Molecular impediments to fate-specifying pioneer factor activity during development

发育过程中决定命运的先锋因子活动的分子障碍

• 批准号: 10363623
• 负责人: Michael P Meers
• 金额: $ 3.8万
• 依托单位: FRED HUTCHINSON CANCER CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-03 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10363623
• 关键词: Acetyl Coenzyme A; Address; Award; Base Pairing; Binding; Biological; Biological Models; Buffers; Cell Differentiation process; Cell Fate Control; Cells; Chromatin; Chromatin Structure; Conflict (Psychology); Consumption; DNA; DNA Binding; DNA Sequence; Development; Developmental Gene; Disease; Downstream Enhancer; Educational Status; Enhancers; Ensure; Environment; Equilibrium; Event; Excision; Foundations; Future; Gene Expression; Gene Expression Regulation; Genes; Genetic Enhancer Element; Genetic Transcription; Genome; Heterogeneity; Histones; Hypoxia; Image; Maintenance; Malignant Neoplasms; Measures; Metabolic; Metabolic Control; Metabolism; Methods; Modeling; Molecular; Mutation; Nucleosomes; Outcome; Oxygen; Phase; Play; Pluripotent Stem Cells; Process; Regulation; Regulator Genes; Research; Resolution; Role; S-Adenosylhomocysteine; Site; Specific qualifier value; Structure; System; Techniques; Technology; Testing; Time; Training; Variant; base; career; cell type; chromatin remodeling; developmental disease; embryonic stem cell; epigenome; epigenomics; expectation; experience; experimental analysis; experimental study; genetic manipulation; improved; in vivo; innovation; insight; metabolomics; novel; prevent; programs; response; screening; stem cell differentiation; stem cells; theories; transcription factor

Project Summary/Abstract Transcription factors act as specifying agents of cell differentiation during development by binding to DNA enhancer sequences and activating them to control developmental gene expression. Enhancer activation is typically associated with the removal of nucleosomes, which decorate eukaryotic genomes and normally wrap roughly 150 base pairs of DNA in a highly stable configuration. A persistent puzzle of developmental gene regulation is how TFs bind and activate their target enhancers when they are initially wrapped in nucleosomes, which typically inhibit TF binding. One hypothesis posits that a special class of “pioneer factors” are able to bind their targets in the context of nucleosomal wrapping and displace the nucleosomes they bind to activate and expose the enhancer for downstream TF binding. However, it has been exceedingly difficult to confirm the presence of nucleosome binding “pioneer activity” in vivo, leaving the developmental roles of pioneer factors in question. We recently used high-resolution epigenome profiling to identify instances of nucleosome binding by pioneer factors that were enriched at enhancers with suboptimal motif binding sequences, presenting the intriguing possibility that pioneer activity is a mechanism to ensure the fidelity of enhancer activation at sites that are vulnerable to natural fluctuations in the local chromatin environment. Pioneer factors often function in early development, which maintains high fidelity despite natural variation in chromatin structure that is sensitive to the metabolic state of the cell. Therefore, pioneer factors may play a direct role in insulating developmental transitions against metabolic variance. However, the potential roles of pioneer factors in developmental fidelity and buffering against metabolic heterogeneity have not been uncovered to date. In this proposal, I will use a controlled pioneer factor expression system to study how pioneer factor-driven developmental changes are buffered against deliberate chromatin and metabolic perturbations. In Aim 1, I will test the hypothesis that pioneer activity facilitates developmental fidelity by observing development after genetically enforcing chromatin barriers to pioneer factor binding and inactivating the nucleosome binding pioneer activity of a specific pioneer factor. In Aim 2, I will use a model system of metabolic control of development to understand how pioneer factor binding responds to metabolic changes, and how specific pioneer factor-enhancer activation events underlie different developmental outcomes in response. These Aims will uncover mechanistic explanations for the disparity between variance in gene regulatory processes on the molecular level and the precision of cell fate outcomes on the developmental level, and my findings will be of direct consequence to diseases such as cancer where extreme heterogeneity overwhelms the checks and balances on cell fate. A K99/R00 Award will be instrumental in addressing these questions and furnishing me with high level training in new methods and biological theory that will prepare me to continue to pursue major research avenues related to pioneer factor and chromatin control of development in my future independent career.

项目摘要/摘要 转录因子通过与DNA结合在发育过程中指定细胞分化的药物的作用 增强子序列并激活它们以控制发育基因表达。增强子激活是 通常与去除真核基因组并通常包裹的核小体的去除有关 高度稳定的配置中大约150个DNA对。发育基因的持续难题 调节是TFS最初包裹在核小体中时结合和激活目标增强子的方式， 通常会抑制TF结合。一个假设认为，特殊类别的“先锋因素”能够约束 它们的目标是在核小体包裹的背景下，并取代它们结合到激活的核小体和 暴露增强子的下游TF结合。但是，很难确认 在体内存在核卫生结合的“先驱活性”，留下了先驱因素的发育作用 问题。我们最近使用高分辨率发作分析来确定核小体结合的实例 在具有次优基序结合序列的增强子上富集的先锋因素，呈现 先锋活动是确保在地点激活增强子激活的机制的有趣可能性 在局部染色质环境中容易受到自然波动的影响。先锋因素通常在早期起作用 开发，维持染色质结构的高富达任务自然变化，这对 细胞的代谢状态。因此，先锋因素可能在绝缘发育中发挥直接作用 反对代谢方差的过渡。但是，先锋因素在发展忠诚中的潜在作用 迄今为止，还没有发现针对代谢异质性的缓冲。在此提案中，我将使用 受控的先驱因子表达系统，以研究先锋因素驱动的发展变化是如何的 反对故意的染色质和代谢扰动。在AIM 1中，我将检验先锋的假设 活性通过观察遗传强化染色质屏障后的发展来促进发展忠诚度 先锋因子结合并灭活特定先锋因子的核小体结合先驱活性。 AIM 2，我将使用一种代谢控制的模型系统，以了解开拓性因素如何绑定 对代谢变化的反应，以及特定的先锋因子 - 增强剂激活事件是不同的 回应的发展结果。这些目标将发现差异的机理解释 在分子水平上基因调节过程的差异与细胞脂肪结果的精度之间 在发展水平上，我的发现将是对癌症等疾病的直接结果 极端异质性压倒了细胞命运的制止和平衡。 K99/R00奖项将是工具性的 在解决这些问题并为我提供新方法和生物学理论的高级培训时 这将使我能够继续购买与先驱因素和染色质控制有关的主要研究途径 我未来独立职业的发展。

项目成果

Multifactorial profiling of epigenetic landscapes at single-cell resolution using MulTI-Tag.

• DOI: 10.1038/s41587-022-01522-9
• 发表时间: 2023-05
• 期刊: NATURE BIOTECHNOLOGY
• 影响因子: 46.9
• 作者: Meers, Michael P.;Llagas, Geneva;Janssens, Derek H.;Codomo, Christine A.;Henikoff, Steven
• 通讯作者: Henikoff, Steven