Iteratively redefining developmental potential through poised enhancers

通过稳定的增强剂迭代地重新定义发展潜力

• 批准号: 9753014
• 负责人: Robert Blelloch
• 金额: $ 37.01万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-07 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9753014
• 关键词: Acetylation; Acute; Address; Binding; Binding Sites; Biochemical Genetics; Cell Differentiation process; Cell Fate Control; Cells; Collaborations; Complex; DNA Binding; Data; Deacetylation; Dependence; Development; Developmental Gene; Developmental Process; Disease; Embryo; Enhancers; Enzymes; Epiblast; Epigenetic Process; Family; Fostering; Future; Gene Activation; Gene Expression; Gene Expression Regulation; Genes; Genetic Epistasis; Goals; Health; Human; Knowledge; Malignant - descriptor; Measures; Mediator of activation protein; Methylation; Mission; Modification; Molecular; Movement; Mutation; Nature; Nucleosomes; Pathway interactions; Phenotype; Population; Post-Translational Protein Processing; Property; Proteins; Public Health; Regulation; Reporter; Research; Role; SMARCA4 gene; Signal Transduction; Site; Stem cells; Structure; System; Testing; Therapeutic; Therapeutic Effect; Time; United States National Institutes of Health; Work; base; cell type; chromatin remodeling; cohesin; design; embryonic stem cell; epigenome; experimental study; forkhead protein; gene product; genetic approach; improved; interest; mutant; novel; programs; promoter; public health relevance; recruit; spatiotemporal; stem; stem cell differentiation; stem cell population; stemness; time use; tool; transcription factor; transcription factor USF

PROJECT SUMMARY/ABSTRACT: There is a fundamental need to understand how transcription factors function together with their co-regulators to redefine a cell’s developmental potential through cell fate transitions. Our current understanding is limited to a small number of examples often based on simple linear pathways with a transcription factor upstream of its co-regulators either activating or repressing an enhancer, which in turn regulates its cognate gene in time and space. However, without a better understanding of the mutual dependencies between transcription factors and their co-regulators including epigenetic enzymes and collaborative DNA binding factors, it will be impossible to predict how manipulation of either will influence cell fate and developmental potential. The long-term goal of the lab is to understand all levels of molecular control of cell fate transitions in order to efficiently reprogram cells to desired phenotypes. The objective here is to focus on a transcription factor, Foxd3, which is essential to maintain the developmental potential of various stem cells. The central hypothesis is that Foxd3 is used iteratively in stem cells to redefine the cell’s development potential by establishing poised enhancers in association with its co-regulators and collaborative cell specific transcription factors. This hypothesis derives from preliminary data establishing a dual functional role for Foxd3 as a simultaneous activator and repressor through its interaction with and regulation of multiple epigenetic factors. As such it poises genes and redefines the developmental potential of different stem cell populations by moving to new enhancer sites. The following specific aims are proposed: 1) Determine the epistatic relationship between Foxd3, H3K4 methylation, nucleosome depletion, and H3K27 acetylation, 2) Uncover the mechanistic basis for Foxd3 movements during the embryonic stem to epiblast cell transition, 3) Identify role of Foxd3 in cohesin recruitment and enhancer- promoter looping during developmental gene activation. In aim 1, epistasis and structure-function analyses will be performed using mutants of the Foxd3 and its coregulators to determine the interdependencies between the factors in establishing a dual-functional complex at bound sites. In aim 2, post-translational modifications and collaboration with other transcription factors will be evaluated to dissect the mechanistic basis of Foxd3 movements. In aim 3, Foxd3’s role in cohesin recruitment and enhancer-promoter looping will be evaluated using time-course and epistasis experiments. The proposal is highly significant as it will provide novel paradigms of gene control central to a cell’s developmental potential. These paradigms are unlikely to be specific to Foxd3, but rather reflect general strategies used by stemness transcription factors to retain or induce a stem cell’s full potential. Such knowledge will allow for better-designed strategies for cell manipulation and improved ability to predict effects of therapeutics aimed at the epigenetic co-regulators.

项目摘要/摘要： 基本需要了解转录因子如何与其共同调节器一起发挥作用 通过细胞脂肪过渡重新定义细胞的发育潜力。我们目前的理解仅限于 少数示例通常基于简单的线性途径，其上游的转录因子上游 共同调节器要么激活或表达增强子，这反过来调节其同源基因的时间和 空间。但是，没有更好地了解转录因子和 他们的共同调节剂在内，包括表观遗传酶和协作DNA结合因子，将不可能 预测两者的操纵将如何影响细胞脂肪和发育潜力。长期目标 该实验室是要了解细胞脂肪转变的所有分子控制，以便有效地重新编程 细胞为所需的表型。这里的目的是关注转录因子FOXD3，这是必不可少的 维持各种干细胞的发育潜力。中心假设是使用FOXD3 在干细胞中迭代以通过在 与其共同调节剂和协作细胞特定转录因子相关联。该假设得出 从初步数据从建立FOXD3作为简单激活器和副本的双重功能作用 通过与多种表观遗传因素的相互作用和调节。因此，它可以使基因并重新定义 通过转移到新的增强子站点，不同干细胞种群的发展潜力。下列 提出了具体目的：1）确定FOXD3，H3K4甲基化的上任关系， 核小体耗竭和H3K27乙酰化，2）发现FOXD3运动的机械基础 胚胎茎到层细胞过渡，3）鉴定FOXD3在粘着素募集和增强子中的作用 发育基因激活期间启动子循环。在AIM 1中，上学和结构功能分析将 使用FOXD3及其核心节的突变体进行确定 在结合位点建立双功能复合物的因素。在AIM 2，翻译后修改和 将评估与其他转录因子的合作，以剖析FOXD3的机械基础 动作。在AIM 3中，将评估FOXD3在凝聚蛋白募集和增强器促销中的作用 使用时间课和上位实验。该提议非常重要，因为它将提供新颖 基因控制的范例是细胞发育潜力的中心。这些范式不太可能 特定于FOXD3，而是反映了干性转录因子使用的一般策略，以保留或 引起干细胞的全部潜力。这些知识将允许改善细胞操作的策略 并提高了预测针对表观遗传共同调节剂的理论影响的能力。