TRANSCRIPTIONAL REGULATION OF TYPE I COLLAGEN GENE EXPRESSION

I型胶原蛋白基因表达的转录调控

• 批准号: 3728072
• 负责人: R RAGHOW
• 金额: --
• 依托单位: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3728072
• 关键词: DNA footprinting; collagen; gel mobility shift assay; gene expression; gene interaction; genetic enhancer element; genetic manipulation; genetic promoter element; genetic regulatory element; genetic transcription; genetically modified animals; intermolecular interaction; messenger RNA; molecular cloning; procollagen; rheumatoid arthritis; southern blotting; transcription factor; transfection

Aberrant regulation of type I collagen gene expression is a hallmark of the fibrotic disorders which are often associated with rheumatoid arthritis. The long term objective of this proposal is to formulate a comprehensive conceptual framework to precisely delineate the DNA-protein interactions which underlie transcriptional regulation of the human Proalpha(I) collagen gene. The more immediate goal will be the molecular dissection of the cis- regulatory sequence motifs that comprise the Proalpha1(I) promoter/enhancer and the trans-acting factors which interact with these sequences. Chimeric collagen promoter/enhancer-reporter gene constructs will be introduced into cultured cells of mesenchymal and non-mesenchymal origin by transient or stable transfection protocols, and also incorporated into the genome of transgenic mice; the expression of the reporter gene will be assessed and compared between in vitro and in vivo models. Putative trans-acting factors involved in the constitutive and modulatory transcriptional regulation of the Proalpha1(I) gene will be extensively characterized and cDNA encoding these factors will be molecularly cloned. Mechanisms by which interactions between the cis-elements and the trans-acting factors ensure appropriate transcriptional regulation of the human Proalpha1(I) gene will be studied by cotransfection of the target promoter/enhancer- reporter constructs and expression vectors containing the putative trans- acting factor(s). Additionally, cell-free systems will be developed to investigate the detailed protein-DNA interactions which determine Proalpha1(I) collagen transcription. These studies will unravel the molecular interactions that orchestrate appropriate tissue and cell specificity of transcriptional regulation of human Proalpha1(I) collagen gene expression. A detailed understanding of the mechanisms of transcriptional control of type I collagen genes will enable us to devise therapeutic interventions to reduce the articular and extraarticular fibrotic manifestations associated with rheumatoid arthritis.

I型胶原基因表达的异常调节是 常与类风湿性关节炎相关的纤维性疾病。 这项建议的长期目标是制定一项全面的 精确描述DNA-蛋白质相互作用的概念框架 它是人类ProAlpha(I)胶原转录调控的基础 吉恩。更直接的目标将是对顺式基因的分子解剖- 组成Proalpha1(I)启动子/增强子的调控序列基序 以及与这些序列相互作用的反式作用因子。嵌合体 胶原蛋白启动子/增强子报告基因构建体将被引入 间质来源和非间充质来源的培养细胞 稳定的转导方案，也被整合到 转基因小鼠；报告基因的表达将被评估和 比较了体外模型和体内模型。推定的反动行为 构成性和调节性转录中涉及的因素 将对Proalpha1(I)基因的调节进行广泛的表征和 编码这些因子的cdna将被分子克隆。机制，由 顺式元件和反式作用因子之间的哪些相互作用 确保对人类Proalpha1进行适当的转录调控(I) 通过将靶启动子/增强子共转染来研究基因。 含推定反式-β-内酰胺酶基因的报告基因构建和表达载体 代理因素(S)。此外，无牢房系统将被开发为 研究决定蛋白质-DNA相互作用的详细信息 Proalpha1(I)胶原蛋白转录。这些研究将揭开 协调适当组织和细胞的分子相互作用 人Proalpha1(I)胶原转录调控的特异性 基因表达。详细了解其作用机制。 I型胶原基因的转录控制将使我们能够设计 关节及关节外复位的治疗性干预 与类风湿性关节炎相关的纤维化表现。