Functional Proteomics ofArabidopsis thalianaGuard Cells Uncovers New Stomatal Signaling Pathways

Abstract

We isolated a total of 3 3 10 8 guard cell protoplasts from 22,000 Arabidopsis thaliana plants and identified 1734 unique proteins using three complementary proteomic methods: protein spot identification from broad and narrow pH range twodimensional (2D) gels, and 2D liquid chromatography-matrix assisted laser desorption/ionization multidimensional protein identification technology. This extensive single-cell-type proteome includes 336 proteins not previously represented in transcriptome analyses of guard cells and 52 proteins classified as signaling proteins by Gene Ontology analysis, of which only two have been previously assessed in the context of guard cell function. THIOGLUCOSIDE GLUCOHYDROLASE1 (TGG1), a myrosinase that catalyzes the production of toxic isothiocyanates from glucosinolates, showed striking abundance in the guard cell proteome. tgg1 mutants were hyposensitive to abscisic acid (ABA) inhibition of guard cell inward K + channels and stomatal opening, revealing that the glucosinolate-myrosinase system, previously identified as a defense against biotic invaders, is required for key ABA responses of guard cells. Our results also suggest a mechanism whereby exposure to abiotic stresses may enhance plant defense against subsequent biotic stressors and exemplify how enhanced knowledge of the signaling networks of a specific cell type can be gained by proteomics approaches.

Antisense Inhibition of the Iron-Sulphur Subunit of Succinate Dehydrogenase Enhances Photosynthesis and Growth in Tomato via an Organic Acid–Mediated Effect on Stomatal Aperture

“…For this purpose, we focused on the line SDH14 and the wild type and hybridized RNA both from whole leaf and epidermal fragments. Evaluation of epidermal fragments has proven highly informative in assessing the transcriptome of guard cells (Leonhardt et al, 2004), while the proteome of guard cell protoplasts has also recently been studied (Zhao et al, 2008). However, our studies revealed no significant changes in the expression of genes in the succinate dehydrogenase antisense line compared with the wild type after adjusting for multiple testing, in keeping with the few significant changes reported for the fumarase antisense lines (Nunes-Nesi et al, 2007b).…”

“…Moreover, analysis of the expression data did not uncover large changes in ABA-responsive genes (Hetherington and Quatrano, 1991;Hildmann et al, 1992;Abe et al, 1997) or of genes/proteins associated with ABA-mediated signal transduction relay (Leonhardt et al, 2004). The recent proteomic study of Zhao et al (2008) highlighted a plethora of proteins that were differentially expressed in guard cells as opposed to mesophyll cells, many of which have previously been characterized as being ABA responsive. Similarly, the experiments in which leaves from the transgenic and wild-type lines were incubated in the presence of the potassium channel blocker CsCl (Ichida et al, 1997) yielded similar results.…”

“…Over many years, considerable research effort has been expended to better comprehend both the formation and physiology of stomata (Hedrich and Neher, 1987;Schroeder and Hagiwara, 1990;Berger and Altmann, 2000;Bergmann et al, 2004;Lee et al, 2008;Zhao et al, 2008;Casson and Hetherington, 2010). Given the relative ease of the isolation of guard cells, which gate stomata, our understanding of these cells exceeds that of many of the other 40 cell types described thus far in plants (Martin et al, 2001;Fernie, 2007).…”

Gene-Sharing Networks Reveal Organizing Principles of Transcriptomes in Arabidopsis and Other Multicellular Organisms

“…First, we found that leptokurtic genes associated with guard cells are enriched in genes with recognized guard cell functions (Table 1): 28 out of 93 genes with known functions in guard cells (Zhao et al, 2008) are present in the list of 2345 guard cell leptokurtic genes (Fisher test, P < 6.3e-8.). Our analysis, based on the standardized transcriptome data set of normal Arabidopsis tissues, reveals that most of these genes are shared between guard cells and other tissues: Only three genes are guard cell specific (i.e., expressed above a basal level solely in guard cell samples), whereas the remaining 25 genes are shared between guard cells and other tissues.…”

Plant Stress Surveillance Monitored by ABA and Disease Signaling Interactions

“…In regulation of merging signals for ABA-and JA-triggered stomatal closures, guard cell abundant myrosinase THIOGLUCOSIDE GLUCO-HYDROLASE1 (TGG1) may have a role. tgg1 showed defects in ABA-inhibition of inward K + -channel activity and stomatal opening (Zhao et al, 2008) and ttg1 ttg2 produced reduced responses in the ABA-and JA-induced stomatal closures (Islam et al, 2009) suggesting a role of glucosinolate metabolism in the guard cell ABA signaling.…”