Wen Chang Chi

Mercury-induced biochemical and proteomic changes in rice roots

Mercury (Hg) is a serious environmental pollution threats to the planet. Accumulation of Hg in plants disrupts many cellular-level functions and inhibits growth and development, but the mechanism is not fully understood. We investigated cellular, biochemical and proteomic changes in rice roots under Hg stress. Root growth rate was decreased and Hg, reactive oxygen species (ROS), and malondialdehyde (MDA) content and lipoxygenase activity were increased significantly with increasing Hg concentration in roots. We revealed a time-dependent alteration in total glutathione content and enzymatic activity of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and peroxidase (POD) during Hg stress. 2-D electrophoresis revealed differential expression of 25 spots with Hg treatment of roots: 14 spots were upregulated and 11 spots downregulated. These differentially expressed proteins were identified by ESI-MS/MS to be involved in cellular functions including redox and hormone homeostasis, chaperone activity, metabolism, and transcription regulation. These results may provide new insights into the molecular basis of the Hg stress response in plants.

Identification of transcriptome profiles and signaling pathways for the allelochemical juglone in rice roots

Juglone (5-hydroxy-1,4-naphthoquinone) is known allelochemical, but its molecular mode of action is not well understood. We found that juglone induced reactive oxygen species production and calcium accumulation. To gain more insight into these cellular responses, we performed large-scale analysis of the rice transcriptome during juglone stress. Exposure to juglone triggered changes in transcript levels of genes related to cell growth, cell wall formation, chemical detoxification, abiotic stress response and epigenesis. The most predominant transcription-factor families were AP2/ERF, HSF, NAC, C2H2, WRKY, MYB and GRAS. Gene expression profiling of juglone-treated rice roots revealed upregulated signaling and biosynthesis of abscisic acid and jasmonic acid and inactivation of gibberellic acid. In addition, juglone upregulated the expression of two calcium-dependent protein kinases (CDPKs), 6 mitogen-activated protein kinase (MAPK) genes and 1 MAPK gene and markedly increased the activities of a CDPK-like kinase and MAPKs. Further characterization of these juglone-responsive genes may be helpful for better understanding the mechanisms of allelochemical tolerance in plants.

Chromium stress response effect on signal transduction and expression of signaling genes in rice.

Hexavalent chromium [Cr(VI)] is a non-essential metal for normal plants and is toxic to plants at high concentrations. However, signaling pathways and molecular mechanisms of its action on cell function and gene expression remain elusive. In this study, we found that Cr(VI) induced endogenous reactive oxygen species (ROS) generation and Ca(2+) accumulation and activated NADPH oxidase and calcium-dependent protein kinase. We investigated global transcriptional changes in rice roots by microarray analysis. Gene expression profiling indicated activation of abscisic acid-, ethylene- and jasmonic acid-mediated signaling and inactivation of gibberellic acid-related pathways in Cr(VI) stress-treated rice roots. Genes encoding signaling components such as the protein kinases domain of unknown function 26, receptor-like cytoplasmic kinase, LRK10-like kinase type 2 and protein phosphatase 2C, as well as transcription factors WRKY and apetala2/ethylene response factor were predominant during Cr(VI) stress. Genes involved in vesicle trafficking were subjected to functional characterization. Pretreating rice roots with a vesicle trafficking inhibitor, brefeldin A, effectively reduced Cr(VI)-induced ROS production. Suppression of the vesicle trafficking gene, Exo70, by virus-induced gene silencing strategies revealed that vesicle trafficking is required for mediation of Cr(VI)-induced ROS production. Taken together, these findings shed light on the molecular mechanisms in signaling pathways and transcriptional regulation in response to Cr stress in plants.

Early signalling pathways in rice roots under vanadate stress.

