OsCdc48与Pik1-H4的互作机制及对稻瘟病抗性的调控  

Interaction mechanism between OsCdc48 and Pik1-H4,and its regulation on rice blast disease resistance

作　　者：黄其伟 张丽娜 王加峰 陈淳 HUANG Qiwei;ZHANG Lina;WANG Jiafeng;CHEN Chun(College of Agriculture,South China Agricultural University/National Engineering Technology Research Center of Plant Space Breeding/Guangdong Provincial Key Laboratory of Plant Molecular Breeding,Guangzhou 510642,China)

机构地区：[1]华南农业大学农学院/国家植物航天育种工程技术研究中心/广东省植物分子育种重点实验室,广东广州510642

出　　处：《华南农业大学学报》2025年第2期164-174,共11页Journal of South China Agricultural University

基　　金：广东省科技计划(2023B0202010003);广东省自然科学基金(2023A1515012101)。

摘　　要：【目的】探究Pik1-H4与细胞周期蛋白OsCdc48的互作机制,明确其在稻瘟病抗性中的作用。【方法】首先,分别利用酵母双杂交试验和荧光素酶互补试验验证Pik1-H4与OsCdc48的相互作用;然后,通过RT-qPCR分析OsCdc48在稻瘟病菌侵染后的表达情况及组织表达特异性;其次,分析OsCdc48的序列保守性、蛋白结构域、系统进化关系、蛋白质三维结构预测及其亚细胞定位;最后,利用CRISPR/Cas9创制OsCdc48突变体,并对其进行稻瘟病抗性鉴定和病程相关基因表达分析。【结果】证实了Pik1-H4与OsCdc48的互作,且OsCdc48受稻瘟病菌侵染诱导表达。OsCdc48在各组织中均有表达,其编码蛋白定位于细胞核与细胞质。OsCdc48在不同物种中序列保守,与玉米和高粱的亲缘关系最近,预测会形成同源六聚体。OsCdc48功能缺失突变体kooscdc48病程相关基因上调表达,对稻瘟病抗性增强。【结论】本研究为深入揭示OsCdc48与NLR蛋白Pik1-H4调控稻瘟病抗性的机制及水稻抗病育种提供了理论基础。【Objective】To investigate the interaction mechanism between Pik1-H4 and cell division cycle protein OsCdc48,clarify its role in rice blast disease resistance.【Method】Firstly,the interactions between Pik1-H4 and OsCdc48 were verified using yeast two-hybrid and luciferase complementation assays.Then,the expression patterns of OsCdc48 after infection of Magnaporthe oryzae and its tissue-specific expression were analyzed by RT-qPCR.Next,the determination of OsCdc48’s sequence conservation,protein domains,phylogenetic relationships,protein 3D structure prediction and subcellular localization were performed.Finally,OsCdc48 mutant was created using CRISPR/Cas9 technology,the resistance identification of transgenic mutant to rice blast disease and expression analysis of pathogenesis-related genes were also conducted.【Result】The interactions between Pik1-H4 and OsCdc48 were confirmed,and OsCdc48 was induced by rice blast fungus infection.OsCdc48 was expressed in all tissues and was localized to the nucleus and cytoplasm.The sequence of OsCdc48 was conserved across different species,with the closest phylogenetic relationship to maize and sorghum,and it might form homologous hexamers.The OsCdc48 loss-of-function mutant ko-oscdc48 up-regulated the expression of disease-related genes and enhanced the resistance to rice blast disease.【Conclusion】This study lays a theoretical basis for further elucidating the mechanism of OsCdc48 and NLR protein Pik1-H4 regulating blast disease resistance and rice disease-resistant breeding.

关 键 词：水稻 稻瘟病 稻瘟病菌 Pik1-H4 OsCdc48 

分 类 号：Q78[生物学—分子生物学] S435.111.1[农业科学—农业昆虫与害虫防治]