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: 6100458
• 负责人: R RAGHOW
• 金额: $ 15.73万
• 依托单位: UNIVERSITY OF TENNESSEE HEALTH SCI CTR
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-09-01 至 1999-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6100458
• 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糖尿病的一个标志。 纤维化疾病，其通常与类风湿性关节炎相关。 这项建议的长远目标，是制订一套全面的 精确描述DNA-蛋白质相互作用的概念框架 其是人前α（I）胶原蛋白转录调节的基础 基因 更直接的目标将是顺式- 包含Proalpha 1（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 胶原启动子/增强子-报告基因构建体将被引入到 间充质和非间充质来源的培养细胞通过瞬时或 稳定的转染方案，并且还掺入到 转基因小鼠;将评估报告基因的表达， 在体外和体内模型之间进行比较。 假定的反式作用 参与组成性和调节性转录的因子 Proalpha 1（I）基因的调节将被广泛表征， 编码这些因子的cDNA将被分子克隆。 机制 顺式元件和反式作用因子之间的相互作用 确保人Proalpha 1（I）的适当转录调控 将通过靶启动子/增强子的共转染来研究基因， 报告构建体和表达载体含有推定的反式- 作用因素。 此外，将开发无细胞系统， 研究决定蛋白质与DNA相互作用的详细信息 Proalpha 1（I）胶原转录。 这些研究将揭开 分子间的相互作用， 人前α 1（I）胶原转录调控特异性 基因表达。 详细了解的机制 I型胶原基因的转录控制将使我们能够设计 治疗干预，以减少关节和关节外 与类风湿性关节炎相关的纤维化表现。