OsJAmyb介导的水稻抗虫机理研究

Our previous studies have shown that a transcriptional factor OsJAmyb plays an important role in rice defense response and herbivore resistance. To elucidate how OsJAmyb regulates the rice defense responses and herbivore resistance, we, in this project, will first investigate the subcelluar location and induced expression patterns of OsJAmyb by fluorescence microscopy and quantitative real time PCR, respectively. Second, the binding sites and transcriptional activation of OsJAmyb will be analyzed using yeast one-hybrid system and electrophoreretic mobility shift assay. Third, the interacting proteins of OsJAmyb will be isolated by combining yeast two-hybrid system, bimolecular fluorescence complementation and pull-down assay. Finally, the effects of OsJAmyb on herbivory-related signaling pathways, defensive compounds and transcriptional profiling, as well as herbivore resistance will be analyzed by combination of molecular biology, transcriptome, metabolome, chemical analysis and bioassay methods. This comprehensive analysis will provide new insights into the mechanism responsible for rice herbivore resistance mediated by OsJAmyb and will provide resistance genes for crop breeding.

本项目将以我们具有很好研究基础的水稻转录因子OsJAmyb为研究对象，通过利用荧光显微技术和定量PCR，研究OsJAmyb的亚细胞定位及其诱导表达特征；利用凝胶阻滞分析及酵母单杂交方法，分析OsJAmyb的结合位点和转录激活活性；利用酵母双杂交、下拉分析、双分子荧光互补等方法，分离鉴定OsJAmyb的互作蛋白；运用分子生物学、转录组学、代谢组学、化学分析以及生物测定等技术手段，揭示OsJAmyb对防御信号分子、防御化合物、转录组及水稻抗虫性的影响；从而深入剖析OsJAmyb介导的水稻防御反应的化学与分子机理，发掘新的水稻抗虫基因，为水稻新型抗虫品种的培育提供理论与技术指导。

本项目以水稻转录因子OsJAmyb为研究对象，综合运用分子生物学、转录组学、生物化学、生物测定等技术手段，系统研究了OsJAmyb在水稻抗虫反应中的作用与机理。OsJAmyb定位于植物细胞核，具有转录激活活性。机械损伤、二化螟（Chilo suppressalis, striped stem borer, SSB）、褐飞虱（Niaparvata lugens, brown planthopper, BPH）为害能够迅速诱导OsJAmyb基因上调，外源茉莉酸（Jasmonic acid, JA）、水杨酸（Salicylic acid, SA）及乙烯处理也能激活OsJAmyb表达。沉默OsJAmyb提高了BPH为害后的JA含量，降低了绿叶性气味（Green leaf volatiles, GLVs）的释放，但不影响水稻本底及虫害诱导的SA、乙烯、H2O2合成，防御化合物胰蛋白酶抑制剂（Trypsin protease inhibitors, TryPIs）活性及挥发物（Volatile organic compounds, VOCs）含量。室内生测和田间试验结果表明，沉默OsJAmyb能够显著增强水稻对二化螟、褐飞虱及白背飞虱（Sogatella furcifera, white-backed planthopper, WBPH）的抗性。综上所述，OsJAmyb在水稻-害虫互作中发挥了重要的调控作用，其沉默突变体具有良好的应用前景。