Capturing the dynamic epigenome using single molecule and single cell approaches

使用单分子和单细胞方法捕获动态表观基因组

• 批准号: 10402130
• 负责人: Kaushik Ragunathan
• 金额: $ 8.23万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-08-01 至 2022-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10402130
• 关键词: Architecture; Award; Binding; Biochemistry; Career Choice; Categories; Cells; Centromere; Chromatin; Chromosome Segregation; DNA; DNA Binding Domain; Defect; Epigenetic Process; Exhibits; Fission Yeast; Funding; Future; Gene Expression; Gene Expression Profile; Gene Silencing; Genetic; Genomic Instability; Goals; Health; Heritability; Heterochromatin; Histone H3; Human; In Vitro; Lead; Lysine; Maintenance; Methylation; Parents; Phenotype; Proteins; Reader; Research; Role; Tetracyclines; Training; United States National Institutes of Health; cancer cell; career; epigenetic silencing; epigenome; genetic approach; graduate student; histone modification; innovation; programs; reconstitution; single molecule; telomere

ABSTRACT Our bodies consist of billions of genetically identical cells that can exhibit distinct phenotypic or epigenetic states. The covalent and reversible modification of histones enables cells to establish heritable gene expression patterns without altering their genetic blueprint. Epigenetic mechanisms that control gene expression are essential to maintain cellular identity and program multicellular differentiation. Histone H3 lysine 9 methylation (H3K9me) is associated with transcription silencing and heterochromatin formation. Fission yeast (S. pombe) has a minimalist heterochromatin architecture that is amenable to high-throughput genetics and biochemistry. A trio of conserved proteins regulates heterochromatin, which includes, 1) an H3K9me specific ''writer,'' Clr4Suv39h that catalyzes H3K9me 2) an H3K9me specific ''reader,'' Swi6HP1 that binds to H3K9me chromatin and silences transcription and, 3) an H3K9me specific ''eraser,'' Epe1JmjC, that opposes heterochromatin assembly and epigenetic inheritance. Fusing Clr4 to the tetracycline-inducible TetR DNA binding domain facilitates rapid and reversible control of heterochromatin assembly. My lab’s innovative genetic strategy has enabled us to identify chromatin-associated factors with unique roles that are restricted to heterochromatin maintenance. As part of the Research Supplements to Promote Diversity in Health-Related Research program, the supplemental funding request will investigate the mechanisms that influence epigenetic silencing downstream of H3K9 methylation establishment. Specifically, the graduate student supported by this application will use in vitro reconstitution and single-molecule approaches to investigate the dynamic chromatin rearrangements that lead to epigenetic silencing. The supplement award will promote the research, career and professional training of a graduate student belonging to an NIH designated underrepresented category. The program of research outlined in this application is well-aligned and collaboratively developed with the candidate’s future career aspirations. Furthermore, all of the scientific goals described here are fully consistent with the proposed research directions of the parent award.

摘要 我们的身体由数十亿个基因相同的细胞组成，这些细胞可以表现出不同的表型或表观遗传学 各州。组蛋白的共价和可逆修饰使细胞能够建立可遗传的基因 在不改变其基因蓝图的情况下，表达模式。控制基因的表观遗传机制 表达对于维持细胞特性和规划多细胞分化是必不可少的。组蛋白H3赖氨酸 9甲基化(H3K9me)与转录沉默和异染色质形成有关。裂解酵母 (S.pombe)有一个最低限度的异染色质结构，它服从于高通量的遗传学和 生物化学。三种保守的蛋白质调节异染色质，包括：1)H3K9me专一性 ‘’编写器，‘’催化H3K9Me的Clr4Suv39h 2)H3K9Me特异的‘’读取器，‘’与H3K9Me结合的Swi6HP1 染色质和沉默转录，以及3)H3K9me特有的“橡皮”，Epe1JmjC，它反对 异染色质组装与表观遗传。CLR4与四环素诱导的TetR DNA融合 结合结构域有助于异染色质组装的快速和可逆控制。我的实验室很有创意 遗传策略使我们能够识别具有独特作用的染色质相关因子，这些因子仅限于 异染色质维持。作为促进与健康相关的多样性的研究补充资料的一部分 研究计划，补充资金申请将调查影响表观遗传学的机制 沉默H3K9甲基化下游的建立。具体地说，这所资助的研究生 应用将使用体外重组和单分子方法来研究动态染色质 导致表观遗传沉默的重排。补充奖将促进研究、职业和 对属于NIH指定的代表性不足类别的研究生进行的专业培训。这个 本申请中概述的研究计划与 候选人未来的职业抱负。此外，这里描述的所有科学目标都是完全一致的 提出了家长奖的研究方向。