拟南芥WRKY57基因参与调控JA诱导叶片衰老的分子机制研究

Leaves are the important spots of photosynthesis in plants and leaf senescence, as the last stage of leaf development, is regulated by diverse developmental and environmental factors. The progression of leaf senescence is accompanied by the rapid loss of photosynthetic capacity resulted to the decreased yield of main crop. Therefore, delaying leaf senescence might be directly increase yield and improve the quality of crop. Recently, several studies on model plants displayed some key factors involved in leaf senescence, however, the molecular mechanism of it remains unknown. In this study, genetic and physiological approaches will used to reveal the molecular basis of WRKY57 transcription factor involved in JA-induced leaf senescence in Arabidopsis. Our previous study results showed that WRKY57 gene of Arabidopsis was significantly induced by JA treatment. Two T-DNA inserted mutant of WRKY57 displayed worse leaves senescence phenotype than wild type leaves, and the overexpression plants displayed opposite tendency.More importantly, the tissue expression pattern confirmed that the expression of WRKY57 was high.These analysis suggested that the WRKY57 transcription factor plays a key role in JA induced leaf senescence. The interaction proteins(JAZ4 and JAZ8)with WRKY57 transcription factor selected by the yeast-two-hybrid system construct the regulating relationship between WRKY57 transcription factor and JA induced leaf senescence signaling pathway. Based on these beforehand analysis, the directly downstream targeting genes will gain based on the ChIP-Seq and then construction the web regulating pattern of WRKY57 and its downstream targeting genes. According to the above research basis and thinking, we will illustrate the molecular mechanism about WRKY57 involved in the JA-induced leaf senescence and, at the same time, rich the signaling pathway of leaf senescence started by JA and most importantly provide some research foundation and approach for controlled the leaf senescence process in the future.

叶片是植物光合作用的重要场所，叶片衰老将导致作物产量下降。近年来对模式植物的研究虽然揭示了一些衰老进程的关键因子，但是对叶片衰老的调控机理还知之甚少。本课题拟研究拟南芥WRKY57参与调控JA诱导叶片衰老的分子机制。前期研究表明，WRKY57显著受JA诱导表达，分析WRKY57的两突变体和超表达植物，发现WRKY57突变导致了叶片在JA处理下比野生型提前衰老，而超表达植物表现出相反的表型。组织表达分析表明WRKY57在衰老叶片中的表达最强。暗示WRKY57在JA诱导的叶片衰老过程中起重要作用。酵母双元杂交证实WRKY57跟JAZ4和JAZ8在蛋白水平上相互作用，建立起WRKY57跟JA信号途径之间的直接联系。在此基础上我们利用基因芯片和ChIP-Seq技术，寻找WRKY57的直接调控靶基因，构建WRKY57跟其靶基因的调控关系，有效阐明WRKY57参与调控JA诱导叶片衰老的分子生物学机制。

叶片是植物进行光合作用的重要场所。叶片一旦衰老，其光合能力将显著下降，从而导致作物的产量下降。对于粮食作物和多数经济作物来说，延缓叶片衰老则可以直接增加农作物产量及品质。因此，研究植物的衰老进程有助于我们利用基因工程技术提高农作物的产量和经济作物的价值。. 在本项目中，我们主要研究四个方面的内容：(1)拟南芥WRKY57基因在调控JA诱导的叶片衰老过程中的功能分析；(2)WRKY57相互作用蛋白的筛选及其协调调控JA诱导的细胞衰老信号转导的分子机制；(3)WRKY57蛋白所调控的下游靶基因筛选及其调控细胞衰老信号转导的分子生物学机制；(4).WRKY57直接靶基因的分子生物学功能分析。. 并取得了一系列研究成果。首先，通过酵母双杂交技术筛选JA处理下跟WRKY57直接相互作用的蛋白，获得了WRKY57的相互作用蛋白JAZ4/8 和IAA29，并阐明JAZ4/8负调控JA诱导的叶片衰老，而IAA29正调控JA诱导的叶片衰老；利用染色质免疫沉淀技术，获得WRKY57转录因子直接调控的下游靶基因SEN4 和SAG12，并构建了拟南芥WRKY57转录因子跟其直接下游靶基因的网络调控模型，并确定WRKY57基因在这个信号途径中的位置，阐明拟南芥WRKY57基因在JA诱导的叶片衰老的信号转导途径中的功能，即拟南芥感受的JA信号后通过竞争性地和JAZ4/8和IAA29相互作用，并在auxin拮抗JA诱导的叶片衰老过程中起关键作用。. 近二十年来的研究表明WRKY基因参与了植物的生物和非生物逆境响应。但是对于WRKY转录因子如何调控叶片衰老过程的研究还甚少。而我们该项目详细地分析WRKY57参与衰老的分子机制，不仅有利于我们深入了解JA信号的转导过程，而且也有利于我们未来利用基因工程技术改善农作物的产量和品质。