The mechanisms of insulin-regulated exocytosis of the GLUT4-containing vesicles

含有 GLUT4 的囊泡的胰岛素调节胞吐作用的机制

• 批准号: 11671107
• 负责人: SHIBATA Hiroshi
• 金额: $ 2.3万
• 依托单位: Gunma University, Institute For Molecular and Cellular Regulation
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671107/
• 关键词: insulin; glucose transporter; GTP-binding protein; Rab4; syntaxin4; SNARE

In the present study, we investigated the interaction of Rab4 with syntaxin4, a t-SNARE protein implicated in the insulin-induced exocytic fusion of the GLUT4-containing vesicle with the plasma membrane. Rab4 and syntaxin4 were coimmunoprecipitated from the lysates of rat adipocytes. The interaction of the two proteins were attenuated by pretreatment of the cells with insulin but enhanced with GTPgS.A GTPase deficient mutant of Rab4, but not a GTP-binding defective mutant was bound to syntaxin 4, suggesting that the interaction between the two proteins were regulated by the guanine-nucleotide-binding state of Rab4. In addition, we found that the presence of munc-18c, a negative regulator of the SNARE comple formation, displaced Rab4 from syntaxin 4, indicating that the dissociation of munc-18c from syntaxin4 is required for Rab4 to bind to syntaxin 4.We also clarified that the actin filaments play a critical role in exocytotic recruitment but not in endocytosis of GLUT4 in isolated rat adipocytes.

在本研究中，我们研究了 Rab4 与 Syntaxin4 的相互作用，syntaxin4 是一种 t-SNARE 蛋白，参与胰岛素诱导的含有 GLUT4 的囊泡与质膜的胞吐融合。 Rab4 和 Syntaxin4 从大鼠脂肪细胞裂解物中进行免疫共沉淀。用胰岛素预处理细胞会减弱两种蛋白质的相互作用，但用 GTPgS 会增强。Rab4 的 GTP 酶缺陷突变体（而非 GTP 结合缺陷突变体）与突触融合蛋白 4 结合，表明这两种蛋白质之间的相互作用是受 Rab4 的鸟嘌呤核苷酸结合状态调节。此外，我们发现munc-18c（SNARE复合体形成的负调节因子）的存在将Rab4从syntaxin 4中取代，表明munc-18c从syntaxin4解离是Rab4与syntaxin 4结合所必需的。阐明肌动蛋白丝在分离的大鼠脂肪细胞中的胞吐募集中发挥关键作用，但在 GLUT4 的内吞作用中不起关键作用。

项目成果

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