Vanadate is beneficial to plant growth at low concentration. However, plant exposure to high concentrations of vanadate has been shown to arrest cell growth and lead to cell death. We are interested in understanding the signalling pathways of rice roots in response to vanadate stress. In this study, we demonstrated that vanadate induced rice root cell death and suppressed root growth. In addition, we found that vanadate induced ROS accumulation, increased lipid peroxidation and elicited a remarkable increase of MAPKs and CDPKs activities in rice roots. In contrast, pre-treatment of rice roots with ROS scavenger (sodium benzoate), serine/threonine protein phosphatase inhibitor (endothall), and CDPK antagonist (W7), reduced the vanadate-induced MAPKs activation. Furthermore, the expression of a MAPK gene (OsMPK3) and four tyrosine phosphatase genes (OsDSP3, OsDSP5, OsDSP6, and OsDSP10) were regulated by vanadate in rice roots. Collectively, these results strongly suggest that ROS, protein phosphatase, and CDPK may function in the vanadate-triggered MAPK signalling pathway cause cell death and retarded growth in rice roots.

Transcriptome profiling and physiological studies reveal a major role for aromatic amino acids in mercury stress tolerance in rice seedlings

Mercury (Hg) is a serious environmental pollution threat to the planet. The accumulation of Hg in plants disrupts many cellular-level functions and inhibits growth and development, but the mechanism is not fully understood. To gain more insight into the cellular response to Hg, we performed a large-scale analysis of the rice transcriptome during Hg stress. Genes induced with short-term exposure represented functional categories of cell-wall formation, chemical detoxification, secondary metabolism, signal transduction and abiotic stress response. Moreover, Hg stress upregulated several genes involved in aromatic amino acids (Phe and Trp) and increased the level of free Phe and Trp content. Exogenous application of Phe and Trp to rice roots enhanced tolerance to Hg and effectively reduced Hg-induced production of reactive oxygen species. Hg induced calcium accumulation and activated mitogen-activated protein kinase. Further characterization of the Hg-responsive genes we identified may be helpful for better understanding the mechanisms of Hg in plants.

Pathways involved in vanadate-induced root hair formation in Arabidopsis.

Root hair formation is controlled by environmental signals. We found significantly increased Arabidopsis root hair density and length in response to low-dose vanadate (V). Reactive oxygen species (ROS) production was induced with V treatment. We investigated the possible role of NADPH oxidase in altering root system architecture induced by V by using diphenylene iodonium (DPI), an inhibitor of NADPH oxidase, and an NADPH oxidase mutant (rhd2/AtrbohC). NADPH oxidase was involved in root hair elongation induced by V. As well, ethylene receptor (ETR1) and ROOT HAIR DEFECTIVE (RHD6) participated in inducing root hair formation induced by V. Furthermore, the kinase inhibitors, genistein (tyrosine kinase inhibitor) and K252a (ser/thr kinase inhibitor), and a phosphatase inhibitor, cantharidin (ser/thr phosphatase inhibitor), suppressed root hair formation induced by V. To elucidate the regulation of gene expression in response to V, we investigated transcriptional changes in roots by microarray assay. Exposure to V triggered changes in transcript levels of genes related to cell wall formation, ROS activity and signaling. Several genes involved in root hair formation were also regulated.

Alleviation of allelochemical juglone-induced phytotoxicity in tobacco plants by proline

Juglone (5-hydroxy-1,4-naphthoquinone) is an important allelochemical in walnut trees (Juglans nigra L.). Its allelopathic potential has been reported in different plant species. We investigated the phytotoxic effects of the allelochemical juglone and the protective role of proline in tobacco seedlings. Juglone inhibited the growth of tobacco seedlings and increased reactive oxygen species content in tobacco roots. Moreover, juglone stress increased proline concentration. The expression of two proline synthesis genes, pyrroline-5-carboxylate synthetase and ornithine aminotransferase, was upregulated and that of a proline catabolism gene, proline dehydrogenase, was downregulated with juglone treatment in tobacco roots. Furthermore, plants pretreated with proline and then exposed to juglone showed attenuated toxic effects in roots. Proline was able to modulate allelochemical juglone-induced stress in tobacco. In summary, this study suggested that increased proline content in the tobacco seedlings treated with juglone may mitigate the deleterious effect of allelochemical stress in plants by inhibiting reactive oxygen species accumulation.