染色质修饰酶的活性调节及其对染色质结构和靶基因表达的调控

Chromatin modifications are important structural and regulatory components of chromatin. In the past two decades, many chromatin modifying enzymes were identified. Their physiological and pathological functions have been extensively studied, and many of them became important potential drug targets. However, studies about the enzymatic activity regulation of these chromatin modifying enzymes are relatively weak, especially in comparison to the studies of kinase activity regulation in the signaling pathways. Recently, a few important progresses have been made in the area of enzymatic activity regulation of chromatin modifying enzymes. These studies promote the idea that the regulation of chromatin modifying enzymes is a critical direction for the area of chromatin structure and gene expression regulation.. In this grant, we propose to study the enzymatic regulation of chromatin modifying enzymes, including histone H3K36-specific methyltransferase KMT2H and DNA methyltransferase. The studies include the enzymatic activity regulation of KMT2H by Eaf3 and Eaf3-L1/L2, the molecular mechanisms of the enzymatic activity regulation of KMT2H by Eaf3 and Eaf3-L1/L2, the regulation of KMT2H chromatin localization by Eaf3 and Eaf3-L1/L2, the biological function regulation of KMT2H by Eaf3 and Eaf3-L1/L2. We also propose to study the identification of regulators of DNA methyltransferase, and the role of TPD2 in regulating the enzymatic activity, chromatin localization and biological function of DNA methyltransferase.

染色质修饰是染色质结构的重要组成和调控因素。过去二十余年来，大量的染色质修饰酶得到发现，其生理意义和病理意义的研究也使得许多染色质修饰酶成为药物研发的热点靶标。近来，若干染色质修饰酶活性调节机制方向上取得了重要的进展，使得染色质修饰酶活性调节机制的研究成为染色质结构和基因表达调控研究领域的重要方向。. 本项目拟对组蛋白H3K36甲基化酶KMT2H、DNA甲基化酶等染色质修饰酶的活性调节机制开展研究。研究内容主要包括KMT2H复合体调节亚基Eaf3和Eaf3-L1/L2对KMT2H的催化活性调节和相关机制、Eaf3和Eaf3-L1/L2对KMT2H的染色质定位调节和Eaf3和Eaf3-L1/L2对KMT2H的生理功能调节。研究内容还包括DNA甲基化酶调节蛋白的鉴定、DNA甲基化酶调节蛋白TPD2对DNA甲基化酶活性、染色质定位以及生理功能的调节。

染色质修饰是染色质结构的重要组成和调控因素，近年来的研究发现了大量的染色质修饰酶，染色质修饰酶活性的调节机制方面的研究，成为染色质结构和基因表达调控领域的重要方向。本项目发现KMT2H与Nurf55、Eaf3在体内形成高度保守的蛋白复合体，其中体外实验证明Eaf3可以激活KMT2H的催化活性，而体内实验证明KMT2H募集Eaf3蛋白至两者的共同靶基因并调控靶基因的转录。KMT2H蛋白的第1288位精氨酸是结合Eaf3蛋白的关键氨基酸，携带该突变的果蝇个体表现出包括平衡棒向翅膀部分转化以及第三对足向第二对足部分转化等一系列同源异形转化的表型，同时过表达Eaf3-Nurf55稳定KMT2H-R1288A与Eaf3的互作后同源异形转化的表型得到部分恢复，表明Eaf3对于KMT2H的调节作用是KMT2H作为TrxG蛋白拮抗PcG蛋白所必需的。此外，该项目解析了人源KMT2H-Eaf3的晶体结构，揭示了Eaf3结合并激活KMT2H活性的分子机制：Eaf与KMT2H的SET催化结构域N端一段区域有两个作用界面，两个蛋白通过疏水相互作用和氢键等相互作用紧密结合，Eaf3结合KMT2H后可解除其自抑制状态，使之转化为激活状态。该研究首次发现KMT2H在体内形成高度保守的蛋白复合体，揭示了Eaf3对KMT2H的酶活调节机制及生理功能，对于理解PcG和TrxG蛋白的作用机制及探索组蛋白甲基化酶的活性调节机制都具有重要意义。.在对TPD2调节DNA甲基转移酶DNMT1蛋白稳定性的研究中，我们发现TPD2结合Cullin泛素化酶复合物，介导DNMT1蛋白的降解。TPD2是卵细胞特异表达的蛋白，定位于卵细胞的细胞核。TPD2缺失的卵细胞细胞核会出现DNMT1和UHRF1 的蛋白信号，暗示TPD2通过Cullin蛋白酶解途径，降解卵细胞细胞核中的DNMT1，是保证卵细胞低甲基化组的冗余机制。