Organizational principles and functional role of 3D enhancer hubs in cell fate decisions

3D增强子中枢在细胞命运决定中的组织原则和功能作用

• 批准号: 10239060
• 负责人: Effie Apostolou
• 金额: $ 44.04万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-15 至 2024-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10239060
• 关键词: 3-Dimensional; Architecture; Binding; Binding Sites; Biological Assay; CRISPR interference; Cancer Biology; Cell Fate Control; Cell Line; Cell Nucleus; Cell physiology; Cells; Chemicals; Chromatin; Chromatin Loop; Code; Communication; Complex; Development; Disease; Down-Regulation; Effectiveness; Elements; Embryo; Embryonic Development; Endoderm; Engineering; Enhancers; Ensure; Epiblast; Epigenetic Process; Family; Foundations; Gene Expression; Genes; Genetic; Genetic Enhancer Element; Genetic Transcription; Genome; Genomics; Goals; Homeostasis; Human; In Vitro; Knowledge; Link; Maintenance; Molecular; Mus; Mutation; Nuclear; Outcome; Play; Proteins; Proteomics; Regenerative Medicine; Regulation; Regulatory Element; Reporter; Resolution; Role; Somatic Cell; Specificity; Stress; Structure; Technology; Testing; Transcription Coactivator; Transcriptional Regulation; Untranslated RNA; Validation; Work; base; blastocyst; blastomere structure; cell type; cofactor; connectome; embryo tissue; embryonic stem cell; experimental study; grasp; in vivo; insight; novel; pluripotency; programs; promoter; spatiotemporal; stem cells; stemness; transcription factor

Project Summary Embryonic development and tissue homeostasis rely on the tight regulation of cell-type specific transcriptional programs by particular transcription factors (TFs) and their target enhancers. The emergence of three- dimensional (3D) genome organization as an important layer of transcriptional control, stresses the fact that understanding how enhancers communicate with target genes to coordinate transcriptional activity requires knowledge of the 3D nuclear topology. In a recent study, we captured a drastic rewiring of three-dimensional regulatory contacts between somatic cells and embryonic stem cells (ESCs) by H3K27ac HiChIP and identified complex 3D “enhancer hubs”, where enhancers are spatially clustered with multiple highly-expressed genes with known or predicted functions in regulation of stemness. Genetic or epigenetic modulation of such enhancers in ESCs resulted in downregulation of all hub-connected genes and partial differentiation, which supports a vital role for these architectural nodes in gene coregulation and cell identity. In addition, we provided proteomics and genetic evidence that KLF family TFs play an important role in the organization and regulation of 3D enhancer hubs in ESCs and identified candidate cofactors. Based on these results, we hypothesize that 3D enhancer hubs function as architectural “headquarters” of cell identity, where cell type-specific genes are sequestered by specific transcriptional regulators to facilitate coordinated gene expression. Here, we will test this hypothesis both in the contexts of mouse ESCs, and of early developmental cell fate decisions using in vitro and in vivo approaches. Specifically, we aim to (1) target systematically enhancers and genes within hubs to determine the functional consequences on the pluripotent transcriptional network and the stability of ESC identity, (2) determine the critical protein factors and activities that control enhancer hub formation and functionality and (3) identify and characterize 3D enhancer hubs that are critical for acquisition and maintenance of each of the early developmental fates. Successful completion of our aims will offer mechanistic insights into the organization and regulation of 3D enhancer hubs, determine their role in cell fate control and reveal novel ways for engineering cell identity by targeting critical architectural nodes and factors.

项目摘要 胚胎发育和组织稳态依赖于细胞类型特异性转录的严格调节 特定转录因子（TFS）及其目标增强器的程序。三个出现 维度（3D）基因组组织是转录控制的重要层，强调了以下事实 了解增强剂如何与目标基因进行交流以协调转录活动需要 3D核拓扑的知识。在最近的一项研究中，我们捕获了三维的急剧重新布线 通过H3K27AC Hichip鉴定的体细胞和胚胎干细胞之间的调节接触（ESC） 复杂的3D“增强子枢纽”，其中增强子用多个高度表达的基因在空间上聚集 具有已知或预测的茎调节功能。此类遗传或表观遗传调节 ESC中的增强子导致所有与HUB连接的基因和部分分化的下调，这 支持这些架构节点在基因结构和细胞身份中的重要作用。另外，我们 提供了蛋白质组学和遗传学证据，表明KLF家族TFS在组织中起重要作用，并且 ESC中的3D增强子中心的调节和确定的候选辅助因子。基于这些结果，我们 假设3D增强子中心充当细胞身份的建筑“总部”，其中细胞 特定类型的基因被特定的转录调节剂隔离，以促进协调的 基因表达。在这里，我们将在小鼠ESC和早期的背景下检验这一假设 使用体外和体内方法的发育细胞脂肪决策。具体而言，我们的目标是（1）目标 集线器中的系统增强子和基因，以确定对多能的功能后果 转录网络和ESC身份的稳定性，（2）确定关键蛋白质因子和活动 控制增强器集线器的形成和功能，（3）识别和表征3D增强子集线器 对于获得和维持每个早期发展命运至关重要。成功完成 我们的目标将提供有关3D增强器中心组织和调节的机械见解，确定 它们在细胞命运控制中的作用，并通过靶向关键建筑，揭示了工程细胞身份的新颖方式 节点和因素。