RABBIT COLLAGENASE GENE REGULATION BY IL-1

IL-1 对兔胶原酶基因的调控

• 批准号: 2651800
• 负责人: ULRIKE BENBOW
• 金额: $ 2.86万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-12-11 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2651800
• 关键词: collagenase; crosslink; fibroblasts; gene expression; genetic regulation; interleukin 1; laboratory rabbit; synovial membrane; tissue /cell culture

The long term goal of this research is to gain a detailed understanding of the molecular mechanisms that regulate collagenase (matrix metalloproteinase 1, MMP-1) expression in response to IL- 1. IL-1 is a potent physiological inducer of collagenase mRNA, and under pathological conditions collagenase, the only enzyme capable to degrade interstitial collagens at neutral pH, is instrumental in cartilage destruction in rheumatoid arthritis. Because of the central role IL-1 plays in the pathophysiology of connective tissue breakdown it is an important area of investigation. An understanding of the transcriptional and post-transcriptional mechanisms may influence the development of therapies designed to abrogate IL-1 induced collagenase gene expression. Therefore, the transcriptional and post- transcriptional mechanisms that regulate collagenase gene expression in response to IL-1 will be investigated using a rabbit model of synovial fibroblast cultures. Both experimentally and on the nucleotide sequence level this model mimics primary cultures of human cells from patients with rheumatoid arthritis. IL-1 responsive elements in the rabbit collagenase gene will be identified by transient transfection reporter constructs and deletional/mutational analysis. The dependency of these fragments on sequences in the proximal promoter, including AP-1 sites will be investigated. Specific DNA/protein interactions occurring on these elements will be investigated using electrophoretic mobility shift analysis, antibodies for specific transcription factors and UV crosslinking. RNA/protein interaction occurring at the 3' untranslated region of the collagenase mRNA will be determined by the ability of native or mutated AUUUA sequences to mediate binding of proteins from IL-1 treated and untreated cells.

这项研究的长期目标是获得详细的 了解调节胶原酶的分子机制 （基质金属蛋白酶 1、MMP-1）响应 IL-1 的表达。 IL-1 是胶原酶 mRNA 的有效生理诱导剂，并且在 病理条件胶原酶，唯一能够 在中性 pH 值下降解间质胶原蛋白，有助于 类风湿性关节炎中的软骨破坏。因为中央 IL-1 在结缔组织破坏的病理生理学中的作用 这是一个重要的研究领域。的理解 转录和转录后机制可能会影响 开发旨在消除 IL-1 诱导的疗法 胶原酶基因表达。因此，转录和后 调节胶原酶基因表达的转录机制 将使用兔模型研究对 IL-1 的反应 滑膜成纤维细胞培养物。无论是在实验上还是在 该模型模拟人类原代培养物的核苷酸序列水平 来自类风湿性关节炎患者的细胞。 IL-1 反应 兔胶原酶基因中的元件将通过以下方式鉴定 瞬时转染报告基因构建和缺失/突变 分析。这些片段对序列的依赖性 将研究近端启动子，包括 AP-1 位点。 这些元件上发生的特定 DNA/蛋白质相互作用将是 使用电泳迁移率变动分析进行研究， 针对特定转录因子和紫外线交联的抗体。 RNA/蛋白质相互作用发生在 3' 非翻译区 胶原酶 mRNA 将由天然或 突变的 AUUUA 序列介导 IL-1 蛋白的结合 处理和未处理的细胞。