细胞色素P450基因CYP709C56上调表达调控看麦娘抗甲基二磺隆的分子机制

看麦娘对甲基二磺隆等常用除草剂的抗性严重威胁我国小麦安全生产，其中代谢抗性是看麦娘产生抗药性的重要原因，但是其内在分子机制尚不清楚。申请人前期研究发现，细胞色素P450s活性增强是看麦娘对甲基二磺隆产生代谢抗性的重要原因之一，而P450基因CYP709C56受甲基二磺隆诱导上调表达且在抗性看麦娘中高表达，推测该基因上调表达调控看麦娘对甲基二磺隆的代谢过程。本项目基于CYP709C56，拟通过异位过表达、基因沉默、酵母体外表达和降解酶活测定明确其解毒甲基二磺隆等除草剂的基因功能，在此基础上分析其启动子功能以探究响应除草剂胁迫的关键顺式作用元件，进而结合酵母单杂交、双荧光素酶报告以及凝胶阻滞迁移率鉴定与之特异性结合的关键转录因子，分析其对CYP709C56的表达调控模式。研究结果将揭示CYP709C56上调表达调控看麦娘抗甲基二磺隆的分子机制，为看麦娘的抗性监测和科学治理提供理论依据。

Resistance to mesosulfuron-methyl and other common herbicides in shortawn foxtail (Alopecurus aequalis Sobol.) severely threatens wheat’s safe production in China. Of them, metabolic resistance is an important reason that leads to herbicide resistance in A. aequalis. However, its internal molecular mechanism remains unclear. Our previous studies have found that the enhanced activity of cytochrome P450s was one of the important reasons for the metabolic resistance of A. aequalis to mesosulfuron-methyl. Also, the P450 gene CYP709C56 could be induced up-regulated expression by mesosulfuron-methyl treatment and was highly expressed in resistant A. aequalis plants. We speculate that the up-regulated expression of CYP709C56 regulated the metabolic process of mesosulfuron-methyl in A. aequalis. Based on CYP709C56, this study intends to analyze its gene function on detoxification of mesosulfuron-methyl and other herbicides through ectopic overexpression, gene silencing, and in vitro yeast expression and degradation enzyme activity assay. On this basis, the function of CYP709C56 promoter will be analyzed to investigate the key cis-elements in response to herbicide stress. Furthermore, the key transcription factors interacting with the promoter will be identified by performing yeast one-hybrid assay, dual-luciferase reporter assay, and electrophoretic mobility shift assay, and its transcriptional regulation patterns on CYP709C56 expression can be further investigated. These results will reveal the molecular mechanism of up-regulated expression of CYP709C56 regulating A. aequalis resistance to mesosulfuron-methyl, providing theoretical basis for herbicide resistance monitoring and scientific management of A. aequalis.