A multidisciplinary approach to elucidate virulence functions of oomycete effetor proteins

植物病原菌为了对寄主植物致病，必须抑制植物的天然免疫。细菌和丝状植物病原体通过将专门的效应蛋白转运到宿主细胞中来操纵植物的免疫反应来实现这一点。本研究旨在阐明感染拟南芥的卵菌病原体阿拉伯透明霜霉（Hyaloperonospora arabidopsidis，Hpa）的效应蛋白的毒力功能。我将使用一个多学科的方法结合结构生物学，分子生物学和植物病理学，揭示卵菌效应如何抑制植物免疫。该提案的轨道I采用X射线晶体学的结构生物学方法来阐明抑制植物防御的Hpa效应物的三维结构。轨道II侧重于识别这些效应器的宿主靶点。已经在酵母双杂交（Y2H）筛选中鉴定了许多候选效应子靶标。这些相互作用将在体外和植物细胞中得到验证。来自拟南芥组织的Hpa效应子的亲和纯化和通过质谱鉴定共纯化蛋白质将用于那些在Y2H中不拾取相互作用蛋白质的效应子。这些综合努力将使人们深入了解Hpa如何操纵植物防御系统，并将确定植物免疫信号的新节点。

In order to cause disease on host plants, plant pathogens have to suppress plant innate immunity. Bacterial and filamentous plant pathogens accomplish this by transporting specialised effector proteins into host cells that manipulate the plant s immune response. This proposal aims to unravel the virulence functions of effector proteins from the oomycete pathogen Hyaloperonospora arabidopsidis (Hpa), that infects Arabidopsis thaliana. I will use a multidisciplinary approach combining structural biology, molecular biology and plant pathology to reveal how oomycete effectors suppress plant immunity. Track I of this proposal takes a structural biology approach by X-ray crystallography to elucidate the threedimensional structures of Hpa effectors that suppress plant defence. Track II focusses on identification of host targets of these effectors. A number of candidate effector targets have been identified in yeast-two-hybrid (Y2H) screens. These interactions will be validated in vitro and in plant cells. Affinity purification of Hpa effectors from Arabidopsis tissue and identification of co-purifying proteins by mass spectrometry will be employed for those effectors that did not pick up interacting proteins in Y2H. These combined efforts will give insights into how Hpa manipulates the plant defence system and will identify new nodes of plant immune signalling.