolved x-ray synchrotron radiation techniques to study biomineralization and metal homeostasis processes in plants. Membrane Lipid Saturation and Chilling Sensitivity: Surprising Results Some Arabidopsis (Arabidopsis thali- ana) mutants that exhibit decreased thylakoid unsaturation are substan- tially indistinguishable from wild type when grown at 22�C, but exhibit defects in biogenesis and maintenance of chlo- roplasts at temperatures below 5�C. A role for thylakoid unsaturation in main- taining photosynthetic function at low temperatures is also suggested by experiments in which transgenic ex- pression of fatty acid desaturases in chilling-sensitive plant species resulted in increased survival of plants at low temperatures. The Arabidopsis mutant fatty acid biosynthesis 1 (fab1) that is par- tially deficient in b-ketoacyl-synthase II (KAS2) activity exhibits a distinct low- temperature phenotype. This mutant contains increased levels of 16:0 due to a mutation in the KAS2 gene, which encodes the condensing enzyme that catalyzes the first step in elongation of 16:0 to 18:0 during fatty acid synthesis. In fab1 mutants, the 16:0 fraction of the thylakoid phospholipid phosphatidyl- glycerol (PG) is increased from 20% in wild type to 41% in fab1. This is signif- icant because increased 16:0 in PG, and more specifically the sum of 16:0 1 18:0 1 16:1, D3 trans (sometimes re- ferred to as high-melting-point fatty acids), has been correlated with chilling sensitivity through surveys of chilling- tolerant and chilling-sensitive plant species. Typically, plants containing more than 60% high-melting-point fatty acids in PG have been shown to be chilling sensitive. The fab1 mutant con- tains 69% high-melting-point fatty acids in PG—a higher percentage than is found in many chilling-sensitive plants. However, fab1 plants were completely unaffected, when compared with wild- type controls, by a range of chilling treatments that quickly killed other chilling-sensitive plants. Instead, fab1 plants are damaged only by long-term (.10 day) exposure to low temperature. Are the elevated levels of high-melting- point fatty acids in PG the direct cause of the damage and death of fab1 plants at 2�C? Surprisingly, the answer is no. Barkan et al. (pp. 1012–1020) have iso- lated suppressor mutations that rescue fab1 from death at low temperatures. One of the suppressors is an allele of fad5, a mutant that has decreased chlo- roplast 16:0 D7-desaturase and, hence, more saturated chloroplast membrane lipids. The overall leaf fatty acid com- position of the rescued line contained 31% 16:0 compared with 23% in fab1 and 17% in wild type. Based on the biophys- ical characteristics of saturated and unsaturated fatty acids, the increased 16:0 in fab1 fad5-2 plants would be expected to exacerbate, rather than ameliorate, low-temperature damage. The authors speculate that changes in the shape of other lipids may compen- sate for disruptive changes in the shape of PG molecules induced by the fab1 mutation, for example, by altering the packing relationships between the thylakoid lipids and membrane pro- teins of the photosynthetic complexes. Clearly, more needs to be learned about the relationship between thylakoid membrane saturation and chilling sen- sitivity. Figure 1. Heavy metal-excreting trichomes appear to be a major source of heavy-metal exposure from tobacco smoke. The painting is Woman with a Cigarette by Pablo Picasso. www.plantphysiol.org/cgi/doi/ 10.1104/pp.104.900196. Plant Physiology, July 2006, Vol. 141, pp. 803–804, www.plantphysiol.org � 2006 American Society of Plant Biologists 803 https://plantphysiol.orgDownloaded on April 5, 2021. - Published by Copyright (c) 2020 American Society of Plant Biologists. All rights reserved. https://plantphysiol.orgDownloaded on April 5, 2021. - Published by Copyright (c) 2020 American Society of Plant Biologists. All rights reserved. https://plantphysiol.orgDownloaded on April 5, 2021. - Published by Copyright (c) 2020 American Society of Plant Biologists. All rights