cis-Acting Elements Regulating Developmental Control of Replication Timing

项目总结/摘要 DNA复制对基因组的结构和功能完整性至关重要，并与大分子生物学密切相关。 染色体的三维结构和细胞谱系的特化，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）， 复制时间（RT）、染色体结构和鼠胚胎干细胞（mESC）中的转录。 ERCE具有乙酰化组蛋白，形成CTCF/粘着蛋白非依赖性3D相互作用，并被CTCF/粘着蛋白共同占据。 多能性转录因子Oct 4、Sox 2和Nanog（OSN）。我们不知道的是这些特性 控制染色体的结构和功能，以及它们的活动是否是可分离的。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。我们眼前的 目的是确定ERCE共同调节RT、染色质结构和转录的机制。我们 核心假设是ERCE相互作用产生组蛋白乙酰化的3D枢纽，从而招募复制 起始因子，同时独立调节转录。我们的基本原理是， 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，并最终在疾病状态下的错误调节。AIM 1基因 剖析ERCE，以确定其相关活动所必需和充分的最小序列。AIM2 检验了Rif 1存在于晚期复制的染色质中，但在RT全基因组范围内是必需的， 将组蛋白乙酰化聚焦于ERCE，以通过其相互作用招募复制起始蛋白Treslin 组蛋白乙酰化结合蛋白Brd 2和Brd 4。Aim 3将解决RT的长期关系 到转录。我们认为细胞类型特异性转录因子创造了组蛋白乙酰化的枢纽 独立于他们在RT和架构中的角色。这一贡献将是重大的，因为它将解除一个主要的 研究染色体结构和功能调节机制及其如何调节的障碍 在细胞命运转变过程中，以及最终，它们如何在人类疾病中被错误调节。这项工作是 创新，因为ERCE的突破性发现引入了新的假设，概念和 基因组结构领域的方法。

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.