Elmo1介导TGF-β1活化Rac1的机制及其在肾间质纤维化中的作用研究

项目名称： Elmo1介导TGF-β1活化Rac1的机制及其在肾间质纤维化中的作用研究

项目编号： No.81200503

项目类型： 青年科学基金项目

立项/批准年度： 2013

项目学科： 医学二处

项目作者： 朱风新

作者单位： 南方医科大学

项目金额： 23万元

中文摘要： 肾间质纤维化（TIF）是各种慢性肾脏病发展至终末期肾病的共同途径，细胞外基质（ECM）沉积是其重要特征。TGF-β1活化Rac1可促进小管上皮细胞合成ECM，但TGF-β1活化Rac1的机制尚不明。申请者前期工作发现TGF-β1以时间依赖模式上调具活化Rac1功能的Elmo1的表达，过表达Elmo1增强TGF-β1对Rac1的活化及ECM的合成，提示TGF-β1-Elmo1-Rac1途径参与调控ECM沉积。为证实上述假说，本项目拟：1）探讨TGF-β1调控Elmo1表达的机制；2）利用Elmo1或Rac1 siRNA，证实Elmo1-Rac1途径参与TGF-β1诱导的ECM的合成；3）构建Elmo1磷酸化位点的突变体，阐明TGF-β1通过促进Elmo1的磷酸化解除其自身抑制，从而活化Rac1。4）利用Elmo1敲基因鼠，进一步证实Elmo1在TIF中的作用，从而为防治慢性肾脏病提供新靶标。

中文关键词： Elmo1；Rac1；TGF-β1；细胞外基质；肾间质纤维化

英文摘要： Tubulointerstitial fibrosis is the common final outcome of almost all progressive chronic kidney diseases. And excessive deposition of extracellularmatrix is the most striking and name-lending feature of tubulointerstitial fibrosis. In the human renal tubular epithelial cell line HKC, Rac1 activity is required for TGF-β1-induced ECM production. But how TGF-β1activates Rac1 is still unclear. Our preliminary study suggests that in NRK52E cells TGF-β1 not only induces higher Rac1 activity but also stimulates the expression of Elmo1. Overexpression of wild-type Elmo1 promotes TGF-β1-stimulated ECM synthesis and Rac1 GTP-loading. These observations suggest that TGF-β1-Elmo1-Rac1 pathway was involved in the regulation of ECM deposition. In order to prove this hypothesis, we unmask the manner on which TGF-β1 regulates the expression of Elmo1. Elmo1 depletion by RNA interference impairs TGF-β1 stimulation of Rac1 activation and ECM deposition. At basal levels, Elmo1 is autoinhibited due to intramolecular EID/EAD interactions. Phosphorylation of Elmo1 could disrupt the autoinhibitory interaction. Tyr18 and Tyr 216, which locate in the EID domain, have been identified as the phosphorylation sites. Mutant forms of Elmo1 lacking these sites were defective in their ability to promote Rac1 activity. Overexpression of these mu

英文关键词： Elmo1；Rac1；TGF-β1；ECM；TIF