reserved. https://plantphysiol.org https://plantphysiol.org https://plantphysiol.org Identification of a Phosphate- Overaccumulating Mutation The Arabidopsis mutant pho2, which is defective in inorganic phosphate (Pi) homeostasis, was identified by screen- ing the Pi content of shoots. Even though normal Pi concentrations are maintained in the roots of pho2 mu- tants, the shoots accumulate excessive amounts of Pi and exhibited Pi-toxic symptoms. Recently, it was reported that a microRNA, miR399, controls Pi homeostasis by regulating the expres- sion of a ubiquitin-conjugating E2 en- zyme (UBC24) in Arabidopsis. The accumulation of UBC24 mRNA was suppressed by the targeting of miR399, whose expression is up-regulated by Pi starvation. Intriguingly, several char- acteristics of Pi toxicity in the pho2 mutant are similar to those in miR399- overexpressing and UBC24 T-DNA knockout plants: both Pi uptake and translocation of Pi from roots to shoots is increased and Pi remobiliza- tion within leaves is impaired. In this issue, both Aung et al. (pp. 1000–1011) and Bari et al. (pp. 988–999) demon- strate that the Pi overaccumulator pho2 is caused by a single nucleotide muta- tion resulting in early termination within the UBC24 gene. No UBC24 protein was detected in the pho2 mu- tant, and the phenotype of the pho2 mutation could be rescued by intro- duction of a wild-type copy of UBC24. The combined results of these two research groups provide many further insights into the workings of this pecu- liar mutant. It was demonstrated by micrografting experiments that a pho2 root genotype is sufficient to yield leaf Pi accumulation, suggesting that Pi toxicity in this mutant arises from increased Pi uptake and translocation of Pi from roots to shoots. Furthermore, miR399 and UBC24 were colocalized in the vascular cylinder. Bari et al. present a working model for the mechanism of Pi sensing in higher plants in which PHR1, a MYB factor that has previously been shown to be required for induc- tion of a small number of genes under Pi starvation, takes a central role and the downstream miR399/PHO2 path- way regulates the expression of only a subset of the phosphate starvation- induced genes. The identification of putative PHO2 orthologs containing five miR399 binding sites in other higher plants and the demonstration of Pi-dependent miR399 expression in rice (Oryza sativa) suggest that this Pi starvation-signaling pathway may be highly conserved throughout the plant kingdom. Brassinosteroid Insensitivity Increases Rice Production The major factor underlying the suc- cess of the Green Revolution was the introduction of high-yielding semi- dwarf cultivars of wheat (Triticum aestivum) and rice. The dwarf pheno- types of the Green Revolution cultivars were largely traceable to disruptions in GA signaling or biosynthesis. Recently, however, another important target for producing high-yielding semidwarf cultivars was identified in barley (Hordeum vulgare). The semidwarf uzu phenotype of barley is brassinosteroid (BR) insensitive and is caused by a missense mutation in HvBRI1, an ortholog of the Arabidopsis gene BRASSINOSTEROID INSENSITIVE1 (BRI1). In contrast to barley, the loss- of-function mutants of a rice BRI1 ortholog (OsBRI1), namely, d61, show a range of phenotypes. Although the weak alleles d61-1 and d61-2 exhibit agronomically useful traits, such as semidwarf stature, erect leaves, and elongated neck internodes, they also exhibit, unfortunately, morphological alterations in their reproductive organs and reduced grain yield. Of nine d61 alleles identified by Morinaka et al. (pp. 924–931), the weakest, d61-7, con- fers agronomically important traits, such as semidwarf stature and erect leaves. The biomass produced by wild type was 38% higher than that of d61-7 at harvest under conventional planting density, whereas, as the situation was reversed at high planting densities, the biomass of d61-7 was 35% higher than that of wild type. However, the small grain size of d61-7 countered any in- crease in grain yield, leading