EPIGENOMIC REGULATION OF GENOMES

基因组的表观基因组调控

• 批准号: 10685469
• 负责人: B FRANKLIN PUGH
• 金额: $ 53.86万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10685469
• 关键词: 3-Dimensional; Acceleration; Adopted; Architecture; Base Pairing; Binding; Biochemical; Biological Assay; Cell Nucleus; Cells; Chromatin; Complex; DNA; Diagnostic; Disease; Event; Excision; Gene Cluster; Gene Expression Regulation; Genes; Genetic Transcription; Genome; Goals; Human; Human Genome; Knowledge; Life; Location; Maps; Measures; Molecular; Mutation; Persons; Phase; Proteins; RNA; Regulation; Research; Resolution; Saccharomyces; Signal Transduction; Specific qualifier value; System; Technology; Testing; Yeasts; cofactor; epigenome; epigenomics; human tissue; insight; programs; promoter; protein function; protein purification; reconstitution; response; tissue/cell culture; transcription factor; ultra high resolution; whole genome

Project Summary/Abstract Gene regulation is central to all life, normal and diseased. The long-term goal of this research program is to un- derstand the molecular mechanisms governing the regulation of all genes in yeast and human systems. This basic knowledge will help produce better diagnostics and treatment options for people. Regulation of the hu- man genome is very complex. Therefore, this research program is focused initially on the simpler yeast Saccha- romyces to experimentally dissect mechanisms of gene regulation that are fundamental and common to all eu- karyotic life. Concepts developed in yeast are ultimately tested in human cells, thereby accelerating discovery. This research program has developed an ultra-high-resolution assay called ChIP-exo to map the bound locations of essentially any protein throughout any genome at base-pair resolution. Using this strategy, a com- prehensive first-of-its-kind epigenome map of the protein-DNA architecture of yeast cells was established. This is now being established in human cells. The epigenome is defined here as the compilation of all molecular in- teractions with DNA and RNA, beyond base-pairing. The next phase of this research is to understand the func- tional interactions among the protein components of the epigenome. This will be achieved in part through dele- tion, mutation, and/or rapid depletion of protein components of the epigenome, particularly those involved in inducible and constitutive transcription. The former is gene-specific and can be hyper-expressed in response to specific signaling events. The latter is general to most genes and typically occurs at low levels. Importantly, the research program here is defining the protein architecture that specifies inducible versus constitutive promot- ers. Once protein components of this architecture are experimentally removed (e.g., sequence-specific tran- scription factors or their cofactors), then the impact of this removal will be measured on chromatin organiza- tion, loading of the core transcription machinery and subsequent transcription. A parallel strategy will be em- ployed in human tissue culture cells to assess conserved paradigms. This research will also continue with its previous biochemical reconstitution of chromatin organization across entire genomes using purified proteins, but now adding in components of the transcription machinery and their regulatory factors. A biochemical system will provide greater control over the experimental parame- ters and therefore provide greater insight into molecular mechanisms of gene control. It is now clear that in- duced genes coalesce in 3D space within the nucleus. However, it remains unclear which genes coalesce into which hubs. Therefore, 3D mapping technologies like SPRITE will be adopted to measure gene clustering. This will provide insight into how multiple genes become coordinately induced by regulatory signals. Taken to- gether, the product of this research program will be a detailed molecular understanding of transcription and its regulation.

项目概要/摘要 基因调控对于所有生命（无论是正常生命还是患病生命）至关重要。该研究计划的长期目标是 了解控制酵母和人体系统中所有基因调节的分子机制。这 基础知识将有助于为人们提供更好的诊断和治疗选择。胡的调节 人类基因组非常复杂。因此，该研究计划最初侧重于更简单的酵母 Saccha- romyces 通过实验剖析对所有欧盟国家来说都是基本且共同的基因调控机制 核生命。在酵母中开发的概念最终在人体细胞中进行测试，从而加速发现。 该研究项目开发了一种称为 ChIP-exo 的超高分辨率测定法，用于绘制结合图谱 基本上任何蛋白质在任何基因组中的碱基对分辨率的位置。使用这种策略，一个公司 建立了酵母细胞蛋白质-DNA 结构的全面的首个表观基因组图谱。这 现在正在人体细胞中建立。表观基因组在这里定义为所有分子信息的汇编 与 DNA 和 RNA 的相互作用，超越碱基配对。这项研究的下一阶段是了解功能 表观基因组蛋白质成分之间的相互作用。这将部分通过删除来实现 表观基因组蛋白质成分的变化、突变和/或快速耗尽，特别是那些涉及 诱导型转录和组成型转录。前者是基因特异性的，并且可以响应于过度表达 特定的信号事件。后者对于大多数基因来说是普遍的，并且通常发生在低水平。重要的是， 这里的研究计划正在定义蛋白质结构，指定诱导型与组成型促进剂 呃。一旦通过实验去除该结构的蛋白质成分（例如，序列特异性转录 转录因子或其辅助因子），然后将测量这种去除对染色质组织的影响 化，核心转录机制的加载和随后的转录。并行策略将是 用于人体组织培养细胞以评估保守范式。 这项研究还将继续其之前的染色质组织生化重建 使用纯化的蛋白质跨整个基因组，但现在添加转录机器的组件 及其调节因素。生化系统将更好地控制实验参数 因此，可以更深入地了解基因控制的分子机制。现在很清楚，在—— 诱导基因在细胞核内的 3D 空间中合并。然而，目前尚不清楚哪些基因融合成 哪些枢纽。因此，将采用SPRITE等3D绘图技术来测量基因聚类。这 将深入了解多个基因如何被调控信号协调诱导。被带到- 总之，该研究计划的产品将是对转录及其转录的详细分子理解 规定。