[CLE peptides regulate plant morphogenesis as intercellular signaling molecules].
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Arabidopsis Proteins↗
Biomedical subjects
Shinichiro Sawa
Publications and source records attributed to Shinichiro Sawa.
Involvement of HLS1 in sugar and auxin signaling in Arabidopsis leaves.
Sugar regulates a variety of genes and controls plant growth and development similarly to phytohormones. As part of a screen for Arabidopsis mutants with defects in sugar-responsive gene expression, we identified a loss-of-function mutation in the HOOKLESS1 (HLS1) gene. HLS1 was originally identified to regulate apical hook formation of dark-grown seedlings (Lehman et al., 1996, Cell 85: 183-194). In hls1, sugar-induced gene expression in excised leaf petioles was more sensitive to exogenous sucrose than that in the wild type. Exogenous IAA partially repressed sugar-induced gene expression and concomitantly activated some auxin response genes such as AUR3 encoding GH3-like protein. The repression and the induction of gene expression by auxin were attenuated and enhanced, respectively, by the hls1 mutation. These results suggest that HLS1 plays a negative role in sugar and auxin signaling. Because AUR3 GH3-like protein conjugates free IAA to amino acids (Staswick et al., 2002, Plant Cell 14: 1405-1415; Staswick et al., 2005, Plant Cell 17: 616-627), enhanced expression of GH3-like genes would result in a decrease in the free IAA level. Indeed, hls1 leaves accumulated a reduced level of free IAA, suggesting that HLS1 may be involved in negative feedback regulation of IAA homeostasis through the control of GH3-like genes. We discuss the possible mechanisms by which HLS1 is involved in auxin signaling for sugar- and auxin-responsive gene expression and in IAA homeostasis.
Arabidopsis↗
CD1d-restricted NKT cell activation enhanced homeostatic proliferation of CD8+ T cells in a manner dependent on IL-4.
CD1d-restricted NKT cells are activated by TCR-mediated stimulation via CD1d plus lipid antigens such as alpha-galactosylceramide (alpha-GalCer). These cells suppressed autoimmunity and graft rejection, but sometimes enhanced resistance to infection and tumor immunity. This double-action phenomenon of NKT cells is partly explained by cytokines produced by NKT cells. Therefore, roles of cytokines from activated NKT cells have been extensively examined; however, their roles on T cell homeostatic proliferation in lymphopenic condition have not been investigated. Here, we showed that alpha-GalCer enhanced homeostatic proliferation of CD8+ but not CD4+ T cells and this effect of alpha-GalCer was required for NKT cells. IL-4 was essential and sufficient for this NKT cell action on CD8+ T cell homeostatic proliferation. Importantly, the expression of IL-4Ralpha and STAT6 in CD8+ T cells was essential for the NKT activity, indicating a direct action of IL-4 on CD8+ T cells. Consistent with this, the level of IL-4Ralpha expression on memory phenotype CD8(+) T cells was higher than that on naive phenotype one and CD4+ T cells. Thus, these results showed the 'involvement' of IL-4 that is produced from activated NKT cells for CD8+ T cell homeostatic proliferation in vivo.
Animals↗
Dodeca-CLE peptides as suppressors of plant stem cell differentiation.
In plants and animals, small peptide ligands that signal in cell-cell communication have been suggested to be a crucial component of development. A bioassay of single-cell transdifferentation demonstrates that a dodecapeptide with two hydroxyproline residues is the functional product of genes from the CLE family, which includes CLAVATA3 in Arabidopsis. The dodecapeptide suppresses xylem cell development at a concentration of 10(-11) M and promotes cell division. An application, corresponding to all 26 Arabidopsis CLE protein family members, of synthetic dodecapeptides reveals two counteracting signaling pathways involved in stem cell fate.
Amino Acid Sequence↗
A plant peptide encoded by CLV3 identified by in situ MALDI-TOF MS analysis.