to the same grain yield as that of wild type even at high density. The authors there- fore produced transgenic rice with par- tial suppression of endogenous OsBRI1 expression. Several of these transfor- mants, although of the same height as wild type, exhibited the desirable phe- notype of erect leaves. The estimated grain yield of these transformants was about 30% higher than that of wild type at high growth densities. These results demonstrate the feasibility of generating erect-leaved plants by modifying the expression of the BR receptor gene in transgenic rice plants. Peter V. Minorsky Department of Natural Sciences Mercy College Dobbs Ferry, New York 10522 804 Plant Physiol. Vol. 141, 2006 https://plantphysiol.orgDownloaded on April 5, 2021. - Published by Copyright (c) 2020 American Society of Plant Biologists. All rights reserved. https://plantphysiol.org CORRECTIONS Vol. 140: 30–48, 2006 Abdulrazzak N., Pollet B., Ehlting J., Larsen K., Asnaghi C., Ronseau S., Proux C., Erhardt M., Seltzer V., Renou J.-P., Ullman P., Pauly M., Lapierre C., and Werck-Reichhart D. A coumaroyl-ester-3-hydroxylase Insertion Mutant Reveals the Existence of Nonredundant meta-Hydroxylation Pathways and Essential Roles for Phenolic Precursors in Cell Expansion and Plant Growth. The authors regret that this article contains a description of immunofluorescence/confocal microscopy methodology that was not actually used for this work. In addition, this methodology was given without proper credit to Sugimoto et al. (K. Sugimoto, R.E. Williamson, G.O. Wasteneys [2000] Plant Physiol 124: 1493–1506), who developed the protocol. The correct immunofluorescence/confocal microscopy methodology used for this article is described below. The authors apologize for this error and any inconvenience it may have caused. Immunofluorescence Visualization of the Microtubules Roots of 2-week-old seedlings were fixed in 2% (v/v) paraformaldehyde and 0.5% (v/v) glutaraldehyde in PEMT buffer (100 mM PIPES, 4 mM EGTA, 4 mM MgSO4, 0.05% [v/v] Triton X-100, pH 7.2) for 40 min, and rinsed in PEMT buffer three times for 10 min. Roots were postfixed in cold methanol (220�C) for 10 min on ice, rehydrated for 10 min in 13 PBS (136 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 2 mM KH2PO4, pH 7.4), and treated for 20 min with NaBH4 (1 mg/mL) diluted in 13 PBS. Fixed roots were digested for 10 min with 0.2% (w/v) pectolyase, 1% (w/v) macerozyme, 3% (w/v) caylase diluted 10 times in digestion buffer (25 mM MES, 8 mM CaCl2, 600 mM mannitol, pH 5.5). After three washes in PBSG buffer (13 PBS, 50 mM Gly), roots were incubated for 20 min in 5% normal goat serum diluted in PBSG to saturate nonspecific sites, then incubated with primary anti- bodies directed against a-tubulin (Molecular Probes) in PBSG at 4�C overnight and washed three times for 5 min in PBSG buffer. Samples were incubated for 1 h at room temperature with secondary antibodies cou- pled to Alexa Fluor 488 (Molecular Probes) and washed three times in PBSG buffer. Roots were mounted in Mowiol containing DABCO (100 mg/mL). Observations were done using a Zeiss LSM510 con- focal laser scanning microscope equipped with argon and helium/neon lasers and with a C-APOCHROMAT (3 63, 1.2 numerical aperture water immersion lens). Excitation/emission wavelengths were 488/bandpass 505 to 550 nm for Alexa 488. Image processing was done using LSM510 version 2.8 (Zeiss), ImageJ (W.S. Rasband; National Institutes of Health), and Photo- shop 6.0 (Adobe Systems). Vol. 141: 803–804, 2006 Plant Physiology regrets that the credit line was not included with the image of Pablo Picasso’s Woman with a Cigarette in July’s On the Inside feature. The Estate of Pablo Picasso has graciously granted permission to ASPB for use of this image in the print and online journal. The credit line for this image is as follows: � 2006 Estate of Pablo Picasso/Artists Rights Society (ARS), New York. 1708 Plant Physiology, August 2006, Vol. 141, p. 1708, www.plantphysiol.org � 2006 American Society of Plant Biologists