稻瘟病菌一候选效应因子PP1的特异细胞定位及其功能研究

Effectors play crucial roles in the interaction process between the rice blast fungus and rice. However, the study of identification, localization and function of effectors is still in a preliminary stage. During that period, the applicant visited Texas A & M university in United State where the putative effector PP1 has been identified with abundant variations in the gene sequences of its paralogues, and the wheat isolate PP1 has been proved to localize around the appressorium pore and in/around the penetration peg. Based on these discoveries, this project will further clarify the precise localization of this effector via multiple detection approaches and systematically analyze its biological function on the pathogenicity on monogenic rice lines resistant to rice blast, and simultaneously, we will also investigate the corresponding traits of PP1 in the rice isolate FJ81278. Then, the key amino acid residues or motifs will be verified through comparative and associated analysis of the sequence polymorphisms and the diverse localizations and functions of the PP1 paralogs from different isolates. The findings can not only broaden the understanding of the functional mode of effectors in the interaction between pathogen and host, but also lay the theoretical foundation for breeding new blast-resistant rice cultivar.

在稻瘟病菌与水稻互作体系中，效应因子对稻瘟病菌致病性起着极为关键的作用，然而关于效应因子的鉴定、细胞定位及功能的研究仍处于起步阶段。申请人在美国德州农工大学访学研究期间，鉴定得到一个基因序列具有多态性的候选效应因子PP1，并发现小麦瘟菌来源的PP1特异地定位于附着胞孔和侵染栓周围。本项目拟在此基础上，利用多种检测手段明确该PP1效应因子的精确细胞定位，并系统分析该效应因子对单基因系抗瘟水稻的致病特征；同时，我们也将对福建分离的稻瘟菌株FJ81278中PP1蛋白的细胞定位及其对单基因系抗瘟水稻的致病特征进行系统分析。通过比较分析麦瘟菌和稻瘟菌中PP1蛋白序列的多态性及其关联的定位和功能差异，探明决定特异性定位和功能的关键氨基酸残基或基序。研究结果不仅可拓宽对效应蛋白在稻瘟病菌与水稻互作过程中作用机制的认识，也可为培育新型抗瘟水稻品种奠定理论基础。

病原菌向寄主组织中分泌效应子来抑制免疫并促进其自身的生长，进而引起植物病害。然而当前关于效应子的鉴定、细胞定位及功能的研究仍处于起步阶段。本项目对一新鉴定到的效应子PP1（来源于麦瘟菌）进行了深入细致地研究。证实PP1蛋白在病原菌分生孢子接种水稻16小时后起始表达。PP1以不依赖于高尔基体转运的方式分泌到侵染钉和水稻质膜的间隙处，并专一性地定位于侵染钉外围和侵染菌丝穿越临近水稻细胞的穿越位点。PP1基因的启动子及其信号肽共同决定这一特异的定位方式。此外，我们发现Mst12转录因子对PP1基因的转录起重要的调控作用。缺失MST12基因，PP1表达水平显著下降并影响蛋白的合成，最终阻断了附着胞介导的穿透和侵染致病。PP1基因在FJ81278（福建分离）和麦瘟菌中均存在，但二者有11氨基酸位点的差异。有趣的是，这一遗传多样性和效应子的定位无直接关系。RNAi干扰及基因敲除FJ81278中PP1基因，均减弱了稻瘟病菌对水稻的侵染致病能力。综上，这些结果揭示PP1是稻瘟病菌中一个新的效应子，特异定位于侵染钉外围并在稻瘟病的侵染致病过程中发挥重要功能。研究结果不仅可拓宽对效应蛋白在稻瘟病菌与水稻互作过程中作用机制的认识，也可为培育新型抗瘟水稻品种奠定理论基础。