Transcriptional Regulation of Chondrogenesis

软骨形成的转录调控

• 批准号: 7216807
• 负责人: HIROSHI ASAHARA
• 金额: $ 32.39万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-07-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7216807
• 关键词: Acetylation; Arthritis; Biological Assay; Biomechanics; CREB-binding protein; CREB1 gene; Cartilage; Cells; Chimera organism; Chondrocytes; Chondrogenesis; Chromatin; Collagen Type II; Complex; Data; Deacetylation; Development; Dominant-Negative Mutation; Embryo; Enzymes; Extracellular Matrix; Gene Expression; Genes; Genetic Transcription; Genital system; Gluconeogenesis; Goals; HMG Domain; In Situ Hybridization; In Vitro; Joint repair; Joints; Limb Development; Mesenchymal Stem Cells; Molecular; Mus; PPAR gamma; Pathway interactions; Phenotype; Phosphorylation; Play; Process; Protein Overexpression; Proteins; Regulation; Research; Research Personnel; Role; Site; Skeletal Development; Staging; Stem cells; Tissue-Specific Gene Expression; Tissues; Transcriptional Regulation; arthropathies; articular cartilage; base; cartilage development; day; embryonic stem cell; glycogenesis; human CREBBP protein; lipid biosynthesis; novel strategies; programs; promoter; reconstitution; transcription factor

DESCRIPTION (provided by applicant): Chrondrogenesis is a multi-step pathway during which multipotential mesenchymals stem cells differentiate into chondrocytes to form articular cartilage. This process is not only essential for cartilage development but also during joint repair. Research on molecular mechanisms of chrondrogenesis will advance our understanding of skeletal development and has the potential to identify new approaches to the treatment of joint diseases. The Sox9 high-mobility group (HMG) domain transcription factor is a key molecule in chondrocyte differentiation. Expression of Sox9 almost parallels that of Col2a1, which encodes cartilage-specific type II collagen during chondrogenesis. In the genital ridge, however, Col2a1 is not expressed, despite high level of endogenous Sox9 expression, suggesting the existence of molecular partner(s) of Sox9 that are required for cartilage-specific Cola1 gene expression. In preliminary studies, we identified PGC-1 as a Sox9 transcriptional co-activator. PGC-1 was previously known as PPAR gamma co-activator and plays a role in adipogenesis and glycogenesis; however, its role in chondrogensis has been unexplored. Whole mount in situ hybridization of day 10-12 mice embryos revealed PGC-1 gene expression only in cells at chondrogenic sites. PGC-1 interacts with Sox9 to enhance Col2a1 promoter activity and promotes chondrogenesis. Furthermore, Sox9 activity on the Col2a1 is tightly regulated by acetylation and deacetylation molecules. CBP (CREB binding protein), one of the chromatin acetylation enzymes, was also shown to interact with Sox9/PGC-1 complex and is involved in this mechanism. These preliminary data provide the basis for our hypothesis that PGC-1 functions as a tissue and developmental stage specific co-activator of Sox9 during chondrogenesis in a mechanism that involves specific chromatin regulation. Toward this end, we propose following Specific Aims: 1. Determine the role of PGC-1 as a co-activator of Sox9 during chondrogenesis and chondrocyte-specific gene expression. 2. Determine the role of Sox9/PGC-1 complex associating HAT activity and subsequent chromatin regulation for tissue specific gene expression and chondrocyte differentiation. 3. Examine mechanisms by which Sox9/PGC-1 complex dependent gene expression is regulated by HDACs and related co-repressor complex during chondrogenesis. 4. Analyze chromatin dependent Sox9/PGC-1 complex transcription mechanism by using in vitro reconstitution of chromatin and transcription assay of the Col2al promoter.

描述（由申请人提供）：软骨形成是多能间充质干细胞分化成软骨细胞以形成关节软骨的多步骤途径。这个过程不仅对于软骨发育至关重要，而且在关节修复过程中也至关重要。对软骨形成分子机制的研究将增进我们对骨骼发育的理解，并有可能找到治疗关节疾病的新方法。 Sox9 高迁移率基团 (HMG) 结构域转录因子是软骨细胞分化的关键分子。 Sox9 的表达几乎与 Col2a1 的表达相似，Col2a1 在软骨形成过程中编码软骨特异性 II 型胶原。然而，在生殖嵴中，尽管内源性 Sox9 表达水平很高，但 Col2a1 并不表达，这表明存在软骨特异性 Cola1 基因表达所需的 Sox9 分子伴侣。在初步研究中，我们确定 PGC-1 是 Sox9 转录共激活因子。 PGC-1 以前被称为 PPAR γ 共激活剂，在脂肪生成和糖生成中发挥作用；然而，其在软骨形成中的作用尚未被探索。第 10-12 天小鼠胚胎的整体原位杂交显示 PGC-1 基因仅在软骨形成位点的细胞中表达。 PGC-1 与 Sox9 相互作用，增强 Col2a1 启动子活性并促进软骨形成。此外，Col2a1 上的 Sox9 活性受到乙酰化和脱乙酰化分子的严格调控。 CBP（CREB ​​结合蛋白）是染色质乙酰化酶之一，也被证明与 Sox9/PGC-1 复合物相互作用，并参与了这一机制。这些初步数据为我们的假设提供了基础，即 PGC-1 在软骨形成过程中作为 Sox9 的组织和发育阶段特异性共激活剂，其机制涉及特定染色质调节。为此，我们提出以下具体目标： 1. 确定 PGC-1 在软骨形成和软骨细胞特异性基因表达过程中作为 Sox9 共激活剂的作用。 2. 确定 Sox9/PGC-1 复合物在 HAT 活性和随后的染色质调节中对组织特异性基因表达和软骨细胞分化的作用。 3. 检查软骨形成过程中 HDAC 和相关共阻遏物复合物调节 Sox9/PGC-1 复合物依赖性基因表达的机制。 4.通过染色质体外重建和Col2al启动子转录测定分析染色质依赖性Sox9/PGC-1复合体转录机制。