5-羟甲基胞嘧啶结合蛋白的鉴定及其对DNA去甲基化的调控和在体细胞重编程中的作用研究

项目名称： 5-羟甲基胞嘧啶结合蛋白的鉴定及其对DNA去甲基化的调控和在体细胞重编程中的作用研究

项目编号： No.91419305

项目类型： 重大研究计划

立项/批准年度： 2015

项目学科： 生物科学

项目作者： 朱冰

作者单位： 中国科学院生物物理研究所

项目金额： 200万元

中文摘要： DNA胞嘧啶甲基化修饰在基因的长期沉默、印记基因的表达调控、X染色体失活和基因组稳定性的维持中起着重要作用。近年来的研究发现， DNA上的甲基化胞嘧啶可以被TET家族蛋白逐步氧化成羟甲基化、醛基化和羧基化的胞嘧啶，最终导致去甲基化。甲基化DNA的氧化产物除了作为去甲基过程的中间产物，其自身也可以排斥或者招募一些DNA结合蛋白。在胚胎干细胞中，5-羟甲基胞嘧啶（5hmC）及其进一步氧化的产物常富集于增强子区域。说明DNA甲基化修饰的动态变化调控，可能参与增强子对基因转录的调控。本申请将利用蛋白质定量质谱技术，鉴定胚胎干细胞特异的5hmC结合蛋白；阐明其选择性结合5hmC的分子机制；利用分子生物学和高通量组学研究5hmC结合蛋白在基因组上的分布，及其与5hmC和TET家族蛋白间的定位关系；此外，还将研究这些蛋白在胚胎干细胞干性维持和体细胞重编程中的作用。

中文关键词： DNA甲基化；TET家族蛋白；DNA去甲基化；SALL4；羟甲基胞嘧啶结合蛋白

英文摘要： 5-methylcytosine (5mC) is an important epigenetic modification that functions in gene silencing, imprinting control, X chromosome inactivation and genome stability control. Recent researches uncovered that TET family enzymes could successively oxidize 5mC to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), which may lead to eventual demethylation by TDG and the base-excision repair pathway. In addition, 5hmC or its further oxidation products may be recognized by putative 5hmC binding proteins. In embryonic stem cells (ESCs), 5hmC and its further oxidated products are often enriched at the enhancers, suggesting that a dynamic regulation of these modifications may play a role in regulating gene expression. In this application, we propose to identify 5hmC binding proteins using quantitative mass spectrometry-assisted DNA affinity purification using mouse ESCs. To explore the in vivo biological functions of our candidate proteins, their genome-wide association and interplay with 5hmC and TET family proteins will be assessed in depth. Given that TET family proteins and DNA oxidation promote cellular reprogramming, we intend to investigate whether our candidate proteins are part of the core transcriptional regulatory network in mouse ESCs and contribute to cell fate decision and re

英文关键词： DNA methylation；TET family proteins；DNA demethylation；SALL4；5-hmC binding protein