Interrogating the Structural Basis of Nuclear Receptor Activation on Chromatin

探究染色质核受体激活的结构基础

• 批准号: 10538474
• 负责人: Timothy Silas Strutzenberg
• 金额: $ 6.72万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-16 至 2025-09-15
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10538474
• 关键词: Adaptive Immune System; Area; Autoimmune Diseases; Binding; Biochemical; Biochemistry; Biological; Biological Assay; Biological Process; Biology; Biophysical Process; Biophysics; Chemicals; Chromatin; Chromatin Structure; Chronic; Chronic small plaque psoriasis; Clinic; Clinical; Clinical Trials; Complement; Complex; Consequentialism; Cryoelectron Microscopy; DNA; Data; Deuterium; Development; Disease; Drug Targeting; Drug usage; EP300 gene; Electron Microscopy; Enzymatic Biochemistry; Foundations; Gene Activation; Gene Expression; Gene Expression Regulation; Genes; Goals; Health; Histone Acetylation; Histones; Home; Hormones; Human; Hydrogen; Immunity; Industrialization; Inflammatory; Investigation; Laboratories; Laboratory Study; Length; Libraries; Ligands; Light; Mass Spectrum Analysis; Measurement; Mediating; Metabolic Diseases; Metabolism; Methodology; Modeling; Modification; Molecular; Molecular Conformation; Monitor; NCOA3 gene; Names; Nature; Nuclear Receptors; Nuclear Structure; Nucleosomes; Pathology; Physiology; Play; Positioning Attribute; Post-Translational Protein Processing; Protein Dynamics; Proteins; Proteomics; Receptor Activation; Regulation; Reproduction; Research; Resolution; Response Elements; Role; Safety; Side; Solvents; Structure; T cell differentiation; Techniques; Technology; Thymus Gland; Training; Transcriptional Activation; Vertebral column; Work; Writing; antimicrobial; base; biophysical techniques; birth control; chromatin remodeling; crosslink; cryogenics; design; drug development; fight against; histone modification; improved functioning; insight; member; next generation sequencing; novel; orphan nuclear receptor ROR-gamma; preference; programs; protein complex; receptor; receptor binding; reconstitution; recruit; side effect; structural biology; targeted treatment; therapeutic target; training project; transcription factor

PROJECT SUMMARY Nuclear receptors (NRs) are ligand-gated transcription factors that control important biological processes including reproduction, development, metabolism, and immunity. NRs are the target of ~16% of drugs used in the clinic for applications that include birth control, inflammatory disease, and metabolic disorders. NRs are thought to function by altering the dynamics of chromatin to alter expression levels of genes through the recruitment of coregulators, chromatin remodelers, and ultimately through the activation of transcriptional machinery. However, the dynamic nature of NRs and NR complexes has made it challenging to study the molecular mechanisms by which full length proteins and protein assemblies regulate gene expression using traditional structural biology techniques. Despite the central role that NRs play in gene expression activation, our mechanistic understanding of how they regulate target gene programs at the molecular level of chromatin is largely unknown. This is attributed, in part, to the dearth of structural biology data for full-length receptors and the near complete absence of biochemical, biophysical, or structural data on NRs in their functionally relevant chromatin context. Building on established foundations in sponsor’s home laboratory, combined with the applicant’s prior expertise in biochemical and biophysical methods and an intimate understanding of the NR field, this project seeks to determine how a model NR named RORγT assembles with coregulatory proteins SRC3 and p300 to alter chromatin dynamics and post-translational modification status of chromatin, a hallmark of NR target gene activation. The proposed research program has two major goals: (1) to determine how RORγT and coregulatory protein complexes engage with nucleosomes, and (2) to determine how RORγT and coregulator protein complexes modify histones and alter nucleosome dynamics. This will be accomplished using a combination of traditional biochemistry, and state-of-the-art technologies including cryogenic electron microscopy (cryo-EM) and structural proteomics. By studying how these protein complexes assemble and act on nucleosomes, we will gain new insight into how RORγT, and more broadly the NR superfamilty, modulates nucleosome dynamics, modification status, and, ultimately, function in gene regulation. The implications of this work will help us understand how an important drug target functions and will pave the way for future research investigating an important class of drug targets.

项目摘要 核受体是一种配体门控的转录因子，控制着重要的生物学过程 包括生殖、发育、代谢和免疫。NRs是约16%的药物的目标， 该诊所的应用包括节育、炎症性疾病和代谢紊乱。NR是 被认为是通过改变染色质的动力学来改变基因的表达水平， 募集辅调节因子，染色质重塑，并最终通过激活转录 机械.然而，NR和NR复合物的动态性质使得研究NR具有挑战性 全长蛋白质和蛋白质组装体调控基因表达的分子机制 传统的结构生物学技术。尽管核受体在基因表达激活中起着核心作用，但我们的研究表明， 对它们如何在染色质的分子水平上调节靶基因程序的机械理解是 大部分未知。这部分归因于缺乏全长受体的结构生物学数据， 几乎完全缺乏关于NR在其功能相关方面的生物化学、生物物理学或结构数据， 染色质背景。在申办者家庭实验室的基础上，结合 申请人在生物化学和生物物理方法方面的先前专业知识以及对NR领域的深入了解， 该项目试图确定一个名为RORγT的NR模型如何与协同调节蛋白SRC 3组装 和p300改变染色质动力学和染色质的翻译后修饰状态，这是NR的标志 靶基因激活。该研究计划有两个主要目标：（1）确定RORγT和 辅调节蛋白复合物与核小体结合，以及（2）确定RORγT和辅调节蛋白如何与核小体结合。 蛋白质复合物修饰组蛋白并改变核小体动力学。这将通过使用 结合传统的生物化学和最先进的技术，包括低温电子 显微镜（cryo-EM）和结构蛋白质组学。通过研究这些蛋白质复合物如何组装和作用 在核小体上，我们将对RORγT以及更广泛的NR超家族如何调节 核小体动力学、修饰状态，以及最终在基因调控中的功能。这意味着 这项工作将帮助我们了解一个重要的药物靶点是如何发挥作用的，并为未来的研究铺平道路。 研究一类重要的药物靶点