The Arabidopsis CLAVATA3 (CLV3) gene encodes a stem cell-specific protein presumed to be a precursor of a secreted peptide hormone. Matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS) applied to in situ Arabidopsis tissues determined the structure of a modified 12-amino acid peptide (MCLV3), which was derived from a conserved motif in the CLV3 sequence. Synthetic MCLV3 induced shoot and root meristem consumption as cells differentiated into other organs, displaying the typical phenotype of transgenic plants overexpressing CLV3. These results suggest that the functional peptide of CLV3 is MCLV3.
Amino Acid Motifs↗
Interaction of auxin and ERECTA in elaborating Arabidopsis inflorescence architecture revealed by the activation tagging of a new member of the YUCCA family putative flavin monooxygenases.
The aboveground body of higher plants has a modular structure of repeating units, or phytomers. As such, the position, size, and shape of the individual phytomer dictate the plant architecture. The Arabidopsis (Arabidopsis thaliana) ERECTA (ER) gene regulates the inflorescence architecture by affecting elongation of the internode and pedicels, as well as the shape of lateral organs. A large-scale activation-tagging genetic screen was conducted in Arabidopsis to identify novel genes and pathways that interact with the ER locus. A dominant mutant, super1-D, was isolated as a nearly complete suppressor of a partial loss-of-function allele er-103. We found that SUPER1 encodes YUCCA5, a novel member of the YUCCA family of flavin monooxygenases. The activation tagging of YUCCA5 conferred increased levels of free indole acetic acid, increased auxin response, and mild phenotypic characteristics of auxin overproducers, such as elongated hypocotyls, epinastic cotyledons, and narrow leaves. Both genetic and cellular analyses indicate that auxin and the ER pathway regulate cell division and cell expansion in a largely independent but overlapping manner during elaboration of inflorescence architecture.
Amino Acid Sequence↗
DRP1A is responsible for vascular continuity synergistically working with VAN3 in Arabidopsis.
In most dicotyledonous plants, vascular tissues in the leaf have a reticulate venation pattern. We have isolated and characterized an Arabidopsis (Arabidopsis thaliana) mutant defective in the vascular network defective mutant, van3. van3 mutants show a discontinuous vascular pattern, and VAN3 is known to encode an ADP-ribosylation-factor-GTPase-activating protein that regulates membrane trafficking in the trans-Golgi network. To elucidate the molecular nature controlling the vein patterning process through membrane trafficking, we searched VAN3-interacting proteins using a yeast (Saccharomyces cerevisiae) two hybrid system. As a result, we isolated the plant Dynamin-Related Protein 1A (DRP1A) as a VAN3 interacting protein. The spatial and temporal expression patterns of DRP1AGUS and VAN3GUS were very similar. The subcellular localization of VAN3 completely overlapped to that of DRP1A. drp1a showed a disconnected vascular network, and the drp1a mutation enhanced the phenotype of vascular discontinuity of the van3 mutant in the drp1a van3 double mutant. Furthermore, the drp1 mutation enhanced the discontinuous vascular pattern of the gnom mutant, which had the same effect as that of the van3 mutation. These results indicate that DRP1 modulates the VAN3 function in vesicle budding from the trans-Golgi network, which regulates vascular formation in Arabidopsis.
ADP-Ribosylation Factors↗
VAN3 ARF-GAP-mediated vesicle transport is involved in leaf vascular network formation.
