Chromatin Organization Regulates Osteogenesis

染色质组织调节成骨

• 批准号: 10316201
• 负责人: Jane B. Lian
• 金额: $ 50.13万
• 依托单位: UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-06 至 2024-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10316201
• 关键词: Acetylation; Acute; Affect; Aging; Alkaline Phosphatase; Binding; Biological Assay; Bone Development; Bone Regeneration; Bone Surface; Bone Tissue; CCCTC-binding factor; CRISPR/Cas technology; Cell Lineage; Cells; Chromatin; Chromatin Structure; Chromosomes; Clustered Regularly Interspaced Short Palindromic Repeats; Competence; Complement; Complex; Control Groups; Data Set; Development; Enhancers; Epigenetic Process; Female; Future; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genome; Genomic Segment; Genomics; Hi-C; Higher Order Chromatin Structure; Histology; Impairment; Implant; Injections; Knowledge; Link; Mature Bone; Mediating; Mesenchymal Stem Cells; MicroRNAs; Modeling; Modification; Mus; Osteoblasts; Osteogenesis; Outcome; Phenotype; Publishing; Regulatory Element; Research; Resolution; Role; Selection Criteria; Supporting Cell; Testing; Therapeutic; Time; Tissues; Transcriptional Regulation; Transplantation; Undifferentiated; base; bone; bone loss; chromosomal location; clinically relevant; cohort; differential expression; functional genomics; gene network; histone modification; in vivo; in vivo evaluation; male; mouse model; novel; osteoblast differentiation; osteogenic; preventive intervention; programs; protein complex; skeletal disorder; stem cells; subcutaneous; transcription factor; transcriptome sequencing; translational applications; treatment strategy

SUMMARY This new R01 builds on discoveries during the R37 period (2008-2018) that established epigenetic mechanisms (miRNAs, histone modifications) regulating osteoblast differentiation. We characterized for the first time a “signature” of specific histone modifications that are associated with dynamic changes in gene expression during the temporal progression of osteogenesis. These histone modifications also predicted “enhancers”, which are critical cis-regulatory elements that contribute to local gene expression. We now propose to examine the recently recognized “super enhancer” domains (SEDs) that include regulatory elements for multiple transcription factors that have emerged as key regulators of cell phenotypes. SEDs function in chromatin organization via long range intra- and inter-chromosomal interactions that coordinate control of gene cohorts responsible for lineage specification and distinct cell identity. Our preliminary studies have identified a subset of SEDs that we now propose are putative “bone-essential super-enhancers” and candidates for the important decision stage of commitment to osteogenesis from MSCs. We hypothesize that super-enhancer domains are differentially activated from the undifferentiated MSC to the osteoblast commitment stage, and function to establish the osteogenic phenotype by coordinately regulating gene networks and contributing to higher order chromatin organization that supports cell identity. Our studies will in: Aim1- analyze the functional effects of prioritized SEDs we have identified related to osteoblastogenesis and mature bone activities through directed inhibition and activation of SEDs using CRISPR/Cas9 in MSCs; Aim 2- determine the chromosomal domains that interact with SEDs to control multiple genes and networks that commit MSCs to the osteoblast phenotype through chromatin organization; and Aim 3- demonstrate in mouse models that using CRISPR activated SEDs in MSCs will stimulate bone formation. Impact: These studies pioneer a new level of gene regulation for MSC lineage commitment to osteogenesis, based on an emerging understanding of SED functions in other tissues but have been minimally studied in bone. By characterizing SED mechanisms related to chromatin organization and stabilization in MSCs, we will discover novel mechanisms of multi-dimensional coordinate control of transcriptional hubs and protein complexes within an SED that is responsible for establishing commitment to the osteoblast phenotype. Importantly, knowledge of the chromatin organization that stabilizes the osteogenic phenotype impacts on future novel treatment strategies for skeletal disorders.

概括 这种新的R01建立在R37时期（2008-2018）的发现上，该发现建立了表观遗传机制 （miRNA，组蛋白修饰）调节成骨细胞分化。我们首次表征了 特定组蛋白修饰的“签名”与基因表达的动态变化相关的特定组蛋白修饰。 成骨的暂时进展。这些组蛋白修饰还预测了“增强子”，这是 关键的顺式调节元素有助于局部基因表达。我们现在建议检查最近 公认的“超级增强子”域（SED），其中包括多个转录因子的调节元素 已成为细胞表型的关键调节剂。 SEDS通过远距离的染色质组织功能 染色体内和染色体间相互作用，这些相互作用协调负责谱系的基因组的控制 规范和不同的细胞身份。我们的初步研究已经确定了我们现在的SED子集 提案是推定的“骨头上的超级增强剂”和候选人的重要决策阶段 对MSC的成骨作用的承诺。我们假设超级增强剂域是差异的 从未分化的MSC激活到成骨细胞承诺阶段，并建立功能 通过协调调节基因网络并有助于高阶染色质，成骨表型 支持细胞身份的组织。我们的研究将在：AIM1-分析优先SED的功能效应 我们已经通过定向抑制和 在MSC中使用CRISPR/CAS9激活SED； AIM 2确定相互作用的染色体结构域 使用SED控制多个基因和网络，这些基因和网络通过 染色质组织； AIM 3-在使用MSC中使用CRISPR激活的SED的鼠标模型中进行了3-演示 将刺激骨形成。 影响：这些研究先驱，一种新的基因调节水平，用于MSC谱系对成骨的承诺， 基于对SED功能的新兴理解，但在骨骼中的研究很少。 通过表征与染色质组织和MSC中稳定相关的SED机制，我们将发现 转录中心和蛋白质复合物多维坐标控制的新机制 负责建立对成骨细胞表型的承诺的SED。重要的是，知识 稳定成骨表型的染色质组织会影响未来的新型治疗策略 用于骨骼疾病。