Genome-wide mechanisms and dynamics of ATP-dependent chromatin remodeling complexes

ATP依赖性染色质重塑复合物的全基因组机制和动力学

基本信息

批准号：
10216103
负责人：
Sandipan Brahma
金额：
$ 10万
依托单位：
FRED HUTCHINSON CANCER RESEARCH CENTER
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-01 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10216103
关键词：
ARID1A gene ATAC-seq Acute Address Affect Automobile Driving Binding Biochemical Bioinformatics Biological Process British Columbia Catalytic Domain Cell physiology Cells Chromatin Chromatin Remodeling Factor Chromatin Structure Complex DNA DNA Packaging Data Defect Development Disease Enhancers Environment Family Foundations Fred Hutchinson Cancer Research Center Frequencies Future Gene Activation Gene Expression Genes Genetic Transcription Genome Genomic Segment Genomics Goals Heterogeneity Histones Human Genome Individual Kinetics Knowledge Label Lead Link Malignant Neoplasms Maps Mentors Microscopy Modeling Molecular Molecular Machines Mus Mutation Nucleosomes Phase Polycomb Positioning Attribute Process Protein Complex Subunit Protein Subunits Proteins Regulation Regulator Genes Regulatory Element Repression Research Research Personnel Research Training Residual state Resolution Role SMARCA4 gene SMARCB1 gene Saccharomycetales Scientist Site Specificity Structure Supervision Techniques Testing Time Tissues Training Transcriptional Regulation Universities Variant base cell type chromatin remodeling chromosomal location design developmental disease embryonic stem cell epigenomics exome sequencing genome-wide in vivo insight molecular imaging nervous system disorder novel novel strategies polybromo preference promoter recruit single molecule stem cell differentiation stem cells targeted treatment transcription factor

项目摘要

PROJECT SUMMARY/ABSTRACT The SWI/SNF family ATP-dependent chromatin remodelers are multi-subunit protein complexes that dynamically position chromosomal nucleosomes to modulate DNA accessibility, transcription-factor binding, and cell-type-specific gene expression. SWI/SNF function is crucial at several stages of mammalian development, and recent human whole-genome and exome sequencing studies revealed striking mutational frequencies in genes encoding SWI/SNF subunits across a range of diseases - from neurologic disorders to over 20% of all cancers. Disease-associated SWI/SNF mutations often cause the loss of a protein subunit that further destabilizes the complex and results in altered subunit composition and functions. Of note, heterogeneity in SWI/SNF subunit composition is also observed naturally within and across cell types, hypothesized to result in complex- and tissue-specific SWI/SNF functions. Despite their importance, how these subunits determine the substrate preference and mechanistic functions of SWI/SNF complexes are not well understood, largely due to the lack of robust approaches to characterize where they bind within the genome and how they interact with chromatin genome-wide. Thus, the main goal of this proposal is to address this key knowledge gap. Since chromatin remodelers directly interact with nucleosomes, it is important to clarify remodeler-bound nucleosome compositions and structures. Standard biochemical characterization of soluble components purified from cellular extracts cannot determine the nucleosome structures associated with chromatin-bound complexes. It is also important to understand the kinetic parameters of remodeler-chromatin interactions, such as how fast a remodeler is recruited to its target sites, and how long does it remain bound at its genomic sites. The candidate proposes to address these questions by using structural and functional epigenomics approaches and live-cell single-molecule imaging, to characterize remodeler-chromatin interactions in the context of the complex and dynamic chromatin environment inside cells. Aim 1 is to determine the genome-wide occupancy of SWI/SNF complexes with distinct subunit compositions. Aim 2 is to determine a) the structures and histone composition of remodeler-bound nucleosomes genome-wide, and b) the kinetic parameters of remodeler-chromatin interactions in live cells. Aim 3 is to study the interaction of SWI/SNF with repressive chromatin. The mentored phase of this project will be completed under the sponsorship of Dr. Steven Henikoff at the Fred Hutchinson Cancer Research Center. The candidate will acquire training in live-cell single-molecule imaging under the supervision of Dr. Sheila Teves at the University of British Columbia. The proposed research and training will provide a strong foundation for the candidate to develop as an independent investigator, studying chromatin remodeling mechanisms and dynamics in the regulation of fundamental cellular processes.

项目概要/摘要 SWI/SNF 家族 ATP 依赖性染色质重塑蛋白是多亚基蛋白复合物，动态定位染色体核小体以调节 DNA 可及性、转录因子结合、和细胞类型特异性基因表达。 SWI/SNF 功能在哺乳动物的几个阶段至关重要的发展，最近的人类全基因组和外显子组测序研究揭示了惊人的突变编码 SWI/SNF 亚基的基因在一系列疾病中的频率 - 从神经系统疾病到占所有癌症的 20% 以上。与疾病相关的 SWI/SNF 突变通常会导致蛋白质亚基的丢失，进一步破坏复合物的稳定性并导致亚基组成和功能改变。值得注意的是， SWI/SNF 亚基组成的异质性也在细胞类型内和细胞类型之间自然观察到，假设导致复杂和组织特异性的 SWI/SNF 功能。尽管它们很重要，但这些亚基决定SWI/SNF复合物的底物偏好和机械功能不佳理解，很大程度上是由于缺乏强有力的方法来表征它们在基因组中的结合位置以及它们如何与全基因组染色质相互作用。因此，本提案的主要目标是解决这一关键问题知识差距。由于染色质重塑剂直接与核小体相互作用，因此澄清这一点很重要重塑剂结合的核小体组成和结构。可溶性物质的标准生化表征从细胞提取物中纯化的成分无法确定与相关的核小体结构染色质结合复合物。了解重塑染色质的动力学参数也很重要交互，例如重塑者被招募到其目标地点的速度，以及它在该地点保持束缚的时间它的基因组位点。候选人建议通过使用结构和功能来解决这些问题表观基因组学方法和活细胞单分子成像，以表征重塑染色质细胞内复杂且动态的染色质环境中的相互作用。目标 1 是确定具有不同亚基组成的 SWI/SNF 复合物的全基因组占有率。目标 2 是确定 a) 全基因组范围内重塑蛋白结合核小体的结构和组蛋白组成，以及 b) 活细胞中重塑剂-染色质相互作用的动力学参数。目标3是研究相互作用 SWI/SNF 具有抑制性染色质。该项目的指导阶段将在 Fred Hutchinson 癌症研究中心的 Steven Henikoff 博士的赞助。候选人将在大学 Sheila Teves 博士的指导下接受活细胞单分子成像培训不列颠哥伦比亚省。拟议的研究和培训将为候选人提供坚实的基础发展成为一名独立研究者，研究染色质重塑机制和动态基本细胞过程的调节。