Nuclear Organization and Function

核组织和功能

• 批准号: 10334480
• 负责人: Victor G. Corces
• 金额: $ 52.91万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-01 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10334480
• 关键词: 3-Dimensional; Affect; Biological Models; Cell Differentiation process; Cell Nucleus; Cells; Cellular biology; Chromatin; Chromatin Fiber; Complex; Computational Biology; Data; Dimensions; Disease; Drosophila genus; Enhancers; Environment; Epigenetic Process; Eukaryota; Frequencies; Gene Expression; Gene Expression Regulation; Genetic; Genetic Diseases; Genetic Materials; Genetic Transcription; Genome; Hi-C; Homeostasis; Human Genetics; Knowledge; Logic; Machine Learning; Malignant Neoplasms; Mammals; Nuclear; Pancreas; Proteins; Resolution; Role; Site; Somatic Cell; Testing; Three-dimensional analysis; Tissues; Transcription Process; Transcriptional Regulation; Vertebrates; Work; base; cohesin; computerized tools; epigenomics; experimental study; human disease; human embryonic stem cell; interdisciplinary approach; mammalian genome; novel; promoter; protein complex; scaffold; stem cell differentiation; stem cell therapy

PROJECT SUMMARY. Proper regulation of gene expression is essential for cell differentiation and homeostasis. Most of our understanding of the mechanisms that control the transcription process comes from studies of the one-dimensional genome i.e. the 10 nm chromatin fiber. However, the genome is folded in the three-dimensional (3D) nuclear space, and the relationship between this organization and gene expression is poorly understood. Using Drosophila as a model system, where it is feasible to obtain 250 bp resolution Hi-C data, we have found that the genome is folded into only one type of domain, which we call compartmental domains. These domains precisely correlate with the transcriptional state of their sequences. Compartmental domains are also found in other lower eukaryotes. Based on this, we propose that compartmental domains represent an evolutionarily conserved principle of genome 3D organization. Drosophila and lower eukaryotes either lack CTCF or this protein is unable to stop cohesin extrusion. However, CTCF can interfere with the progression of cohesin extrusion in vertebrates, which in turn affects other types of interactions in the genome. Here we suggest extending concepts learned from the analysis of 3D organization in Drosophila to mammals by proposing an ambitious and substantive multi-disciplinary approach combining genetics, epigenomics, computational biology, and differentiation of human embryonic stem cells (hESC) into disease-relevant tissues. The hypothesis underlying the proposed experiments is based on the idea that, rather than the prevalent view of large compartments containing smaller TADs, the mammalian genome is organized by conserved principles into relatively small compartmental domains. Cohesin extrusion operates on top of the compartmental domain scaffold and affects its organization. To test this novel hypothesis, we will deplete specific proteins present in complexes required for various aspects of the transcription process. We will also deplete protein complexes responsible for H3K27me3- and H3K9me3-dependent silencing. We will then use Micro-C XL to obtain very high-resolution interaction data and examine effects of protein depletion on the formation of self-interacting domains and in the interactions between these domains. These effects will be examined in the presence and absence of cohesin in order to understand the contribution of loop extrusion to enhancer-promoter interaction frequency. We will examine the predictability of 3D genome organization from one-dimensional epigenetic information using machine learning computational tools. We will study the logic of CTCF loop formation by analyzing the local chromatin environment around CTCF sites able or unable to form loops of different strengths using a new computational tool we have developed. Principles learned from these experiments will be tested by analyzing changes in 3D organization and their relationship to gene expression during the differentiation of hESCs into pancreatic cells. Results from this work will fill critical gaps in our understanding of the relationship between 3D chromatin organization and transcription, and its possible role in human disease.

项目摘要。适当调节基因表达对于细胞分化至关重要 稳态。我们对控制转录过程机制的大多数理解都来自 一维基因组的研究，即10 nm染色质纤维。但是，基因组折叠在 三维（3D）核空间以及该组织和基因表达之间的关系是 理解不佳。使用果蝇作为模型系统，在该系统中获得250 bp分辨率HI-C是可行的 数据，我们发现基因组仅折叠成一种类型的域，我们称之为隔室 域。这些域与序列的转录状态完全相关。隔间 在其他较低的真核生物中也发现了域。基于此，我们提出了隔区域 代表基因组3D组织的进化保守原则。果蝇和较低的真核生物 缺乏CTCF，或者该蛋白无法阻止凝聚素挤出。但是，CTCF可以干扰 脊椎动物中粘蛋白挤出的进展，进而影响基因组中的其他类型的相互作用。 在这里，我们建议扩展从果蝇3D组织分析到哺乳动物的概念 通过提出一种雄心勃勃的实质性多学科方法，结合了遗传学，表观基因组学， 计算生物学以及人类胚胎干细胞（HESC）与疾病相关组织的分化。 提出的实验的基础假设是基于这样的观念，即不是普遍的观点 在包含较小TAD的大室中，哺乳动物基因组由保守的原理组织 进入相对较小的隔室域。粘蛋白挤出在隔室结构域的顶部 脚手架并影响其组织。为了检验这个新的假设，我们将耗尽存在的特定蛋白质 转录过程的各个方面所需的复合物。我们还将耗尽蛋白质复合物 负责H3K27ME3-和H3K9ME3依赖性沉默。然后，我们将使用Micro-C XL获得非常 高分辨率相互作用数据并检查蛋白质耗竭对形成自我相互作用的影响 域以及这些域之间的相互作用。这些效果将在存在的情况下进行检查， 缺乏粘着蛋白以了解环路挤压对增强子促销相互作用的贡献 频率。我们将检查一维表观遗传学的3D基因组组织的可预测性 使用机器学习计算工具的信息。我们将通过研究CTCF循环形成的逻辑 分析CTCF站点周围的局部染色质环境能够或无法形成不同的循环 使用我们开发的新计算工具的优势。从这些实验中学到的原则将 通过分析3D组织的变化及其与基因表达的关系来测试 将hESC分化为胰腺细胞。这项工作的结果将填补我们对 3D染色质组织与转录之间的关系及其在人类疾病中的可能作用。