Functional Analysis Of AtST39an AtST39H In Salt And Drought Stress

Abiotic stresses such as high salty, drought and low temperature have brought a significant impact on agriculture, they can result in reduction of crops seriously. Therefore, it becomes more and more important to study the resistance mechanisms and isolate resistance-related genes. Arabidopsis, as a model plant, has its unique advantages for research. Previous studies have obtained that AtST39(At3g46110) and AtST39H (At5g59790), two genes of DUF966gene families, may involve in stress resistance pathway. My research continues on studying the functions of AtST39and AtST39H.Subcellular localization assay suggest AtST39is a cytoplasm and plasma membrane-localized protein. The tissue expression pattern of AtST39is examined by performing histochemical staining for GUS activity in various tissues of AtST39promoter-GWS1plants. The study results show that AtST39is expressed in roots, stems, lesve in seeding stage, and veins, styluses,anthers and noth of stems. Expression profile analysis of adversity show that the expression of AtST39is induced by drought, low temperature and salt stresses, especially salt stress. We study the effects of salt stress on root growth of transgenic plants on seeding stage. The results show that AtST39-RNAi plants are more sensitive:under salt treatment, the taproot length are much shorter than the wild type; the microscope observation on root tip cells, we find: without salt stress the root tips of AtST39-RNAi are already swollen(wild-type apical swells by stress), and the root tips extremely collapse with salt stress; the activity of CAT is nearly six times higher than the wild type. We also find that overexpression of AtST39plants are less sensitive to salt and drought than the wild type, while T-DNA insertion mutants and interference lines are more sensitive to salt and drought than the wild type. These results show that:AtST39genes may involve in salt and drought stress resistance pathway, and may play a positive role.Expression profile analysises by qRT-PCR show that in AtST39-KNAi plants RD29A, RD29B, RD22, RAB, ERD1, COR15are raised significantly; Auxin-related transcription factor ARF5, Auxin synthase related genes YUC1, YUC4also significantly increased, indicating auxin signaling pathway of AtST39-RNAi plants is activated. To detect endogenous auxin distribution, we obtain hybrids of transgenetic plants and DR5::GUS plants, and find after NaCl treatment, the endogenous auxin levels of AtST39-OE plants and wild-type plants are increased, and increased as time passes; while after NaCl treatment, the endogenous auxin levels of3-3and AtST39-RNAi plants are increased, but decreased as time passes. These results suggest AtST39may participate in or influence Auxin signaling pathway.Subcellular localization assay suggest AtST39H is a plasma membrane and nucleus-localized protein. The tissue expression pattern of AtST39H is examined by performing histochemical staining for GUS activity in various tissues of AtST39H promoter-GUS plants. The results show that AtST39H is expressed in leaves at seeding stage, and styluses, sepals at adult stage. Expression profile analysis show that the expression of AtST39H is induced by salt stresses. But we find there is no difference between mutant5-5and wild type with salt and drought stress. Expression profile analysises by qRT-PCR show that the expression level of defense-related genes in AtST39H mutant,5-5, are downregulated, but RD22, ERD1, PAD3increased significantly. Auxin synthase related genes YUC1, YUC4also significantly increased. We construct prokaryotic expression vector His-AtST39H and GST-AtST39H, and purify the strain BL21-pET28a-AtST39H inclusion bodies, to obtain the protein of AtST39H. Then we can get the antibody, and detect on protein level.The double mutant hybrids of AtST39and AtST39H have no difference with wild type under normal circumstances. Expression profile analysises by qRT-PCR show that the expression levels of defense-related genes are increased, especially RD29A and RAB. Auxin-related transcription factor ARF5and Auxin synthase related genes YUC1are also significantly increased, indicating Auxin signaling pathway of the double mutant hybrids may be activated under normal circumstances.