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.

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.

Autotoxicity mechanism of Oryza sativa: transcriptome response in rice roots exposed to ferulic acid.

BACKGROUND: Autotoxicity plays an important role in regulating crop yield and quality. To help characterize the autotoxicity mechanism of rice, we performed a large-scale, transcriptomic analysis of the rice root response to ferulic acid, an autotoxin from rice straw. RESULTS: Root growth rate was decreased and reactive oxygen species, calcium content and lipoxygenase activity were increased with increasing ferulic acid concentration in roots. Transcriptome analysis revealed more transcripts responsive to short ferulic-acid exposure (1- and 3-h treatments, 1,204 genes) than long exposure (24 h, 176 genes). Induced genes were involved in cell wall formation, chemical detoxification, secondary metabolism, signal transduction, and abiotic stress response. Genes associated with signaling and biosynthesis for ethylene and jasmonic acid were upregulated with ferulic acid. Ferulic acid upregulated ATP-binding cassette and amino acid/auxin permease transporters as well as genes encoding signaling components such as leucine-rich repeat VIII and receptor-like cytoplasmic kinases VII protein kinases, APETALA2/ethylene response factor, WRKY, MYB and Zinc-finger protein expressed in inflorescence meristem transcription factors. CONCLUSIONS: The results of a transcriptome analysis suggest the molecular mechanisms of plants in response to FA, including toxicity, detoxicification and signaling machinery. FA may have a significant effect on inhibiting rice root elongation through modulating ET and JA hormone homeostasis. FA-induced gene expression of AAAP transporters may contribute to detoxicification of the autotoxin. Moreover, the WRKY and Myb TFs and LRR-VIII and SD-2b kinases might regulate downstream genes under FA stress but not general allelochemical stress. This comprehensive description of gene expression information could greatly facilitate our understanding of the mechanisms of autotoxicity 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.

Protein tyrosine dephosphorylation during copper-induced cell death in rice roots.

Early signalling events that control the process of heavy metal-induced cell death are largely unknown in plants. In mammals protein tyrosine phosphorylation plays an important role in the activation of programmed cell death. We thus examined the involvement of tyrosine phosphorylation in Cu-induced rice cell death. This investigation demonstrates that Cu induces cell death and DNA fragmentation in rice root cells. In the presence of Cu, the level of phosphotyrosine accumulation declined in the band of 45 kDa, p45. To analyze the role of tyrosine dephosphorylation for the regulation of Cu-induced cell death more precisely, we increased levels of tyrosine phosphorylation using the protein tyrosine phosphatase inhibitor, sodium orthovanadate (Na(3)VO(4)). Treatment of rice roots with Na(3)VO(4) blocked Cu-induced cell death and protein tyrosine dephosphorylation. In addition, the antioxidant GSH and the calcium chelator EGTA significantly abolished Cu-induced cell death and protein tyrosine dephosphorylation. These results provide evidence that dephosphorylation of a tyrosine-phosphorylated protein, p45, is an important step in the Cu-triggered signalling transduction pathway.

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.

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.

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.

Possible involvement of MAP kinase pathways in acquired metal-tolerance induced by heat in plants.

Cross tolerance is a phenomenon that occurs when a plant, in resisting one form of stress, develops a tolerance to another form. Pretreatment with nonlethal heat shock has been known to protect cells from metal stress. In this study, we found that the treatment of rice roots with more than 25 muM of Cu(2+) caused cell death. However, heat shock pretreatment attenuated Cu(2+)-induced cell death. The mechanisms of the cross tolerance phenomenon between heat shock and Cu(2+) stress were investigated by pretreated rice roots with the protein synthesis inhibitor cycloheximide (CHX). CHX effectively block heat shock protection, suggesting that protection of Cu(2+)-induced cell death by heat shock was dependent on de novo protein synthesis. In addition, heat pretreatment downregulated ROS production and mitogen-activated protein kinase (MAPK) activities, both of which can be greatly elicited by Cu(2+) stress in rice roots. Moreover, the addition of purified recombinant GST-OsHSP70 fusion proteins inhibited Cu(2+)-enhanced MAPK activities in an in vitro kinase assay. Furthermore, loss of heat shock protection was observed in Arabidopsis mkk2 and mpk6 but not in mpk3 mutants under Cu(2+) stress. Taken together, these results suggest that the interaction of OsHSP70 with MAPKs may contribute to the cellular protection in rice roots from excessive Cu(2+) toxicity.