cis-Acting Elements Regulating Developmental Control of Replication Timing

项目摘要/摘要 DNA复制是基因组结构和功能完整性的核心，并与大型- 标尺3D染色体组织和细胞谱系规范，但我们对其调控知之甚少。 我们已经确定了特定的顺式元件，称为早期复制控制元件(ERCEs)，它调节 小鼠胚胎干细胞的复制时间(RT)、染色体结构和转录。 ERCEs含有乙酰化组蛋白，形成CTCF/粘附素不依赖的3D相互作用，并与 多能转录因子Oct4、Sox2和Nanog(OSN)。目前尚不清楚的是这些特性是如何 控制染色体的结构和功能，以及它们的活动是否可分离。我们的长期目标是 了解RT与染色体结构、表观遗传状态和疾病的关系。我们的直接客户 目的是确定ERCEs共同调节RT、染色质结构和转录的机制。我们的 中心假说是，ERCEs相互作用，创建组蛋白乙酰化的3D中枢，从而招募复制 启动因子，同时独立地调节转录。我们的理论基础是通过以下方式阐明机制 哪些ERCEs共同调节RT、转录和基因组结构将为研究 染色体的结构和功能，以及最终在疾病状态下的错误调节。Aim1将从基因上 解剖ERCEs以确定其相关活动所必需和充分的最小序列。AIM2 Rif1位于后期复制的染色质中，但在整个基因组范围内是必需的， 将组蛋白乙酰化与ERCEs结合，通过相互作用招募复制起始蛋白Treslin 与组蛋白乙酰化结合蛋白Brd2和Brd4结合。Aim3将解决RT的长期关系 转到抄写。我们认为，细胞类型特异的转录因子产生组蛋白乙酰化中心 独立于他们在RT和体系结构中的角色。这一贡献将是重大的，因为它将提升一个重大的 研究染色体结构和功能的调控机制及其调控方式的障碍 在细胞命运转变期间，以及最终它们在人类疾病中是如何被错误调控的。这项工作是 创新是因为ERCEs的突破性发现引入了新的假设、概念和 基因组体系结构领域的方法。

PROJECT SUMMARY / ABSTRACT DNA replication is central to the structural and functional integrity of the genome and intimately tied to large- scale 3D chromosome organization and cell lineage specification, but we understand little about its regulation. We have identified specific cis-elements, termed Early Replication Control Element (ERCEs), that regulate replication timing (RT), chromosome architecture, and transcription in murine embryonic stem cell (mESCs). ERCEs harbor acetylated histones, form CTCF/cohesin-independent 3D interactions and are co-occupied by pluripotency transcription factors Oct4, Sox2 and Nanog (OSN). What is not known is how these properties control chromosome structure and function and whether their activities are separable. Our long-term goal is to understand the relationship of RT to chromosome architecture, epigenetic states and disease. Our immediate goal is to identify mechanisms by which ERCEs co-regulate RT, chromatin architecture and transcription. Our central hypothesis is that ERCEs interact to create 3D hubs of histone acetylation that recruit replication initiation factors while independently regulating transcription. Our rationale is that elucidating mechanisms by which ERCEs co-regulate RT, transcription and genome architecture will open new horizons for studies of chromosome structure and function and, ultimately, its mis-regulation in disease states. Aim1 will genetically dissect ERCEs to identify minimal sequences necessary and sufficient for their associated activities. Aim2 tests the hypothesis that Rif1, which resides in late replicating chromatin but is necessary RT genome-wide, focuses histone acetylation to ERCEs to recruit the replication initiation protein Treslin through its interaction with histone acetylation binding proteins Brd2 and Brd4. Aim3 will address the longstanding relationship of RT to transcription. We propose that cell type specific transcription factors create hubs of histone acetylation independent of their roles in RT and architecture. This contribution will be significant because it will lift a major barrier to the study of mechanisms regulating chromosome structure and function, how they are regulated during cell fate transitions and, ultimately, how they are mis-regulated in human disease. This work is innovative because the breakthrough discovery of ERCEs introduces novel hypotheses, concepts and approaches to the genome-architecture field.