黄萎病是棉花生长过程中最具有毁灭性的病害之一，严重影响棉花的产量和品质。开展棉花抗黄萎病分子机制研究，对利用基因工程手段提高陆地棉黄萎病抗性有重要意义。4-香豆酰-CoA连接酶（4CL）是苯丙烷类衍生物代谢途径的关键酶，调控苯丙烷衍生物代谢方向，与多种抗菌物质代谢有关。项目前期研究发现，黄萎病病菌诱导后，棉花Gh4CL1基因表达显著提高。抑制Gh4CL1基因表达可显著提高棉花抗病及感病品种对黄萎病抗性，同时发现棉株茎秆中有明显红色物质积累。本项目拟在此基础上，通过对黄萎病菌侵染前后不同时段棉花对照、Gh4CL1过表达、Gh4CL1抑制表达和基因编辑敲除材料的转录组分析、抗病信号通路相关基因的表达分析、防御相关的次生代谢产物的含量变化分析，以及对Gh4CL1相互作用蛋白的分析，明确Gh4CL1基因在棉花抗黄萎病中的功能及调控分子机制，为利用基因工程手段提高棉花对黄萎病抗性奠定基础。

Verticillium wilt is one of the most destructive diseases in Upland cotton, which severely affects the yield and quality of cotton. Understanding the molecular mechanism of Verticillium wilt resistance in cotton is of great significance for improving Verticillium wilt resistance in Upland Cotton by genetic engineering. 4-coumarin-CoA ligase (4CL) is a key enzyme in the metabolic pathway of phenylpropane derivatives. It regulates the outcomes of the metabolic pathway of phenylpropane derivatives and is related to the metabolism of various antimicrobial substances. Our previous study found that the expression level of the Gh4CL1 gene was significantly increased in cotton seedlings infected by Verticillium dahliae (Vd). Inhibition of Gh4CL1 expression can significantly improve cotton resistance to Verticillium in both Vd-resistant and Vd-susceptible cotton varieties, and a large amount of red color metabolites was found to be accumulated in the stem of the Gh4CL1-silenced cotton plants. To investigate the molecular bases of the observed phenomena caused by the down-regulation of the Gh4CL1 expression level, this study will generate stable transgenic cotton plants with a down-regulated level of Gh4CL1 using RNAi and/or genome editing as well as transgenic overexpressing Gh4CL1. The generated transgenics will be used as experimental materials to investigate their responses to Vd infection in comparison to the wild-type plants. The function and molecular mechanism of Gh4CL1 in response to Verticillium dahlia will be analyzed by investigation of transcriptome, the expression of defense-related genes and signaling pathway, the content changes of defense-associated secondary metabolites and proteins interacting with Gh4CL1. We expect to uncover the gene networks responsible for the increased Vd resistance observed in the Gh4CL1-silenced cotton plants and the molecular mechanisms underlying Gh4CL1-mediated Vd-response in cotton, and to finally provide strategies for genetic improving cotton’s resistance to Verticillium wilt.