以Set1为核心的COMPASS甲基转移酶复合体调控黄曲霉毒素合成的分子机理

黄曲霉毒素是毒性和致癌性最强的真菌毒素，严重威胁食品安全，却难以早期防控。我们前期工作发现敲除黄曲霉菌中的组蛋白赖氨酸甲基转移酶Set1后，黄曲霉毒素合成被抑制了，提示以Set1为核心酶的COMPASS组蛋白甲基转移酶复合体参与了黄曲霉毒素合成的调控，但是其分子机制尚不清楚。本研究拟采取免疫共沉淀和酵母双杂交等技术分析黄曲霉菌中构成COMPASS复合体的蛋白亚基及其结构，通过晶体X-射线衍射分析Set1分子结构，利用基因敲除、结构域敲除和点突变等方法确定各亚基与黄曲霉毒素合成的关系；运用ChIP-seq等手段淘选COMPASS甲基转移酶复合体调控的组蛋白所在DNA靶位点，及其分布规律，并结合RNA-seq等技术揭示COMPASS甲基转移酶复合体调控黄曲霉毒素合成的分子机制。本研究为早期防控黄曲霉毒素污染提供理论依据，并为真菌次生代谢表观遗传调控机制提供重要参考。

Aflatoxins mainly produced by Aspergillus flavus are the most toxic and carcinogenic known mycotoxins. Aflatoxins popularly contaminate crops, fresh milk and dairy products, pickled foods, fermented foods, and nuts between 35 degrees of north and south latitude, and they seriously threaten the food safety of human beings and animals, and the contamination caused by them is hard to be removed and early prevented. Our previous work has found that the synthesis of aflatoxins from A. flavus was inhibited when histone lysine methyltransferase Set1 was absent. The results of our work revealed that the epigenetic factor - Set1 mediated histone lysine methylation modification is deeply involved in the regulation of aflatoxins synthesis, and the detail molecular regulatory mechanism about it needs to be clarified urgently. This project is designed to screen the protein subunits of the COMPASS (Complex of Proteins Associated with Set1), and to construct the structure model of the complex in A. flavus through Co-Immunoprecipitation (Co-IP) and yeast two-hybrid (Y2H), to anaylze the melecular structure of Set1 by X-ray crystallography, and to reveal the relationship between the protein subunits of the COMPASS complex and aflatoxins synthesis by gene knockout, domain knockout and point mutation operations. In this project, ChIP-seq and ChIP-q-PCR are chosen to identify the target chromatin framents and target genes，and the distribution patterns of these target chromatin framents. RNA-seq or qRT-PCR would be performed to revealed the expression pattern of the target genes under the regulation of the COMPASS complex idetified by the analysis of ChIP-seq and ChIP-q-PCR. The project will reveal the molecular mechanism of the COMPASS methyltransferase complex regulating aflatoxins synthesis, and will supply theoretical basis for the early prevention and control of aflatoxins contamination, which will provide important reference for the epigenetic regulation mechanism in fungal secondary metabolism.