Deciphering the mechanism of long-range gene regulation in vivo

破译体内长程基因调控机制

• 批准号: 10473041
• 负责人: Evgeny Kvon
• 金额: $ 141.3万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-09 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10473041
• 关键词: 3-Dimensional; Address; Affect; Base Pairing; Binding Proteins; Chromatin; Communication; DNA; Development; Disease; Distant; Elements; Enhancers; Foundations; Gene Expression; Gene Expression Profile; Gene Expression Regulation; Gene Transfer Techniques; Genes; Genetic Transcription; Genome; Genomics; Human Genome; Imaging technology; Knowledge; Link; Maps; Mediating; Methods; Mus; Mutation; Nucleotides; Pathologic; Regulation; Regulatory Element; Reporter; Resolution; Role; Technology; Time; Untranslated RNA; Variant; Work; gain of function; genetic regulatory protein; genome editing; human disease; in vivo; interdisciplinary approach; novel; promoter; remote control; transcription factor

PROJECT SUMMARY Distant-acting (“remote”) transcriptional enhancers bound by transcription factors (TFs) drive gene expression patterns in space and time. The importance of this function is underscored by mounting evidence linking mutations affecting remote enhancers to human disease. What remains unclear is how these remote enhancers regulate their target genes over long genomic distances, often skipping intervening genes. Remote enhancers typically interact with target promoters with the support of higher-order chromatin organization, but disruption of this chromatin organization does not abolish enhancer–promoter interactions. What additional factors regulate these long-range regulatory interactions remains largely unexplored. To address this central question, the proposed project will use a novel gain-of-function approach to identify cis-regulatory sequences that mediate long-range enhancer–promoter communication in the context of mammalian development. Traditional methods that assess in vivo enhancer activity rely on enhancer-reporter transgenesis; however, technical limitations preclude this approach from assessing long-range enhancer activity. The proposed work overcomes this limitation by utilizing efficient enhancer replacement technology in mice to systematically characterize in vivo long-range enhancer activity and identify cis-regulatory sequences that are critical for this long-range activity. To determine the mechanism of long-range regulation by these remote control elements and identify regulatory proteins that bind them, this work will use a multidisciplinary approach to map and visualize enhancer–promoter contacts at macromolecular resolution. Functional characterization of remote control elements will expand the repertoire of known cis-regulatory elements, provide a novel general mechanism for long-range enhancer– promoter communication, and substantially advance knowledge of the role of 3D-genome organization in the context of development and disease.

项目摘要 远处（“远程”）的转录增强子受转录因子（TFS）驱动基因表达 时空的模式。通过安装链接的证据来强调此功能的重要性 影响远程增强子对人类疾病的突变。尚不清楚这些远程增强器如何 在长长的基因组距离上调节其靶基因，通常会跳过干预基因。远程增强器 通常在高阶染色质组织的支持下与目标启动子进行互动，但 该染色质组织不会废除增强子 - 促进剂的相互作用。哪些其他因素调节 这些远程监管相互作用在很大程度上仍然是出乎意料的。为了解决这个中心问题， 拟议的项目将使用新型的功能获得方法来识别介导的顺式调节序列 在哺乳动物发育的背景下，远程增强子 - 启动器通信。传统方法 体内增强子活性的评估依赖于增强子 - 重生蛋白转基因；但是，技术限制 排除这种方法无法评估远程增强剂活性。拟议的工作克服了这一点 通过在小鼠中使用有效的增强剂替代技术来系统地表征体内的限制 远程增强剂活性并识别对这种远程活性至关重要的顺式调节序列。到 通过这些遥控元素确定远程调节的机制并确定调节 绑定它们的蛋白质，这项工作将使用多学科的方法来映射和可视化增强剂 - 促进剂 大分子分辨率的接触。遥控元素的功能表征将扩展 已知的顺式调节元素的曲目为远程增强子 - 发起人的沟通，并大大提高了3D基因组组织在 发展和疾病的背景。