PRC1在胚胎干细胞生长和分化中识别和抑制靶基因的分子机制

多梳抑制复合体（PRC1或2）通过重塑染色质构象在表观遗传上抑制靶基因表达，从而定义和维持机体内多种细胞类型的特异表征和功能。CBX蛋白是参与PRC1识别靶基因的核心成员。我们对Cbx基因敲除或稳定表达外源Cbx基因及突变体的胚胎干细胞（ESC）株系的鉴定发现：CBX与非编码RNA的结合存在未知的复杂性和普遍性；Cbx同源基因及转录剪切体功能有所差异。我们推测CBX的多样性及其与非编码RNA的结合直接参与调节PRC1对靶基因的识别和抑制。本课题以小鼠ESC及其分化体系为模型，分离鉴定与PRC1特异性结合的非编码RNA分子，澄清作用的结构基础及非编码RNA调节PRC1靶向募集的分子机理；在表达不同CBX的遗传背景下剖析CBX的多样形式与PRC1特异性识别靶基因的相互关联。阐明PRC1随细胞分化动态识别靶基因的分子机理对揭示其如何在多种细胞类型和生理、病理状态下发挥特异性调控作用至关重要。

Polycomb Repressive Complexes 1 and 2 (PRC1, PRC2) are epigenetic regulatory complexes that cause transcriptional repression of target genes through modifying chromatin. The PRC1/2-mediated gene silencing mechanism is required for the maintenance of the core properties of embryonic stems cells (ESC), pluripotency and self renewal. The spectra of target genes for PRC complexes are dynamically changing along with cell differentiation during developmental processes. One of the key questions in the field is how PRCs selectively recognize and switch targets in multiple cell types and under various physiological or pathological conditions. To address this question, our research has been focused on CBX protein(s), one of the core components in PRC1 that is responsible for the chromatin targeting of the complex. We generated and analyzed a number of ES cell lines with the expression of Cbx genes manipulated. Our preliminary data have provided us several lines of valuable information. 1) The interaction between CBX7 and ncRNA is structurally complicated and possibly prevalent in cells; 2) Cbx homologs and their transcription splicing variants expressed in ESC may have distinct functions. Combining previously published data and our new evidence, we propose that both the diversity of CBX proteins and their interaction with ncRNA contribute to the targeting specificity of PRC1. In this study, we will employ approaches in biochemistry, molecular and cellular biology, together with genome-wide analyses to identify and verify the ncRNA species that specifically interact with PRC1. We will further elucidate the structural foundation of these interactions and their role in selectively targeting PRC1 to chromatin. In addition, we plan to characterize main features of PRC1 and analyze its function in maintaining ESC pluripotency and regulating differentiation in established or newly generated ESC lines with manipulated Cbx expression. The data will help us make functional comparisons among Cbx homologs and their transcription splicing variants. Overall, our study will help elucidate the targeting and working mechanisms of PRC1.