Diabetes and Obesity Program, Garvan Institute of Medical Research, Sydney, 2010, Australia; St Vincent's Clinical School, University of New South Wales, Sydney, 2052, Australia.
Traffic. 2012 Oct;13(10):1429-41. doi: 10.1111/j.1600-0854.2012.01397.x. Epub 2012 Jul 29.
Insulin stimulates glucose transport in adipocytes by triggering translocation of GLUT4 glucose transporters to the plasma membrane (PM) and several Rabs including Rab10 have been implicated in this process. To delineate the molecular regulation of this pathway, we conducted a TBC/RabGAP overexpression screen in adipocytes. This identified TBC1D13 as a potent inhibitor of insulin-stimulated GLUT4 translocation without affecting other trafficking pathways. To determine the potential Rab substrate for TBC1D13 we conducted a yeast two-hybrid screen and found that the GTP bound forms of Rabs 1 and 10 specifically interacted with TBC1D13 but not with eight other TBC proteins. Surprisingly, a comprehensive in vitro screen for TBC1D13 GAP activity revealed Rab35 but not Rab10 as a specific substrate. TBC1D13 also displayed in vivo GAP activity towards Rab35. Overexpression of constitutively active Rab35 but not constitutively active Rab10 reversed the block in insulin-stimulated GLUT4 translocation observed with TBC1D13 overexpression. These studies implicate an important role for Rab35 in insulin-stimulated GLUT4 translocation in adipocytes.
胰岛素通过触发 GLUT4 葡萄糖转运蛋白向质膜（PM）易位来刺激脂肪细胞中的葡萄糖转运，几种 Rab 蛋白，包括 Rab10，已被牵连到这个过程中。为了描绘这个途径的分子调控，我们在脂肪细胞中进行了 TBC/RabGAP 过表达筛选。这鉴定出 TBC1D13 是胰岛素刺激的 GLUT4 易位的有效抑制剂，而不影响其他运输途径。为了确定 TBC1D13 的潜在 Rab 底物，我们进行了酵母双杂交筛选，发现 Rab1 和 Rab10 的 GTP 结合形式与 TBC1D13 特异性相互作用，但与其他八个 TBC 蛋白没有相互作用。令人惊讶的是，针对 TBC1D13 的全面体外 GAP 活性筛选显示 Rab35 而不是 Rab10 是特定的底物。TBC1D13 还在体内对 Rab35 显示出 GAP 活性。过表达组成性激活的 Rab35，但不是组成性激活的 Rab10，逆转了 TBC1D13 过表达观察到的胰岛素刺激的 GLUT4 易位的阻断。这些研究表明 Rab35 在胰岛素刺激的脂肪细胞中 GLUT4 易位中起着重要作用。
