Mechanisms of chromatin regulation of transcription

染色质转录调控机制

• 批准号: 10490242
• 负责人: Michael Guy Poirier
• 金额: $ 73.2万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-17 至 2026-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10490242
• 关键词: Aging; Binding Proteins; Binding Sites; Biochemical; Biology; Biophysics; Cell Differentiation process; Cellular biology; Chemicals; Chromatin; Collaborations; Complex; Development; Disease; Drug resistance; Epigenetic Process; Fluorescence; Foundations; Gene Expression Regulation; Genetic; Genetic Transcription; Genome; Histone H3; Histones; Human; In Vitro; Individual; Ligation; Malignant Neoplasms; Methods; Molecular; Nature; Nucleosomes; Pharmacotherapy; Post-Translational Protein Processing; Reader; Research; SAGA; Site; Spectrum Analysis; Testing; Transcriptional Regulation; Work; XCL1 gene; base; chromatin remodeling; design; embryonic stem cell; epigenetic drug; epigenetic therapy; in vivo; insight; novel therapeutics; programs; single molecule; tool; transcription factor; tumorigenesis

MECHANISMS OF CHROMATIN REGULATION OF TRANSCRIPTION PROJECT SUMMARY Over the past decade, the Poirier lab has worked to contribute to the fields of chromatin biology and transcription regulation by developing a research program that is focused on the mechanisms by which chromatin and nucleosome structural dynamics regulate genome accessiblity to transcription regulatory complexes. We have developed and applied on our own and through collaborations, a wide range of experimental tools that enable mechanistic studies including single molecule fluorscence, single molecule force spectroscopy, ensemble fluorescence, and histone chemical ligations. In combination, we have quantitatively investigated how chromatin regulators including histone post translational modifications (PTMs), PTM binding proteins (readers), chromatin remodelers, linker histones, and transcripton factors function to control genome accessiblity to chromatin regulatory complexes. Building off this work, we propose to investigate two central questions in the fields of chromatin biology and transcription in the coming 5 years: (i) How do pioneer transcription factors target their recognition sites within compact nucleosomes and chromatin, and then facilitate chromatin decompaction? (ii) How do epigenetic regulators function together to synergistically or antagonistically regulate genome accessibility to transcription regulatory complexes? Pioneer factors (PFs) are master regulators of cell differentiation, are correlated with nucleosome depletion, and somehow access their binding sites within compact chromatin that are inaccessible to canonical transcription factors (TFs). Furthermore, PF disfunction is strongly correlated with disease most notably cancer. By combining single molecule and ensemble studies we will directly test distinct mechanisms of PF function, and determine what differentiates PFs from canonical TFs. Furthermore, through collaborative work we will investigate the same PFs in vivo to determine how the PF mechanisms that emerge from our in vitro studies impact their functions in vivo. This first direction will provide key insights into how pioneer factors gain access to their sites within compact chromatin and what differentiates PFs from canonical TFs. Epigenetic regulators including histone PTMs, and their readers and writers are critical to organismal development, aging and numerous diseases including cancer. We and others have investigated these regulators individually, yet how they function in combination remains largely unknown. Leveraging our ability to prepare core histone and most recently linker histones with any combination of PTMs, histone H3 readers, and most recently biochemical quantities of endogenous human SAGA and ATAC complexes, we will use single molecule, ensemble and ES cell-based methods to determine how biologically relevant combinations of these regulators control accessibility to canonical TFs and influence PFs. This second direction will provide a mechanistic and functional foundation for understanding how key epigenetic transcriptional regulators differentially control chromatin dynamics, accessibility and transcription.

染色体易位的染色质调节机制 项目摘要 在过去的十年中，Poirier实验室一直致力于为染色质生物学领域做出贡献， 转录调控通过开发一个研究计划，重点是机制， 染色质和核小体结构动力学调节基因组对转录调节的可接近性 配合物我们已经开发和应用了我们自己和通过合作，广泛的 实验工具，使机制的研究，包括单分子荧光，单分子力 光谱学、系综荧光和组蛋白化学连接。结合起来，我们在数量上 研究了染色质调节剂，包括组蛋白翻译后修饰（PTM），PTM结合 蛋白质（阅读器），染色质重塑，连接组蛋白和转录因子的功能，以控制基因组 染色质调节复合物的可接近性。在这项工作的基础上，我们建议调查两个中心 在染色质生物学和转录领域的问题在未来5年：（i）如何做先锋 转录因子将它们的识别位点定位在紧密的核小体和染色质内，然后 促进染色质分解？(ii)表观遗传调节因子如何协同作用， 拮抗性地调节基因组对转录调节复合物的可及性？ 先锋因子（Pioneer Factor，PF）是细胞分化的主要调节因子，与核小体耗竭相关， 并以某种方式进入它们在紧凑的染色质中的结合位点，这些位点是典型的 转录因子（TF）。此外，PF功能障碍与疾病密切相关， 癌通过结合单分子和系综研究，我们将直接测试PF的不同机制 函数，并确定PF与规范TF的区别。此外，通过合作 我们将在体内研究相同的PF，以确定我们的体外实验中出现的PF机制如何。 研究影响其在体内的功能。这第一个方向将提供关键的见解如何先锋因素获得 进入致密染色质内的位点以及PF与典型TF的区别。 包括组蛋白PTM在内的表观遗传调节因子及其阅读器和编写器对生物体的 发育、衰老和包括癌症在内的多种疾病。我们和其他人已经调查了这些 这些监管机构单独运作，但它们如何结合起来发挥作用仍在很大程度上未知。利用我们的能力， 用PTM、组蛋白H3读取器的任何组合制备核心组蛋白和最近的接头组蛋白，以及 最近的内源性人佐贺和ATAC复合物的生化量，我们将使用单一的 分子，系综和ES细胞为基础的方法，以确定如何生物相关的组合，这些 调节器控制对典型TF的可接近性并影响PF。这第二个方向将提供一个 机制和功能的基础，了解关键的表观遗传转录调控 差异控制染色质动力学、可及性和转录。