Within the leaf of an angiosperm, the vascular system is constructed in a complex network pattern called venation. The formation of this vein pattern has been widely studied as a paradigm of tissue pattern formation in plants. To elucidate the molecular mechanism controlling the vein patterning process, we previously isolated Arabidopsis mutants van1 to van7, which show a discontinuous vein pattern. Here we report the phenotypic analysis of the van3 mutant in relation to auxin signaling and polar transport, and the molecular characterization of the VAN3 gene and protein. Double mutant analyses with pin1, emb30-7/gn and mp, and physiological analyses using the auxin-inducible marker DR5::GUS and an auxin transport inhibitor indicated that VAN3 may be involved in auxin signal transduction, but not in polar auxin transport. Positional cloning identified VAN3 as a gene that encodes an adenosine diphosphate (ADP)-ribosylation factor-guanosine triphosphatase (GTPase) activating protein (ARF-GAP). It resembles animal ACAPs and contains four domains: a BAR (BIN/amphiphysin/RVS) domain, a pleckstrin homology (PH) domain, an ARF-GAP domain and an ankyrin (ANK)-repeat domain. Recombinant VAN3 protein showed GTPase-activating activity and a specific affinity for phosphatidylinositols. This protein can self-associate through the N-terminal BAR domain in the yeast two-hybrid system. Subcellular localization analysis by double staining for Venus-tagged VAN3 and several green-fluorescent-protein-tagged intracellular markers indicated that VAN3 is located in a subpopulation of the trans-Golgi network (TGN). Our results indicate that the expression of this gene is induced by auxin and positively regulated by VAN3 itself, and that a specific ACAP type of ARF-GAP functions in vein pattern formation by regulating auxin signaling via a TGN-mediated vesicle transport system.
ADP-Ribosylation Factors↗
Comprehensive comparison of auxin-regulated and brassinosteroid-regulated genes in Arabidopsis.
Although numerous physiological studies have addressed the interactions between brassinosteroids and auxins, little is known about the underlying molecular mechanisms. Using an Affymetrix GeneChip representing approximately 8,300 Arabidopsis genes, we studied comprehensive transcript profiles over 24 h in response to indole-3-acetic acid (IAA) and brassinolide (BL). We identified 409 genes as BL inducible, 276 genes as IAA inducible, and 637 genes in total. These two hormones regulated only 48 genes in common, suggesting that most of the actions of each hormone are mediated by gene expression that is unique to each. IAA-up-regulated genes were enriched in genes regulated in common. They were induced quickly by IAA and more slowly by BL, suggesting divergent physiological roles. Many were early auxin-inducible genes and their homologs, namely SAUR, GH3, and IAA. The comprehensive comparison also identified IAA- and BL-specific genes, which should help to elucidate the specific actions of each hormone. The identified genes were classified using hierarchical clustering based on the similarity of their responses to the two hormones. Gene classification also allowed us to analyze the frequency of cis-elements. The TGTCTC element, a core element of the previously reported auxin response element, was not enriched in genes specifically regulated by IAA but was enriched in the 5'-flanking region of genes up-regulated by both IAA and BL. Such gene classification should be useful for predicting the functions of unknown genes, to understand the roles of these two hormones, and the promoter analysis should provide insight into the interaction of transcriptional regulation by the two hormones.
Arabidopsis↗
A novel rice PR10 protein, RSOsPR10, specifically induced in roots by biotic and abiotic stresses, possibly via the jasmonic acid signaling pathway.
Plant roots have important roles not only in absorption of water and nutrients, but also in stress tolerance such as desiccation, salt, and low temperature. We have investigated stress-response proteins from rice roots using 2-dimensional polyacrylamide-gel electrophoresis and found a rice protein, RO-292, which was induced specifically in roots when 2-week-old rice seedlings were subjected to salt and drought stress. The full-length RO-292 cDNA was cloned, and was determined to encode a protein of 160 amino acid residues (16.9 kDa, pI 4.74). The deduced amino acid sequence showed high similarity to known rice PR10 proteins, OsPR10a/PBZ1 and OsPR10b. RO-292 mRNA accumulated rapidly upon drought, NaCl, jasmonic acid and probenazole, but not by exposure to low temperature or by abscisic acid and salicylic acid. The RO-292 gene was also up-regulated by infection with rice blast fungus. Interestingly, induction was observed almost exclusively in roots, thus we named the gene RSOsPR10 (root specific rice PR10). The present results indicate that RSOsPR10 is a novel rice PR10 protein, which is rapidly induced in roots by salt, drought stresses and blast fungus infection possibly through activation of the jasmonic acid signaling pathway, but not the abscisic acid and salicylic acid signaling pathway.
