Biochemical studies of aberrant chromatin regulation in cancer

癌症中异常染色质调控的生化研究

• 批准号: 10305344
• 负责人: Nicholas Prescott
• 金额: $ 4.75万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10305344
• 关键词: ATP Hydrolysis; Address; Architecture; Binding; Biochemical; Biochemistry; Biology; Biophysics; Cancer Biology; Case Study; Cell Line; Cell physiology; Cells; Cellular biology; Chemicals; Chromatin; Chromatin Fiber; Chromatin Remodeling Factor; Chromatin Structure; Chronic; Chronic Hepatitis B; Circular DNA; Complex; Cryoelectron Microscopy; DNA; Data; Detection; Disease; Enzymes; Epigenetic Process; Eukaryotic Cell; Faculty; Family; Foundations; Functional disorder; Gatekeeping; Gene Expression; Gene Expression Profiling; Genes; Genetic Transcription; Genome; Goals; Hepatitis B Virus; Hepatocyte; Histones; Homeostasis; Host Defense; Immune; In Vitro; Individual; Investigation; Knock-out; Libraries; Light; Maintenance; Malignant Neoplasms; Malignant neoplasm of liver; Mass Spectrum Analysis; Molecular Biology; Molecular Conformation; Molecular and Cellular Biology; Mutate; Mutation; Nucleosome Core Particle; Nucleosomes; Oncogenic; Oncornaviruses; Output; Pathologic; Phase; Play; Population; Positioning Attribute; Post-Translational Protein Processing; Primary carcinoma of the liver cells; Proteins; Proteomics; Recombinants; Regulation; Regulatory Pathway; Research; Research Project Grants; Resolution; Rest; Role; Signal Transduction; Stimulus; Structure; System; Techniques; Testing; Therapeutic Intervention; Viral Proteins; Virus; Virus Replication; Work; biophysical analysis; cancer type; career; chromatin protein; chromatin remodeling; chronic infection; clinically relevant; crosslink; epigenome; epigenomics; established cell line; experience; improved; insight; member; mutant; novel strategies; novel therapeutics; overexpression; reconstitution; recruit; response; skills; structural biology; structural genomics; transcription factor; transcriptome; tumorigenesis; ATP Hydrolysis; Address; Architecture; Binding; Biochemical; Biochemistry; Biology; Biophysics; Cancer Biology; Case Study; Cell Line; Cell physiology; Cells; Cellular biology; Chemicals; Chromatin; Chromatin Fiber; Chromatin Remodeling Factor; Chromatin Structure; Chronic; Chronic Hepatitis B; Circular DNA; Complex; Cryoelectron Microscopy; DNA; Data; Detection; Disease; Enzymes; Epigenetic Process; Eukaryotic Cell; Faculty; Family; Foundations; Functional disorder; Gatekeeping; Gene Expression; Gene Expression Profiling; Genes; Genetic Transcription; Genome; Goals; Hepatitis B Virus; Hepatocyte; Histones; Homeostasis; Host Defense; Immune; In Vitro; Individual; Investigation; Knock-out; Libraries; Light; Maintenance; Malignant Neoplasms; Malignant neoplasm of liver; Mass Spectrum Analysis; Molecular Biology; Molecular Conformation; Molecular and Cellular Biology; Mutate; Mutation; Nucleosome Core Particle; Nucleosomes; Oncogenic; Oncornaviruses; Output; Pathologic; Phase; Play; Population; Positioning Attribute; Post-Translational Protein Processing; Primary carcinoma of the liver cells; Proteins; Proteomics; Recombinants; Regulation; Regulatory Pathway; Research; Research Project Grants; Resolution; Rest; Role; Signal Transduction; Stimulus; Structure; System; Techniques; Testing; Therapeutic Intervention; Viral Proteins; Virus; Virus Replication; Work; biophysical analysis; cancer type; career; chromatin protein; chromatin remodeling; chronic infection; clinically relevant; crosslink; epigenome; epigenomics; established cell line; experience; improved; insight; member; mutant; novel strategies; novel therapeutics; overexpression; reconstitution; recruit; response; skills; structural biology; structural genomics; transcription factor; transcriptome; tumorigenesis

