Rheb调节Glut4胞膜表达分子机制研究

项目名称： Rheb调节Glut4胞膜表达分子机制研究

项目编号： No.30800388

项目类型： 青年科学基金项目

立项/批准年度： 2009

项目学科： 金属学与金属工艺

项目作者： 杜晓霞

作者单位： 四川大学

项目金额： 20万元

中文摘要： 葡萄糖转运蛋白 4（Glut4）是脂肪和骨骼肌细胞协助葡萄糖转运的主要蛋白质，胰岛素刺激下从胞浆转位至胞膜，Glut4功能障碍是胰岛素抵抗和糖尿病的重要机制。胰岛素信号通路是调节脂肪细胞中Glut4 膜表达的主要途径，而Rheb在该信号通路中起关键作用，Rheb敲除小鼠中AKT激酶活性增加，而AKT活性在 Glut4 转位及葡萄糖摄取中起重要作用。本项目着重研究 Rheb是否通过AKT对Glut4膜表达起调节作用。 通过本研究，我们成功诱导3T3-L1前体细胞分化为脂肪细胞，胰岛素刺激后Glut4在转染CHO-K1细胞及诱导脂肪细胞中发生易位。但体外实验表明在诱导脂肪细胞中过表达Rheb1对胰岛素刺激下的Glut4膜表达无抑制作用。为了进一步研究体内Rheb1对Glut4膜表达有无抑制作用，我们还制备和鉴定了过表达野生型和活性形式的Rheb1转基因小鼠，体内实验表明过表达野生型和活性形式Rheb1均会抑制AKT激酶活性。但由于转基因小鼠数量较少，因此部分实验尚未能开展，这也将是下一步的研究目标。通过本研究将在Ⅱ#22411;糖尿病因学研究、新的药物靶点、Ⅱ#22411;糖尿病动物模型等方面提供重要依据。

中文关键词： Glut4；胰岛素信号通路；Rheb1；AKT；基因敲入；

英文摘要： Glucose transport protein 4 (Glucose transporter-4, Glut4) is the main protein of Glucose transport in adipocytes and skeletal muscle cells, translocation from cytoplasm to the cell membranes at insulin stimulation, Glut4 function obstacle is a cause of insulin resistance and is the important mechanism of diabetes. Insulin signaling is a main pathway adjusting Glut4 membrane expression in the adipocytes, and Rheb plays a key role in insulin signal transduction, Rheb knockout mice AKT kinase activity increased significantly, and AKT activity in the Glut4 inversion to cell membrane and glucose absorb plays an important role. This project focuses on the regulating action of Rheb in Glut4 membrane expression and translocation mediated by AKT. In this study, we successfully induction the 3T3-L1 precursor cells differentiation become adipocytes. After insulin stimulation, HA-Glut4-GFP fusion protein translocated to membrane in transfected CHO-K1 cells and in adipocytes induced differentiation from 3T3-L1 cells. But in vitro studies show that, overexpression of Rheb1 in adipocytes induced differentiation from 3T3-L1 cells have no inhibition to the Glut4 membrane expression after insulin stimulating, this is not consistent with the original concept. In order to further research if Rheb1 inhibit in Glut4 the Glut4 membrane expression in vivo, we also prepared and identified two kinds of transgenic mice which can overexpress wild type or active form (S16H) Rheb1.In vivo experiments show that Whether the wild type or the active Rheb1 can inhibit AKT kinase activity, that is consistent with the in vitro experimens. But because the transgenic mice now get a smaller number of, some experiments have not been able to develop, that will also be the next research contents. Through this research will be deepen understanding of pathogenesis of Ⅱype of diabetes, found new drug targets, and establishedⅡtype of diabetes animal model.

英文关键词： Glut4; Insulin signaling pathways; Rheb1; AKT; knockin;