Chromatin-mediated mechanisms of transcription regulation in ES cells

项目摘要/摘要 我们的研究兴趣集中在不同物种染色质结构的异同上 细胞类型以及调节这些差异的染色质重塑因子如何调节细胞命运。长的- 我们实验室的学期目标是全面了解功能、目标、调节和 具有关键功能的非编码RNA和染色质调节因子的作用机制 基因调控网络。 大约75%的哺乳动物基因组是转录的。而编码区只占 ~2%的基因组，其余的基因间和基因内转录产生大量的非 编码RNA(NcRNAs)。两个富含非编码转录的区域是活性增强子(其中ncRNAs 是双向产生的)和启动子(其中ncRNA在反义方向上产生自 蛋白质编码基因)。虽然一些单独的ncRNA已被证明在基因激活中发挥作用，但它们的 基因调控方面的活动还没有得到系统的解决。最近，我们展示了esBAF，一种 结合增强子和启动子的核小体重塑复合体是抑制 胚胎干细胞基因组中大量的ncRNAs。当esBAF耗尽时，核小体占据 开放染色质的侧翼区域减少，导致ncRNA转录增加。基于这些 研究表明，我们处于一个独特的位置来构建核小体重塑因子对ncRNA的调控网络 在ES细胞中。具体地说，我们将确定(1)esBAF如何调节高阶染色质结构，(2) 增强子特异的ncRNAs在增强子环和基因调控中的作用，(3)机制 NcRNAs对mRNAs的潜在调控，以及(4)核小体重塑复合体网络 调控ncRNA的表达。这些研究将加深我们对以染色质为中心的 对ncRNA的调控。 在接下来的五年里，我们的实验室将确定调控ncrna的核小体重构体。 表达，探讨核小体重塑因子在调控高阶染色质结构中的作用 增强子-启动子环的水平，并决定两类未鉴定的ncRNAs的功能 在ES细胞中。这项拟议的研究具有重要意义，因为它将揭示 编排染色质动力学与ncRNA功能的相互作用，从而为 了解胚胎干细胞的命运决定。

PROJECT SUMMARY/ABSTRACT Our research interests focus on the similarities and differences in chromatin structure among different cell types and how chromatin remodeling factors that modulate these differences regulate cell fate. The long- term goals of our laboratory are to comprehensively understand the functions, targets, regulation, and mechanisms of action of non-coding RNAs (ncRNAs) and chromatin regulatory factors with critical functions in gene regulatory networks. Approximately 75% of the mammalian genome is transcribed. While coding regions account for only ~2% of the genome, the remaining intergenic and intragenic transcription generates a large collection of non- coding RNAs (ncRNAs). Two regions rich with non-coding transcription are active enhancers (where ncRNAs are generated bi-directionally) and promoters (where ncRNAs are generated in the antisense orientation from protein-coding genes). While a few individual ncRNAs have been shown to function in gene activation, their activities in gene regulation have not been systematically addressed. Recently, we showed that esBAF, a nucleosome remodeling complex that binds enhancers and promoters, is necessary for repression of numerous ncRNAs throughout the genome in ES cells. Upon depletion of esBAF, nucleosome occupancy flanking regions of open chromatin was reduced, resulting in elevated ncRNA transcription. Based on these studies, we are in a unique position to build a network of ncRNA regulation by nucleosome remodeling factors in ES cells. Specifically, we will determine (1) how esBAF regulates higher order chromatin structure, (2) the functions of enhancer-specific ncRNAs (eRNAs) in enhancer looping and gene regulation, (3) the mechanisms underlying regulation of mRNAs by ncRNAs, and (4) the network of nucleosome remodeling complexes regulating ncRNA expression. These studies will enhance our understanding of the chromatin-centered regulation of ncRNAs. Over the next five years, our laboratory will identify nucleosome remodelers that regulate ncRNA expression, explore the role of nucleosome remodeling factors in regulating higher order chromatin structure at the level of enhancer-promoter looping, and determine the function of two uncharacterized classes of ncRNAs in ES cells. The proposed research is significant because it will uncover fundamental mechanisms that choreograph the interplay of chromatin dynamics with ncRNA function, consequently providing a crucial step in understanding ES cell fate decisions.