Project Summary/Abstract Eukaryotic cells require the tight regulation of global gene expression to maintain homeostasis and respond to environmental stimuli. DNA spools around histone proteins form this vital structure, chromatin, and provide a platform for the sophisticated tuning of gene expression through physical and chemical regulation. Unsurprisingly, the disruption of these chromatin regulatory mechanisms is particularly prevalent in cancers as a driver of disease. Completion of the proposed projects will shed light on the mechanisms of healthy chromatin regulation and its disruption in disease, providing the insight necessary to develop improved therapeutic interventions in a variety of cancers. In the F99 phase of this proposal, I study disrupted chromatin signaling by Hepatitis B Virus (HBV), a leading cause of hepatocellular carcinoma worldwide. HBV maintains chronic infections within hepatocytes by establishing an independent minichromosome, termed covalently closed circular DNA (cccDNA), that largely evades immune detection and conventional chromatin regulatory mechanisms. Further contributing to this evasion is the viral protein HBx, which has documented roles redirecting numerous chromatin effectors, including transcription factors, degradation machinery, and epigenome modifiers. So far in my thesis work, I have developed a platform to reconstitute cccDNA in vitro for biochemical and biophysical studies, determined that histone occupancy in cccDNA is required for HBx expression, and shown that HBx binds directly to nucleosomes. The remainder of my thesis work will be spent testing the biochemical effects of other known interactors on cccDNA compaction and gene expression and illuminating the cccDNA landscape in cells using locus-specific proteomic and epigenomic studies. The K00 phase shifts focus to ATP-dependent chromatin remodeling enzymes, a class of proteins shown to be mutated or overexpressed in more than 20% of cancers. In particular, I intend to study the CHD family of remodelers, which have been implicated as drivers of a variety of cancer types. I will apply my expertise in chromatin biochemistry and expand my technical repertoire to include cryo-electron microscopy as a means to study the structure and function of CHD chromatin remodelers. In parallel, I will develop skills in gene editing techniques to knockout wild-type enzyme and introduce clinically-relevant CHD mutants into cells for epigenomic analyses of remodeler dysfunction in disease. Combining these new approaches with my background in biochemistry, chemical biology, and biophysics will position me to address pressing questions in chromatin and cancer biology throughout the rest of my career as I pursue an independent, cancer-focused faculty position.

项目摘要/摘要 真核细胞需要严格调节全球基因表达以维持稳态 并应对环境刺激。 DNA周围围绕组蛋白蛋白启动这一重要 结构，染色质，并为基因表达的复杂调整提供了一个平台 通过物理和化学调节。毫不奇怪，这些染色质的破坏 作为疾病驱动力的癌症，调节机制尤其普遍。完成 拟议的项目将阐明健康染色质调节及其的机制 疾病的破坏，提供了改善治疗性所必需的见解 各种癌症的干预措施。 在该提案的F99阶段，我研究了丙型肝炎的染色质信号传导 病毒（HBV），全球肝细胞癌的主要原因。 HBV保持慢性 通过建立独立的微型小体，称为共价，肝细胞内感染 闭合圆形DNA（CCCDNA），在很大程度上逃避了免疫检测和常规染色质 监管机制。进一步促进这种逃避的是病毒蛋白HBX，它具有 记录了重定向众多染色质效应子的角色，包括转录因子， 降解机械和表观基因组修饰符。到目前为止，在我的论文工作中，我已经开发了 在体外重构CCCDNA的生化和生物物理研究的平台，确定 HBX表达需要CCCDNA中的组蛋白占用率，并表明HBX直接结合 到核小组。我的其余论文工作将用于测试的生化效应 其他有关CCCDNA压实和基因表达的已知相互作用者，并照亮了 使用基因座特异性蛋白质组学和表观基因组研究中的CCCDNA景观。 K00相移到ATP依赖性染色质重塑酶，一类 在超过20％的癌症中显示出突变或过表达的蛋白质。特别是我 打算研究冠心病家族的重塑家族，这些家族被认为是一种多样性的驱动因素 癌症类型。我将在染色质生物化学上运用我的专业知识并扩展我的技术 曲目包括冷冻电子显微镜作为研究的手段 CHD染色质改造。同时，我将开发基因编辑技术的技能来淘汰 野生型酶，并将与临床相关的CHD突变体引入表观基因组学细胞 疾病重塑功能障碍的分析。将这些新方法与我的 生物化学，化学生物学和生物物理学的背景将使我定位以解决压力 在我追求的过程中 独立，以癌症为中心的教师职位。