Chemical strategies to investigate biochemical crosstalk in human chromatin

研究人类染色质生化串扰的化学策略

• 批准号: 10621634
• 负责人: Champak Chatterjee
• 金额: $ 36.69万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-01 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10621634
• 关键词: Base Pairing; Biochemical; Biological; Biological Assay; Biophysics; Cells; Chemicals; Chromatin; Chromatin Structure; Complex; Disease; Gene Expression Regulation; Genes; Goals; Heterogeneity; Histone H2B; Histone H4; Histones; Human; Methylation; Modification; Molecular; Nucleoproteins; Play; Post-Translational Protein Processing; Protein Family; Proteins; Regulation; Research Project Grants; Role; Site; Sumoylation Pathway; TP53 gene; Techniques; cellular engineering; gene function; genetic information; histone modification; human disease; new therapeutic target; synthetic protein; therapeutic target; tool; transcription factor

PROJECT SUMMARY Chromatin is a massive nucleoprotein complex that packages about three billion base pairs of genetic information in nucleated cells. Histones and transcription factors (TFs) are two important families of proteins associated with chromatin that play key roles in organizing, protecting and activating our genes. A conserved feature among histones and TFs is the critical role that post-translational modifications (PTMs) play in controlling their diverse functions. Many gaps remain in our understanding of the mechanistic roles for specific histone and TF PTMs that are either low in abundance or where the necessary molecular biological and chemical tools are unavailable for mechanistic studies. This is especially challenging when studying regulation of the tumor suppressor p53 that is rapidly turned over by the proteasomal machinery in our cells. The proposed research project seeks to overcome challenges arising from the low abundance and heterogeneity of histone and p53 modifications by applying a combination of chemical and molecular biological tools to generate site-specifically modified proteins. Specifically, sumoylated histones H2B and H4 and methylated p53 will be generated by new protein semisynthesis techniques. The semisynthetic proteins will be subjected to a range of biophysical and biochemical assays in order to elucidate the mechanistic roles for sumoylation and methylation in regulating chromatin structure and function. Results from biochemical assays will be further validated in cell- based studies to arrive at a complete picture of the molecular mechanisms underlying gene regulation by histone sumoylation and p53 methylation. The long-term goal of this project is to identify new biochemical relationships, or crosstalk, in cellular chromatin that may be controlled to engineer cellular fates and selectively therapeutically targeted in human diseases.

项目概要 染色质是一种巨大的核蛋白复合物，包裹着约 30 亿个核蛋白 有核细胞中遗传信息的碱基对。组蛋白和转录因子 (TF) 是与染色质相关的两个重要蛋白质家族，它们发挥着关键作用 在组织、保护和激活我们的基因中发挥着重要作用。其中一个保守的特征 组蛋白和 TF 是翻译后修饰 (PTM) 在 控制它们的不同功能。我们的理解仍然存在许多差距 丰度较低的特定组蛋白和 TF PTM 的机制作用 或无法获得必要的分子生物学和化学工具的地方 机理研究。当研究监管时，这尤其具有挑战性。 肿瘤抑制因子 p53 可以通过我们体内的蛋白酶体机制快速转换 细胞。拟议的研究项目旨在克服以下挑战： 组蛋白和 p53 修饰的低丰度和异质性 结合化学和分子生物学工具来生成位点特异性 修饰的蛋白质。具体来说，组蛋白 H2B 和 H4 以及甲基化 p53 将通过新的蛋白质半合成技术产生。半合成的 蛋白质将接受一系列生物物理和生化测定，以便 阐明苏酰化和甲基化在调节染色质中的机制作用 结构和功能。生化检测结果将在细胞中进一步验证 基于研究来全面了解潜在的分子机制 通过组蛋白 sumoylation 和 p53 甲基化进行基因调控。长期目标是 该项目旨在识别细胞中新的生化关系或串扰 可以控制染色质来改造细胞命运并选择性地 人类疾病的治疗目标。