弯孢叶斑病菌铁离子通道关键酶对玉米致病性影响研究

Maize (Zea mays L.) is the important food crop,and relates to the food security directly in China. Corn curvularia leaf spot is a major leaf disease, its pathogenicity is the key factor of prevalence. The preliminary studys show that iron is related to pathogenicity of Curvularia lunata. But the relationship between iron uptake systems and pathogenicity of C.lunata is still unknown. Applicants have studied key enzymes of iron acquisition systems,and obtained Nonribosomal Peptide Synthetase 6(NPS6) mutant and Ftr1 knockout vector..We attempt to understand the relationship between iron acquisition systems and pathogenicity of C.lunata by Agrobacterium tumefaciens Mediated Transformation (ATMT), Green Fluorescent Protein(GFP) and tissue dyeing technologies in this project. We analyze the process that corn is infected by mutants NPS6 and Ftr1 of C.lunata, then propose pathogenesis mechanism of C.lunata based on iron nutrition metabolization, and enrich molecular pathogenesis mechanism of C.lunata.For fungal requirement nutrition,it is new idea for openning up environment-friendly fungicides,that made foundation for controlling CuLS effectively.

玉米（Zea mays L.）是主要粮食作物，直接关系到我国粮食安全。玉米弯孢叶斑病（Curvularia lunata (Wakker)Boed.）是重要叶部病害，致病性强弱是影响其流行的关键因素。前期研究初步证明铁离子与玉米弯孢叶斑病菌致病性相关。但铁离子通过何种分子途径影响玉米弯孢叶斑病菌致病性仍缺少研究。申请者对玉米弯孢叶斑病菌铁离子通道关键酶基因进行了深入研究，已获得△nps6和Ftr1基因敲除载体。.本项目拟采用ATMT遗传转化、绿色荧光蛋白标记和组织染色等多项技术，对铁离子通道与玉米弯孢叶斑病菌致病性的关系进行研究，在此基础上分析比较铁载体和RIA途径对玉米弯孢叶斑病菌侵染过程的影响，提出基于铁营养代谢的玉米弯孢叶斑病菌致病机理，进而丰富玉米弯孢叶斑病菌致病分子机制。为从营养代谢角度开发环境友好型抑菌剂提供依据和新思路，从而为有效控制玉米弯孢叶斑病奠定基础。

铁离子是丝状真菌调控致病性的重要营养元素，因此在低铁条件下，研究Curvularia lunata铁离子吸收途径的关键酶基因ClNPS6和ClFtr1作用对于全面揭示C.lunata致病机理具有重要意义。本项目主要以DHN型黑色素生物合成途径中的1,3,8-三羟基萘酚还原酶(Brn1)基因为靶标建立了C.lunata靶基因定向敲除体系，明确了分生孢子萌发时间、共培养温度和时间是提高转化效率的关键条件；ClNPS6、ClFtr1和ClNPS6Ftr1缺失后导致C.lunata的致病力显著下降，在铁离子匮乏和饱和培养条件下培养的C.lunata野生菌株CX-3致病力也下降，证明ClNPS6、ClFtr1是C.lunata保持致病性的关键因子；ClNPS6、ClFtr1促进分生孢子早期萌发和保持细胞壁降解酶中PG和PMTE活性是维持C.lunata致病力的重要机制。ClNPS6缺失后Fit1p、Fit3p和Sit1p基因表达量显著下调，导致铁离子无法从胞外进入胞内参与生理和生化功能。总之，本研究通过研究ClNPS6、ClFtr1的作用与功能，明确了铁离子是维持C.lunata致病性的重要营养元素。