OsEPSPS3调控水稻稻瘟病抗性的分子机制

Rice blast, caused by Magnaporthe oryzae (M. oryzae), is one of the destructive diseases leading to large losses in yield and quality of rice, the main staple food for half population of the world. It’s important to improve blast resistance to ensure the food security. It is well known that 5-Enolpyruvylshikimate-3-phosphate (EPSP) synthases are mainly involved in the synthesis of EPSP in planta. Whether EPSP synthase regulates innate immunity remains undetermined. Our previous study has identified that the OsEpsps3 gene from EPSP family in rice is highly induced in expression by M. oryzae. The OsEpsps3RNAi transgenic plants generated by RNA interfere approach in silence of OsEpsps3 exhibit enhanced susceptibility compared with the wild type rice, suggesting that the EPSP synthase gene OsEpsps3 positively regulates rice blast resistance. This study proposes these aims: (1) generate OsEpsps3 knock-out (OsEpsps3KO) and overexpression (OsEpsps3OE) transgenic plants and test their blast resistance; (2) determine and compare the dynamic infection processes of M. oryzae among the OsEpsps3RNAi, OsEpsps3KO and OsEpsps3OE plants with the wild type rice; (4) biochemically characterize the OsEPSPS3 enzymatic activity on EPSP synthesis; and (5) screen the antimicrobial metabolites or hormone regulated by OsEPSPS3 pathway and determine their regulation on blast disease resistance. Through this study, we will elucidate the molecular mechanism of OsEpsps3 on rice blast resistance, provide novel insights into rice immunity and offer novel approaches for breeding disease resistant rice and developing ecological pesticides.

稻瘟病严重危害水稻的产量和品质，提高水稻对稻瘟病的抗性是粮食安全的重要保障。通过转录组分析，我们发现水稻中一个5-烯醇式丙酮酰莽草酸-3-磷酸合酶（EPSP合成酶）编码基因OsEpsps3特异地受稻瘟病菌诱导表达。利用RNAi技术抑制该基因表达后，水稻植株更感稻瘟病。该结果表明EPSP合成酶OsEPSPS3对稻瘟病抗性起正调控作用。本项目拟开展如下研究：（1）通过转基因敲除和过量表达试验，对该基因的稻瘟病抗性调控作用进行印证；（2）观察比较转基因植株和野生型间稻瘟病菌侵染过程的差异；（3）明确OsEPSPS3的生化功能和特点；（4）筛选下游受影响的抗菌代谢物或激素并分析差异代谢物或激素的抗菌机理。在此基础上阐述OsEPSPS3调控稻瘟病抗性的分子机制。由于尚未有EPSP合成酶对植物免疫和抗性调控研究方面的报道，本项目的研究将丰富水稻抗病分子机理，为水稻抗病育种以及开发生态农药提供理论依据。

稻瘟病号称水稻“癌症”，严重威胁水稻的产量和品质，利用水稻自身抗性来应对该病害既经济有效又生态环保。项目前期发现水稻莽草酸代谢途径中一个关键酶——EPSP合成酶编码基因OsEPSPS3在受到稻瘟病侵染的水稻中表达量显著升高，说明该基因及其相关的代谢过程在响应稻瘟病入侵的过程中具有重要的作用，但尚未见与之相关的报道。项目以此为基础，通过构建OsEPSPS3基因干涉和过量表达水稻材料，接种稻瘟病菌，证明了OsEPSPS3基因对稻瘟病抗性的负调控作用；鉴定了OsEPSPS3基因表达量抑制的水稻材料对稻瘟病的抗谱，发现该水稻材料具有广谱稻瘟病抗性；通过细胞学观察，比较了稻瘟病菌侵染OsEPSPS3基因干涉材料和野生型水稻时的差异，发现稻瘟病菌在侵染该基因干涉水稻时，附着胞和侵染钉等侵染结构的形成较慢，侵染菌丝在细胞间的扩增速度降低，从细胞学水平揭示了该材料稻瘟病抗性增强的机制；通过比较代谢组分析，鉴定了在OsEPSPS3基因干涉材料中特异富集的代谢物，筛选鉴定到代谢物A和代谢物B对水稻稻瘟病菌具有明显的抑制作用和田间防治效果。项目研究成果发现了一个稻瘟病抗性调控新基因并解析了可能的调控机理，丰富了水稻抗病分子理论；研究还鉴定到了两种来自水稻自身、对稻瘟病具有防治作用的天然代谢产物，为后续深入的抑菌机理研究提供了重要材料，也为开发新型绿色生态农药奠定了坚实的基础。