水稻水通道PIP蛋白识别与传导黄单胞菌Hpa1蛋白信号的机制

Plant aquaporin proteins basically mediate water transport through cell membranes, and also affect plant-microbe interactions and plant defense responses. Molecular mechanisms that underlie the dual function are unclear. Recently we have determined that rice and Arabidopsis aquaporins OsPIP1;2 and AtPIP1;4 interact with Hpa1, a protein secreted by type III pathway in Xanthomonas oryzae pathovars oryzae and oryzicola, which cause bacterial blight disaese and bacterial leaf streak disease of rice, respectively. When applied to plants or produced in transgenic plants, Hpa1 localizes to the apoplast and induces hydrogen peroxide generation in the same cellular location. The generation of hydrogen peroxide is dependent of the NADPH oxidase located at the plasma membrane. In response to Hpa1, moreover, the apoplastic hydrogen peroxide translocates to the cytoplasm with subcequent effects of enhancing plant defense responses to bacterial pathogens. The present proposal is composed for studies that will be done based on established models in regard to the topological stucture of aquaporins, the performance of bacterial type III secretion, and subcellular trafficking of hydrogen peroxide in plants. We will identify the functional domain for OsPIP1;2 interacting with Hpa1, and elucidate molecular features and structural basis of the OsPIP1;2-Hpa1 interaction. Then, we will investigate the role of OsPIP1;2-Hpa1 interaction in modulating translocation of apoplastic hydrogen peroxide to the cytoplasm and thus connect the signal translocation with defense responses in rice. Next, we will explore whether the OsPIP1;2-Hpa1 interaction plays a role in facilitating the translocation of X. oryzae type III effector proteins from the bacterial cell to the plant cell, and how the effector translocation, if occurs, affects the bacterial pathogenesis. The ultimate goal of these studies is to further reveal regulatory mechanisms of plant-pathogen interactions, offerring a significant extension of aquaporins' functions over what we have known up to date.

最近有研究发现，植物水通道蛋白PIP不仅担负细胞间或细胞内外水分子输导的基本功能，还参与植物-微生物互作与植物防卫反应，这种双重功能的调控机制备受关注。我们研究表明，水稻PIP1;2和拟南芥PIP1;4可以与水稻黄单胞III型泌出蛋白Hpa1互作，Hpa1定位于植物细胞的质外体，诱导过氧化氢在质外体产生及向原生质转运，进而影响植物防卫反应与对病原细菌的抗性。根据植物水通道蛋白拓扑结构与病原细菌III型分泌系统工作模型，本课题拟鉴定水稻PIP1;2与Hpa1互作的功能域，试验阐明互作的特征与分子基础；揭示PIP1;2-Hpa1互作对过氧化氢信号从植物细胞的质外体向原生质转运的调控作用及对植物防卫反应的影响；试验探究PIP1;2-Hpa1互作对水稻黄单胞菌III型效应蛋白从细菌细胞向植物细胞转运的调控作用。研究结果将深入阐释植物-病原物互作调控机制，为水通道蛋白功能调控提供新的见解。

本课题拟鉴定水稻细胞外膜上的水通道蛋白质 PIP 与白叶枯病菌III型转位子蛋白质 Hpa1 互作的功能域，试验阐明互作的特征与分子基础；揭示 PIP-Hpa1 互作对过氧化氢信号从植物细胞的质外体向原生质转运的调控作用及对植物防卫反应的影响；试验探究 PIP-Hpa1 互作对水稻黄单胞菌 III 型效应蛋白从细菌细胞向植物细胞转运的调控作用。研究结果将深入阐释植物-病原物互作调控机制，为水通道蛋白功能调控提供新的见解。. 包括水稻白叶枯病菌在内的革兰氏阴性细菌借助III型分泌系统向真核寄主胞内注射毒性或无毒效应蛋白，但穿透寄主细胞膜的转运过程还需要转位子（translocon）的协助。转位子通常由疏水性与亲水性转位子蛋白（translocators）组成，在寄主细胞膜上还存在能够识别转位子的受体分子，受体分子的性质及其在效应蛋白转运过程中的作用尚无研究。本项目发现，水稻白叶枯病菌III型转位子蛋白Hpa1与水稻水通道蛋白OsPIP1;3之间存在特异性互作，调控类转录激活子效应蛋白PthXo1与AvrXa10从病菌细胞向水稻细胞内转运。而进一步的研究发现，OsPIP1;3不影响Hpa1Xoo激活的PTI与SAR信号通路。上述研究结果深入解析了Hpa1-OsPIP1;3互作调控水稻与白叶枯病菌抗感性互作的机理。. 过氧化氢的产生是植物成功识别病原物相关分子模式(PAMP) 与病原物的标志，而病原物或PAMPs诱导产生的过氧化氢的通常在植物细胞胞外(外质体)积累，然后通过细胞外膜上的特殊通道进入细胞质，参与胞内免疫信号通路从而调控抗病性，如 SAR和 PTI。前人研究表明，植物细胞膜通道可能包括水通道蛋白质的作用。本项目在拟南芥上进行研究发现，一种细胞膜嵌入蛋白质AtPIP1;4具有介导过氧化氢的从细胞外向细胞质转运的作用。AtPIP1;4的这一功能，在植物细胞外质体内诱导产生的过氧化氢的与其对免疫反应的调控作用之间，建立了前后关联的功能关系。