SIGNAL PATHWAY--INTERSTIT COLLAGE TRANSCRIPT

信号通路--校际拼贴成绩单

• 批准号: 2683246
• 负责人: MATTHEW P VINCENTI
• 金额: $ 8.62万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-04-01 至 2002-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2683246
• 关键词: DNA binding protein; biological signal transduction; collagenase; enzyme activity; enzyme biosynthesis; fibroblasts; gel mobility shift assay; gene expression; genetic promoter element; immunoprecipitation; interleukin 1; interstitial; laboratory rabbit; mitogen activated protein kinase; molecular cloning; pathologic process; polymerase chain reaction; protein kinase; rheumatoid arthritis; site directed mutagenesis; southern blotting; synovial fluid; transcription factor; transfection; yeast two hybrid system

Interstitial collagenase is an enzyme produced by synovial fibroblasts which contributes to the pathophysiology of rheumatoid arthritis. In this disease, synovial fibroblasts are activated by inflammatory signals, such as interleukin-1 (IL-1 and produce inappropriately high levels of callagenase, contributing to the degradation of cartilage, ligament and bone. Despite the importance of collagenase gene activation in RA, the signaling pathways leading to increased collagenase transcrption have not been defined. Recent studies I have conducted demonstrate that in addition to IL-1 by v-src is a potent activator of collagenase transcription. Furthermore, IL-1 and v-src can work synergistically to activate the collagenase promoter, suggesting these stimuli share common signaling intermediates. To begin to define the pathway (s) leading to collagenase transcription, the transcription factors activated b v-src and interact with the proximal collagenase promoter will be characterized. Downstream mediators of v-src, such as Map kinases will be identified and the function of these downstream kinases will be assayed in synovial fibroblasts expressing v-src. The synergism between v-src and IL-1 may be due to a src-related kinase which is an intermediate in the the IL-1 dependent pathway. To explore this possibility, a dominant negative v-src willbe tested as a specific inhibito of IL-1 induced collagenase transcription. Alternatively, transcription factors which are activated by v-src and IL-1 may physically interact in the nucleus to cooperatively recruit RNA polymerase II to the collagenase promoter. Thus, potential protein-protein interactions between v-src and IL-1 induced transcription factors will be tested through use of the yeast two hybrid system. This approach will also be used to identify signal transducing proteins, such as Map kinases which interact with activate these transcription factors. This work will, for th first time, descrive the specific protein kinases and transcription factor which interact the signaling pathways involved in collagenase transcription will lead to a better understanding of the molecular mechanisms of connective tissue disease.

间质胶原酶是滑膜成纤维细胞产生的一种酶 这有助于类风湿性关节炎的病理生理学。在这个 当疾病发生时，滑膜成纤维细胞会被炎症信号激活，例如 作为白细胞介素-1 (IL-1) 并产生不适当高水平的 卡拉酶，导致软骨、韧带和软骨的降解 骨。 尽管胶原酶基因激活在 RA 中很重要， 导致胶原酶转录增加的信号通路尚未 被定义。 我最近进行的研究表明，在 v-src 除了 IL-1 之外，也是胶原酶的有效激活剂 转录。 此外，IL-1 和 v-src 可以协同作用 激活胶原酶启动子，表明这些刺激共享 常见的信号中间体。 开始定义路径 导致胶原酶转录，转录因子被激活 v-src 并与近端胶原酶启动子相互作用 特点。 v-src 的下游介质，例如 Map 激酶，将 被鉴定并且这些下游激酶的功能将被确定 在表达 v-src 的滑膜成纤维细胞中进行测定。 之间的协同作用 v-src 和 IL-1 可能是由 src 相关激酶引起的，该激酶是 IL-1依赖性途径的中间体。 为了探索这个 可能性，显性失活 v-src 将作为特定抑制剂进行测试 IL-1 诱导胶原酶转录。 或者，转录 由 v-src 和 IL-1 激活的因子可能在物理上相互作用 细胞核协同招募RNA聚合酶II 胶原酶启动子。 因此，潜在的蛋白质-蛋白质相互作用 将测试 v-src 和 IL-1 诱导转录因子之间的关系 通过使用酵母两种杂交系统。 这一做法也将 用于识别信号转导蛋白，例如 Map 激酶 相互作用并激活这些转录因子。 这项工作将为 第一次描述具体的蛋白激酶和转录因子 与胶原酶相关的信号通路相互作用 转录将有助于更好地理解分子 结缔组织疾病的机制。