Mechanisms of response to environmental stresses with active oxygen species-detoxification in higer plants

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Salt tolerance in plants has been attributed to many mechanisms, including accumulation of osmoregulatory solutes inside cells, enhanced ability of cells to take up water or K^+ from surrounding solutions of higher salt concentration, elimination of excess NaCl, and the ability to maintain photosynthetic activity under salt stress. To further investigate the mechanisms of salt tolerance, we have begun to screen and characterize mutants of Arabidopsis thaliana tolerant to high NaCl concentrations during photoautotrophic growth. Several mutants of A. thaliana with altered responses to salt have been reported, including mutants tolerant of salt during germination and mutants that are salt-hypersensitive. However, there have been no reports of mutants whose photoautotrophic growth is salt tolerant.We selected pst (photoautotrohic salt tolerance) mutant lines of Arabidopsis, and characterized pst1 plants in which activities of active oxygen species-detoxifying enzymes were enhanced (Plant Cell, 11, 1195-1206, 1999). However, it was difficult to specify the event crucial for conferring salt tolerance on pst2 plants. The cDNA macroarray and oligo-microarray were performed for pst2 and wild-type lines grown under a non-stress condition. Higher expression of a transcription factor with bHLH motif in pst2 plants was detected, and further confirmed by real-time RT-PCR. There was another specie of mRNA produced by alternative splicing under the salt stress. The RACE has clarified that transcription initiation site is single. The salt-induced mRNA retained an intron on C-terminal side among two introns, and the newly generated stop codon of first ORF and its subsequent initiation codon of new second ORF may encode two separate peptide molecules which looked like cleaved forms of the original protein. Protein species derived from the bHLH gene are being analyzed by immuno-blotting, and the function of bHLH transcription factor is also examined with its transgenic plants.