Abscisic Acid↗
Brassinolide induces IAA5, IAA19, and DR5, a synthetic auxin response element in Arabidopsis, implying a cross talk point of brassinosteroid and auxin signaling.
Despite numerous physiological studies addressing the interactions between brassinosteroids (BRs) and auxins, little is known about the underlying molecular mechanisms. We studied the expression of IAA5 and IAA19 in response to treatment with indole acetic acid (IAA) or brassinolide (BL), the most active BR. Exogenous IAA induced these genes quickly and transiently, whereas exogenous BL induced them gradually and continuously. We also found that a fusion of DR5, a synthetic auxin response element, with the GUS (beta-glucuronidase) gene was induced with similar kinetics to those of the IAA5 and IAA19 genes in response to both IAA and BL treatment of transgenic plants. These results suggest that the IAA genes are induced by BL, at least in part, via the activation of the auxin response element. Endogenous IAA levels per gram fresh weight did not increase when seedlings of Arabidopsis wild type (WT) or the BR-deficient mutant det2 were treated with BL. Furthermore, the levels of IAA transcripts were lower in the det2 mutant than in the WT, even though endogenous IAA levels per gram fresh weight were higher in the det2 mutant than in the WT. In conclusion, the lack of evidence for auxin-mediated activation of early auxin-inducible genes in response to BL suggests that the BR and auxin signaling pathways independently activate the transcriptional system of the IAA and DR5-GUS genes.
Arabidopsis↗
Overexpression of the AtmybL2 gene represses trichome development in Arabidopsis.
Leaf trichome formation is known to be regulated by the TTG, GL1, GL2, and GL3 genes in Arabidopsis. GL1 and GL3 encode proteins with Myb and bHLH domains, respectively. Overexpression of the AtmybL2 gene, which encodes a single Myb-like DNA-binding domain, repressed trichome development in transgenic Arabidopsis plants. The amount of GL2 transcription was clearly reduced in the transgenic plants. Consistent with this, overexpression of AtmybL2 decreased beta-glucuronidase (GUS) activity in transgenic plants carrying a GUS-reporter gene regulated by the GL2 promoter. These findings, together with the results from our yeast two-hybrid analysis, suggest that GL3 gene function and overexpression of AtmybL2 act synergistically to inhibit trichome formation by negatively regulating GL2 expression.
Arabidopsis↗
The HAT2 gene, a member of the HD-Zip gene family, isolated as an auxin inducible gene by DNA microarray screening, affects auxin response in Arabidopsis.
The plant hormone, auxin, regulates many aspects of growth and development. Despite its importance, the molecular mechanisms underlying the action of auxin are largely unknown. To gain a more comprehensive understanding of the primary responses to auxin, we analyzed the expression of genes in Arabidopsis seedlings treated with indole-3-acetic acid (IAA) for 15 min. We identified a single gene that is downregulated early, and 29 genes that are upregulated early. Several types of typical transcription factors are identified as early upregulated genes, suggesting that auxin signals are mediated by a master set of diverse transcriptional regulators. Of the genes that responded to auxin, the expression of the homeobox gene, HAT2, was induced rapidly. Furthermore, we show that the expression of HAT2 is induced by auxin, but not by other phytohormones. To analyze the function of HAT2 in the plant's response to auxin, we generated 35S::HAT2 transgenic plants. These produced long hypocotyls, epinastic cotyledons, long petioles, and small leaves, which are characteristic of the phenotypes of the auxin-overproducing mutants, superroot1 (sur1) and superroot2 (sur2). On the other hand, 35S::HAT2 plants showed reduced lateral root elongation, and reduced auxin sensitivity compared to wild-type plants. Together with the results of RNA blotting and biochemical analyses, these findings suggest that HAT2 plays opposite roles in the shoot and root tissues in regulating auxin-mediated morphogenesis.
Amino Acid Sequence↗
[Mechanisms of auxin biosynthesis].
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Aldehyde Oxidase↗