Mechanism of chromatin accessibility, 3D chromosome organization, and their functions in gene regulation

染色质可及性机制、3D 染色体组织及其在基因调控中的功能

基本信息

批准号：
10887047
负责人：
Lu Bai
金额：
$ 1.77万
依托单位：
PENNSYLVANIA STATE UNIVERSITY, THE
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-01-01 至 2024-12-31
项目状态：
已结题

项目摘要

Project Summary/Abstract The overall theme of my research program is to understand the formation mechanism of chromatin structure and its role in gene regulation. We plan to address this problem at two different levels: 1) at the chromatin / nucleosome level, we will identify pioneer factors (PFs) and investigate the mechanism underlying nucleosome displacement and chromatin opening, and 2) at the chromosome level, we will study the mechanism of gene regulation by high-order 3D chromosome organization. In the past five years, supported by two NIGMS R01 grants, we have made significant progress in both directions. New observations and insights we gained from these studies, as well as several novel methods we developed, form the foundation of this proposal. Theme1: Pioneer factors (PFs) can invade nucleosome and increase chromatin accessibility near their binding sites and therefore play critical roles in gene regulation. Mis-regulation of PFs is highly linked to cancer and developmental disease. Despite their essential functions, only a handful of PFs have been identified and the mechanism underlying the pioneer activity is unclear. The long term goal of this theme is to systematically characterize PFs and their pioneer activities in health and disease cells, and to develop a full understanding of the mechanism of these activities. In our recent PF studies, we have innovated an Integrated Synthetic Oligo (ISO) assay to investigate PF function in a high-throughput manner. In the next five years, we plan to 1) adapt the ISO assay into human cell lines and pluripotent stem cells to dissect the genetic rules underlying PF binding and nucleosome displacement, and 2) further our understanding of the pioneering activity by investigating the co-factors of PFs, the sequence of events during nucleosome displacement, and the kinetic rates of these events. Theme2: Imaging and 3C-based methods have revealed 3D chromosome organization with extensive long-distance chromosomal interactions. The long-term goal of this theme is to understand the formation mechanism of these high-order chromosome structures and their roles in gene regulation. Our recent studies show that long-distance chromosomal interactions contribute to gene regulation in yeast. More specifically, some allelic or co-regulated genes cluster in the 3D space, and such clustering is correlated with higher gene activities. The former case is analogous to the “transvection” phenomenon in Drosophila. These novel observations revealed a new layer of gene regulation in yeast and opened the opportunity of using the powerful genetic system to investigate the 3D genome function. Currently we have little understanding of what mediates the cluster formation and how the clusters enhance gene expression. In the next five years, we plan to 1) use an unbiased genetic screen to identify factors that negatively regulate transvection, and investigate the underlying mechanism, and 2) test the hypothesis that some activators condense though liquid-liquid phase separation, leading to the clustering of co-regulated genes and enhanced expression.

项目概要/摘要我的研究计划的总体主题是了解染色质结构的形成机制及其在基因调控中的作用我们计划在两个不同的层面上解决这个问题：1）在染色质/核小体水平，我们将识别先锋因子（PF）并研究其潜在机制核小体位移和染色质打开，2）在染色体水平上，我们将研究高阶3D染色体组织的基因调控机制在过去的五年中，在两项 NIGMS R01 资助的支持下，我们在两个方向上都取得了重大进展。我们从这些研究中获得的观察和见解，以及我们开发的几种新方法，构成本提案的基础。主题1：先锋因子（PF）可以侵入核小体并增加其结合附近的染色质可及性因此，PF 的错误调控与癌症密切相关。尽管它们具有重要的功能，但只有少数 PF 已被识别。该主题的长期目标是：系统地表征 PF 及其在健康和疾病细胞中的先驱活动，并开发完整的在我们最近的 PF 研究中，我们创新了一种方法来了解这些活动的机制。集成合成寡核苷酸 (ISO) 测定以高通量方式研究 PF 功能在接下来的五年中。年，我们计划 1) 将 ISO 测定应用于人类细胞系和多能干细胞，以剖析遗传 PF 结合和核小体置换的基本规则，2）进一步加深了我们对开创性的理解通过研究 PF 的辅因子、核小体位移过程中的事件序列以及这些事件的动力学速率。主题2：成像和基于 3C 的方法揭示了广泛的 3D 染色体组织该主题的长期目标是了解长距离染色体相互作用。这些高阶染色体结构的机制及其在基因调控中的作用。表明长距离染色体相互作用有助于酵母中的基因调控。一些等位基因或共同调控的基因在3D空间中聚集，并且这种聚集与更高的基因相关前一种情况类似于果蝇中的“平移”现象。观察结果揭示了酵母中新的基因调控层，并为使用目前我们对研究 3D 基因组功能的强大遗传系统知之甚少。介导簇的形成以及簇如何增强基因表达在未来五年中，我们计划。 1）使用公正的遗传筛选来识别负向调节横传的因素，并调查潜在的机制，2) 检验一些活化剂通过液-液凝结的假设相分离，导致共同调节的基因聚集并增强表达。