Program Project: BONE CELL STRUCTURE AND GENE EXPRESSION

总体说明（由申请人提供）：我们的计划 继续讨论核结构参数 （染色质组织和核基质的组装和活性） 相关的亚核转录位点）有助于基因表达 介导骨细胞表型的发生、发展和维持， 骨骼发育和体内平衡所需的特性。我们现在搞的 一个综合的，团队的方法，结合分子，细胞，生物化学和 体内遗传分析，以确定亚核组织 核酸和调节蛋白的结合促进了 支持骨骼基因表达能力的生理信号， 体内骨发育。我们的计划已经：1）证明重塑 与发育诱导相关的染色质和核小体组织 和类固醇激素增强骨特异性转录; 2）鉴定骨 整合骨骼肌活性的特异性激活剂和共调节剂 基因启动子元件; 3）证明Runx 2（Cbfal/AML 3）因子是 点状定位于离散的核基质相关的亚核位点; 和4）结构和功能特征的独特的核内 引导Runx 2到达转录位点的靶向信号 在体内进行骨骼发育。我们现在将研究 支持了我们的基本观测， 细胞核内调节复合物的组织和组装是 控制骨形成的基因表达所必需的。项目1将 研究指导骨特异性转录因子和 共调节蛋白质到细胞核内的位点， 发生类固醇激素应答基因表达。项目2将决定 Runx 2的亚核组织与协同调节的机制 蛋白质在功能上与调节信号的整合相结合， 控制成骨细胞核中的基因表达， 破骨细胞核。项目3将寻求控制重塑的机制 骨特异性基因的染色质结构，使启动子元件 能够进行生理反应调节蛋白质相互作用。 体外细胞核结构基因表达相互关系的评价 将提供调节参数的生物学验证， 对于骨骼发育是必需的，并且可能在代谢骨中受损 疾病

DESCRIPTION OF THE OVERALL PROGRAM (provided by applicant): Our program continues to address the hypothesis that parameters of nuclear structure (chromatin organization and the assembly and activity of nuclear matrix associated subnuclear sites for transcription) contribute to gene expression that mediates the onset, progression and maintenance of bone cell phenotypic properties required for skeletal development and homeostasis. We are pursuing an integrated, team approach that combines molecular, cellular, biochemical and in vivo genetic analyses to define mechanisms by which subnuclear organization of nucleic acids and regulatory proteins facilitates integration of physiological signals to support competency for skeletal gene expression and bone development in vivo. Our program has: 1) demonstrated remodeling of chromatin and nucleosome organization that is linked to developmental induction and steroid hormone enhanced bone specific transcription; 2) identified bone specific activators and co regulators that integrate activities at skeletal gene promoter elements; 3) demonstrated that the Runx2 (Cbfal/AML3) factor is punctately localized at discrete nuclear matrix associated subnuclear sites; and 4) structurally and functionally characterized a unique intranuclear targeting signal that directs Runx2 to sites of transcription as a requirement for skeletal development to proceed in vivo. We will now examine mechanisms that support our fundamental observations that spatial and temporal organization and assembly of regulatory complexes within the nucleus are necessary for expression of genes that control bone formation. Project 1 will investigate mechanisms that direct bone specific transcription factors and coregulatory proteins to sites within the nucleus where developmental and steroid hormone responsive gene expression occurs. Project 2 will determine mechanisms by which the subnuclear organization of Runx2 with coregulatory proteins is functionally coupled to the integration of regulatory cues that control gene expression in the nucleus of osteoblasts and in the multiple nuclei of osteoclasts. Project 3 will pursue mechanisms that control remodeling of chromatin structure of bone specific genes to render promoter elements competent for physiologically responsive regulatory protein interactions. Evaluation of nuclear structure gene expression interrelationships in vitro and in vivo will provide biological validation of regulatory parameters that are obligatory for skeletal development and may be compromised in metabolic bone disease.