组蛋白及其变体H2A.Z和H3.3体外装配核小体的动力学特征研究

The nucleosome is the fundamental repeating unit of eukaryotic chromatin. In addition to the canonical histones, many eukaryotes possess histone variants, especially variant H2A.Z and H3.3. By changing the access of trans-acting factors to their template and controlling the degree of chromatin opening or folding, nucleosome assembly regulates many biology processes occurred in chromatin level. High dynamic processes of nucleosome assembly, removal and sliding are known to be affected by many factors in vivo. Reconstituted nucleosome with canonical histone and histone variant on different DNA templates in vitro is an effective method to investigate the dynamical characteristics of nucleosome assembly. In this project, considering the intrinsic sequence preference to form canonical nucleosome, sequence feature of ±1 nucleosome region flanking TSS, promoters and enhancers deposited histone variant, the nucleosome assembly, removal and competition dynamics of canonical histone, variant H2A.Z and H3.3 is analyzed. The proposed research project also attempt to establish the dynamic model of canonical nucleosome assembly. This work will lead to a better understanding of nucleosome assembly and positioning in vivo, chromatin dynamics, histone variant dynamics and function.

核小体是构成真核生物染色质的基本结构单元，H2A.Z和H3.3是体内常见的组蛋白变体。通过影响反式调控因子与顺式序列元件的结合以及改变染色质结构开放或压缩状态，核小体组装调控着几乎所有DNA层次上的生物学过程。体内核小体组装、解聚及滑动等高度动态变化的过程受到诸多因素的综合影响。利用体外实验系统研究组蛋白及其变体H2A.Z和H3.3装配核小体过程的动力学特征，是阐明组蛋白及其变体装配核小体规律较为有效的手段。本项目拟结合影响常规核小体装配的DNA序列特征以及富集组蛋白变体的启动子、增强子和转录起始位点附近±1核小体区域DNA序列特征，多角度分析组蛋白及其变体H2A.Z和H3.3体外核小体装配、解聚以及竞争等过程的动力学规律与差异性，尝试构建常规核小体组装过程动力学模型。项目的开展期望能够为体内核小体装配及定位、染色质动力学、组蛋白变体动力学及功能等相关问题的研究奠定一定的基础。

核小体是构成真核生物染色质的基本结构单元，H2A.Z和H3.3是体内常见的组蛋白变体。通过影响反式调控因子与顺式序列元件的结合以及改变染色质结构开放或压缩状态，核小体组装调控着几乎所有DNA层次上的生物学过程。体内核小体组装、解聚及滑动等高度动态变化的过程受到诸多因素的综合影响。利用体外实验系统研究组蛋白及其变体H2A.Z和H3.3装配核小体过程的动力学特征，是阐明组蛋白及其变体装配核小体规律较为有效的手段。本项目利用体外组装核小体的方法，建立了体外组装核小体的动力学理论模型，证明了核小体组装与解聚过程遵循化学反应动力学规律，阐明了组装效率与组装物浓度、组装时间以及反应速率常数之间的关系；证明了序列模体和TA 10.5-bp周期性规律对常规核小体、含组蛋白变体H2A.Z核小体和含组蛋白变体H3.3核小体装配的影响；从解聚的角度体外研究了核小体动力学特征，表明解聚过程分为三个阶段；分析了组蛋白变体与常规组蛋白装配核小体的竞争动力学关系；基于DNA弯曲能等物理原理，发展了一个核小体定位的理论计算模型，开发了一个网络在线版核小体预测的工具。项目的结果能够为体内核小体装配及定位、染色质动力学、组蛋白变体动力学及功能等相关问题的研究奠定一定的基础。