Molecular analysis of virulence and avirulence activities of the bacterial effector protein XopH and related proteins

微生物植物病原体对全球粮食安全构成了一条永久的线索。植物病原微生物通过III型(T3)分泌系统将效应蛋白注入宿主细胞，这对致病力至关重要，因为它们操纵植物细胞生理，使病原体受益。一些原本敏感的寄主物种的某些植物基因型含有抗性(R)蛋白，这些蛋白识别微生物效应器，进而激活植物的防御系统。被植物免疫系统识别的效应器被称为无毒蛋白(AVR)，因为它们的存在将毒力转化为无毒菌株。对AVR-R相互作用的分子分析是阐明植物免疫的机理基础，并为基于知识的抗虫作物育种提供关键。最近的研究表明，Xe中的XopH能使植酸盐(肌醇六磷酸，InsP6)脱磷，植酸盐是植物中主要的磷储存化合物。XopH是第一个被证明具有植酸酶活性的T3效应器，也是自然界中第一个被描述的1-植酸酶(在C1位去磷酸化)。植物含有大量的肌醇多磷酸盐，目前尚不清楚哪些依赖于XopH的肌醇多磷酸盐促进疾病。XopH还可以在含有同源R基因BS7的辣椒植株中引发免疫反应。初步工作表明，BS7检测的是XopH的生物活性，而不是结构。下面的提议有两个截然不同但相互关联的目标。1)从生化、细胞和结构水平研究XopH及其相关微生物效应因子。这里的一个主要目的是鉴定依赖XopH的肌醇多聚磷酸盐，它促进疾病和/或触发植物防御。2.)用定位克隆的方法分离辣椒R基因BS7。具有肌醇多磷酸水解酶活性的微生物T3效应器及其匹配的宿主R蛋白的可用性为首次阐明这种活性如何转化为植物免疫反应提供了独特的机会。拟议的研究将在Lahaye实验室(Tübingen)和Schaaf实验室(波恩)的合作下进行。Schaaf实验室在肌醇衍生代谢物的生化分析方面拥有全面的专业知识。拉哈耶实验室在微生物效应器的功能分析和从作物物种中定位克隆植物R基因方面拥有长期经验。我们预计，两个实验室的互补专业知识将产生强大的协同效应，使人们能够从机制上深入了解XopH和相关蛋白质的毒力和无毒活动，如果没有这种协调一致的努力，这是不可能的。

Microbial plant pathogens pose a permanent thread to global food security. Phytopathogenic microbes inject effector proteins into host cells via type III (T3) secretion systems, which are crucial to virulence since they manipulate plant cell physiology to the pathogen’s benefit. Some plant genotypes of otherwise susceptible host species contain resistance (R) proteins that recognize microbial effectors and in turn activate the plant’s defense system. Effectors that are recognized by the plant immune system have been coined as avirulence (Avr) proteins, since their presence converts virulent into avirulent strains. Molecular analysis of Avr-R interactions is imperative to elucidate the mechanistic basis of plant immunity, and provides the key for knowledge-based breeding of pest-resistant crops.We intend to study virulence and avirulence activities of XopH, an effector from the pepper and tomato pathogen Xanthomonas euvesicatoria (Xe). Recent studies revealed that XopH from Xe dephosphorylates phytate (myo-inositol-hexakisphosphate, InsP6), the major P storage compound in plants. XopH is the first T3-effector for which phytase activity has been demonstrated and is the first 1-phytase described in nature (dephosphorylation at position C1). Plants contain numerous inositol polyphosphates, and it remains to be clarified which of the XopH-dependent inositol polyphosphates promotes disease. XopH also triggers an immune reaction in pepper plants containing the cognate R gene Bs7. Preliminary work suggests that Bs7 detects the biological activity, rather than the structure of XopH.The following proposal has two distinct but interconnected aims. 1.) To study XopH and related microbial effectors at the biochemical, cellular and structural level. One major aim here is the identification of XopH-dependent inositol polyphosphates that promote disease and/or trigger plant defense. 2.) To isolate the pepper R gene Bs7 via positional cloning. Availability of a microbial T3 effector with inositol polyphosphate hydrolase activity and its matching host R protein provides the unique opportunity to elucidate for the first time how such an activity can translate into a plant immune reaction.The proposed research will be carried out in a collaborative effort of the Lahaye lab (Tübingen) and the Schaaf lab (Bonn). The Schaaf lab has comprehensive expertise in the biochemical analysis of myo-inositol-derived metabolites. The Lahaye lab has long-term experience in the functional analysis of microbial effectors and the positional-cloning of plant R genes from crop species. We expect that the complementary expertise of both labs will generate strong synergistic effects that will allow a deep mechanistic understanding of virulence and avirulence activities of XopH and related proteins that would not be possible without this concerted collaborative effort.