大丽轮枝菌PKS3基因指导双环芳香类聚酮的合成机制

次级代谢物大多是分子结构比较复杂的化合物，主要受聚酮合成酶（PKS）基因所指导合成，是分析化学与分子生物学交叉学科研究的热点。本实验室前期发现，大丽轮枝菌PKS3基因在病原菌侵染寄主初期表达量显著上升，该基因突变后化合物发生明显变化，其中双环芳香类聚酮可引起棉花叶片的萎蔫反应。因此，病原菌PKS3基因指导双环芳香类聚酮的生物合成网络有待深入研究。在已有的工作基础上，本项目拟采用模块重排等技术明确双环芳香类聚酮在病原菌致病力中的作用；利用分段缺失及荧光素报告基因揭示PKS3基因启动子核心区；通过EMSA和酵母单杂交揭示转录因子与其启动子结合区；采用亚细胞定位分析转录因子作用位点；通过缺失和过表达解析其对双环芳香类聚酮的调控方式；利用转录组、qRT-PCR和VIGS技术揭示双环芳香类聚酮与植物的互作信号通路。本研究将有助于进一步阐述病原菌致病机理和为病原菌的综合防治提供重要的化合物及基因资源。

Most secondary metabolites are complicated molecular structure compounds, which are mainly synthesized under the guidance of polyketone synthase (PKS) gene. They are the focus of interdisciplinary research between analytical chemistry and molecular biology. In previous study, we found that the expression level of Verticillium dahliae PKS3 gene during the infected stage. The compound obviously changed after PKS3 was disturbed. Among of them, dicyclic aromatic ketones could induced the cotton leaves to wilt. Therefore, the biosynthesis network of dicyclic aromatic ketones guided by PKS3 gene is urgent to be further studied. Base on the data, we will utilize module rearrangement and other techniques to clarify the role of dicyclic aromatic ketones in the pathogenicity of V. dahliae. The PKS3 promoter core region was revealed by segmenting deletion and fluorescein reporter gene. The transcription factor and its promoter binding region were investigated by EMSA and yeast single hybridization. The transcription factor action sites were analyzed by subcellular localization. The regulation mode of dicyclic aromatic ketones was analyzed by deletion and overexpression of the transcription factor. Furthermore, we investigate the interaction signaling pathways between the two aromatic ketones and plants by transcriptome analysis, qRT-PCR and VIGS techniques. This study will contribute to further elucidate the pathogenic mechanism of V. dahliae and provide important compounds and gene resources for the comprehensive prevention and treatment of pathogens.