辣椒含DWE的WRKY基因功能相关及其在协同应答高温及青枯菌中作用解析

土传病害多的典型茄科植物辣椒在高温高湿下发病严重是制约其生产效益的重要限制因素，剖析辣椒高温高湿下抗病机制有利于其抗病遗传改良。项目前期发现DWE在WRKY基因启动子协同应答青枯菌及高温高湿中起关键作用，且发现43个含有DWE的WRKY蛋白（DWE-WKs），暗示它们可能构成辣椒协调耐高温高湿和抗病调控网络。基于此，本项目进一步分析各DWE-WKs亚细胞定位、结合W盒活性，并采用荧光定量PCR分析、在辣椒植株上瞬间超表达、病毒诱导基因沉默、染色质免疫沉淀（ChIP）、ChIP-seq等手段，分析这些DWE-WKs在辣椒应答高温高湿和抗青枯菌中的表达、作用、彼此相互转录激活、与彼此启动子的相互结合等。通过实施本项目，建立DWE-WKs构成的辣椒协同应答高温高湿和青枯菌侵染的WRKY网络，找出其中的关键调控节点，为辣椒等茄科植物高温高湿下抗病遗传改良提供合理技术策略和具有应用价值的基因。

Pepper（Capsicum annuum） is a one of the most important vegetable worldwide and a typical solanaceaes. Heavy loss in pepper production is frequently caused by diseases, which is usually aggravated by high temperature and high humidity. A better understanding of the signal crosstalk between high temperature & high humidity and pathogen response is helpful for the genetic improvement of disease resistance of pepper and other Solanaceae crops. Previously we found that a duouble W-box element,which is present in the promoter of CaWRKY40, a WRKY transcription factor of Capsicum annuum acting as an important regulator in pepper basal thermotolerance and resistance to Ralstonia solanacearum attack, confered largely the synergistic response of CaWRKY40 to high temperature & high humidity and pathogen infection. In addition, the DWE also found in the promoter of 43 members of the pepper WRKY family,implying that these members may constitute a network synergistically regulating pepper’s response to high temperature & high humidi and pathogen infection. Based on this, the common feathures such as W-box binding, and nuclei targeting will be analyzed on the DWE-WKs in the present study. Additionally, by approaches such as realtime PCR, transient expression and VIGS in pepper plants, chromatin immunoprecipitation, ChIP-seq, the functions of these DWE-WKs in pepper’s synergistic response to high temperature & high humidity and pathogen infection, their transcriptonal activation on each other, and their binding to the promoters of each other will be assayed. Based on the results of the project, the regulatory WRKY network on pepper’s synergistic response to high temperature & high humidity and pathogen infection will be established and the crucial nodes in this networks will be identified, which will not only benefit the development of strategy but also can pruovide useful genes for efficient genetic improvement of Solanaceae disease resistance.