疫霉菌新型效应蛋白SCR2调节植物免疫反应的作用机制研究

Phytophthora infestans is the casual agent of potato late blight disease which is one of the most destructive crop diseases in China. It has been well known that Phytophthora pathogens delivered a variety of effector proteins into host cell to manipulate host physiological and immune process. The study on effector biology is important for understanding the disease mechanism and plant resistance. Previously, high-throughput screening discovered SCR2, a Phytophthora conserved effector protein that triggers host immune response. Furthermore, inhibition assay uncovers that plant serine proteases are required for SCR2 function. However, the molecular mode of action of SCR2 and the interplay with serine proteases remains unclear. In this proposal, we aim to capture SCR2 host targets by Co-immunoprecipitation followed by peptide identification by LC-MS/MS. The functional study on target protein and the way how SCR2 manipulate target protein will be beneficial to understand the virulence mechanism of SCR2. Meanwhile, genetic screening will be performed to verify the plant serine protease that is required for SCR2 function. Hypothesis that SCR2 maturation by proteases is important for SCR2 triggered immunity will be tested. A model how protease participate effector mediated plant immunity will be expected. In summary, the results will not only help us to understand the molecular mode of action of a conserved Phytophthora effector, but also provide novel insight for screening natural resistance and engineering durable resistant crops.

由致病疫霉菌引发的晚疫病是我国农业生产中的毁灭性病害之一。效应子是致病疫霉菌向植物细胞输送的一类分泌蛋白，具有调节寄主生理和免疫反应的重要功能，是解析病害发生机制和植物抗病性状的关键。申请者前期筛选到一个能激活植物免疫反应的致病疫霉菌效应子SCR2，发现SCR2在疫霉菌种间具广谱保守性，其激活过程需要植物丝氨酸蛋白酶参与，但是SCR2具体的生化功能及其作用机制尚不清楚。本项目拟一方面利用免疫沉淀结合蛋白质谱的方式筛选SCR2靶蛋白，探究靶蛋白在植物免疫中的生物学功能及其受SCR2效应子调节的作用方式，明确SCR2的毒性机理；另一方面拟通过遗传筛选鉴定参与SCR2激活植物免疫反应的丝氨酸蛋白酶，验证蛋白酶通过切割SCR2从而激活免疫反应的假说，以期阐明一种蛋白酶与效应子互作的新模式。研究结果有望揭示一类广谱效应子的生化功能和作用机制，并为抗晚疫病的作物分子设计育种和抗病资源筛选提供科学依据。

受本基金项目资助，我们发现了一种由致病疫霉分泌的富含半胱氨酸的小分泌蛋白SCR2（又称PC2，下文中均称PC2），进一步研究发现植物丝氨酸蛋白酶P69B能够切割PC2蛋白，而这种切割活性能够被丝氨酸蛋白酶化学抑制剂阻断。在番茄中沉默P69B或在烟草中沉默其同源基因能够减弱PC2诱导免疫反应的强度。本项目发现植物丝氨酸蛋白酶P69B及其同源物能够切割PC2，而PC2诱导植物免疫的活性依赖丝氨酸蛋白酶P69B的活性。本研究鉴定到一个新型的胞间SCR类效应蛋白PC2，并以此为研究材料开展了PC2诱导植物免疫反应机制的研究，研究结果初步揭示一类广谱效应子的生化功能和作用机制，有望为晚疫病的抗病资源筛选和作物抗病育种设计提供科学依据。项目资助成果在New Phytologist、PNAS等学术期刊上发表。