基于冷冻电镜的组蛋白H3K4甲基转移酶COMPASS复合体的结构与功能研究

Methylation of histone H3 lysine 4 (H3K4) is catalyzed by the multi-component COMPASS complex, which is highly conserved from yeast to human, and plays essential roles in gene expression and transcription, cell cycle progression, and DNA repair. COMPASS complex have been the most intensively studied because of its involvement by chromosomal translocations in a variety of acute lymphoidand myeloid leukaemias and several types of human tumours. To identify the structural elements underpinning COMPASS assembly, several crystal structures of the key components and subcomplexes have been determined and provided information about its subunit interaction network. However, due to the lack of the complete structure of COMPASS, the entire spectrum of interactions controlling COMPASS assembly remains unclear, which hinders our understanding of the mechanisms underlying its substrate recognition and H3K4 methylation. We have overexpressed all the seven subunits of COMPASS in yeast, and for the first time, we determined the complete structure of S. ceravisa COMPASS by cryo-EM. By utilizing the emerging powerful cryo-EM technique, the research objectives include: To determine the high resolution structure of complete yeast COMPASS complex, reveal the subunit organization and full molecular architecture of yeast COMPASS; to uncover the interaction mechanism of nucleosome core particle with COMPASS complex, and reveal the mechanism of histone H3K4 methylation, which could provide essential structure basis for the diagnosis and treatment of related human diseases.

组蛋白H3第四位赖氨酸（H3K4）三甲基化是转录起始的标志，酵母中该过程由甲基转移酶COMPASS复合体催化完成。该复合体从酵母到人高度保守。人源COMPASS亚基的缺失或突变将引起白血病、肿瘤等诸多人类疾病。COMPASS组成复杂且极具动态性，给结构生物学研究带来极大挑战，至今尚无该复合体完整结构信息。这也极大阻碍了人们对COMPASS复合体的整体构架结构，各亚基如何组装，如何相互协调完成底物核小体识别与结合，进而完成组蛋白H3K4甲基转移酶活性等科学问题的深入理解。本项目将在酵母中超表达并纯化酵母COMPASS复合体，解析完整COMPASS复合体及其结合核小体形成复合体的高分辨率冷冻电镜三维结构，以期揭示其亚基排布方式和相互作用网络，及其识别底物核心核小体并且催化H3K4甲基化的分子机制，为相关白血病或肿瘤等疾病的诊疗提供新思路和靶标。

组蛋白H3第四位赖氨酸（H3K4）甲基化由组蛋白甲基转移酶复合体COMPASS催化完成。该复合体从酵母到人高度保守，人源COMPASS复合体亚基的缺失或突变将引起白血病、肿瘤等人类疾病。COMPASS组成复杂且极具动态性，给结构生物学研究带来极大挑战。至今尚无该复合体的完整结构，限制了对COMPASS复合体的整体构架结构，各亚基如何相互协调识别底物核小体、进而完成组蛋白H3K4甲基转移酶活性机制的理解。本项目研究进展如下：（1）首次解析了完整的酵母COMPASS 复合体的冷冻电镜结构，应用自行发展的复合体亚基标签定位方法结合交联质谱，揭示了COMPASS复合体亚基排布方式及其作用网络（Sci Rep 2018, 见附件，已致谢本项目）；通过优化蛋白纯化和制样条件以及数据处理方法，已推进完整COMPASS的较高分辨率冷冻电镜结构解析（文章撰写中）。（2）完成了不同物种及Ub修饰状态的核小体核心颗粒制备，组装并解析了完整COMPASS结合核小体形成复合体的冷冻电镜结构；（3）构建了一系列Set1亚基N端截短的COMPASS复合体表达菌株，及调节亚基外源表达菌株，展开基于结构的生化和功能验证；（4）发展了冷冻电镜亲和载网方法（文章撰写中），便于高效原位纯化完整COMPASS复合体。上述研究为揭示完整COMPASS识别核小体并催化H3K4甲基化的分子机制奠定基础，为相关白血病或肿瘤的诊疗提供新思路。（5）本团队还在新冠病毒及其值得关注突变株的结构及其受体结合机制和抗体发展等课题中取得系列成果（Nature 2022; Nat Commun 2021a,b, 2022a; Sci Advances 2021等）。受本项目资助，在Nature，Mol Cell，Sci Advances，Nat Commun，PNAS，Sci Rep等国际学术期刊发表通讯或共同通讯作者论文12篇。