EPIGENOMIC REGULATION OF GENOMES

基因组的表观基因组调控

• 批准号: 10797418
• 负责人: B FRANKLIN PUGH
• 金额: $ 9.46万
• 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/10797418
• 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 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.

项目摘要 基因调控是所有生命的核心，无论是正常的还是患病的。这项研究计划的长期目标是 了解酵母和人类系统中所有基因调控的分子机制。这 基础知识将有助于为人们提供更好的诊断和治疗选择。对胡适的监管-- 人类基因组是非常复杂的。因此，这项研究计划最初集中在更简单的酵母Saccha- 从实验上剖析了基因调控的机制，这些机制是所有欧盟的基本和共同的。 核生物生命。在酵母中开发的概念最终在人类细胞中进行测试，从而加速了发现。 这项研究计划开发了一种名为CHIP-EXO的超高分辨率分析来绘制边界 在碱基对分辨率下，基本上任何蛋白质在任何基因组中的位置。使用此策略，COM- 首次建立了酵母细胞蛋白质-DNA结构的表观基因组图谱。这 现已在人类细胞中建立起来。表观基因组在这里被定义为所有分子在- 与DNA和RNA的相互作用，超越了碱基配对。这项研究的下一个阶段是了解 表观基因组中蛋白质组分之间的相互作用。这将在一定程度上通过Dele- 表观基因组的蛋白质组分的缺失、突变和/或快速耗尽，特别是与 可诱导的和构成的转录。前者是基因特异性的，并且可以在对 特定的信令事件。后者对大多数基因来说是普遍的，通常发生在低水平。重要的是， 这里的研究计划是定义蛋白质结构，指定诱导性启动子和构成启动子- 艾尔斯。一旦这种结构的蛋白质组分被实验移除(例如，序列特异性的反式- 划痕因子或其辅助因子)，则将测量这种去除对染色质组织的影响。 启动、加载核心转录机器以及随后的转录。一个平行的战略将是EM- 应用于人类组织培养细胞以评估保守的范例。 这项研究还将继续其先前的染色质组织的生化重组 在整个基因组中使用纯化的蛋白质，但现在添加了转录机器的组件 以及他们的监管因素。生化系统将对实验参数提供更好的控制- 从而对基因控制的分子机制提供了更多的洞察力。现在很明显，在- 诱导基因在细胞核内的3D空间中聚合。然而，目前还不清楚哪些基因结合在一起。 哪个中心。因此，将采用像Sprite这样的3D作图技术来测量基因聚集性。这 将提供对多个基因如何由调控信号协调诱导的洞察。带到- 这项研究计划的成果将是对转录和其基因的详细分子理解 监管。