Aledo J C, Darakhshan F, Hundal H S
Department of Anatomy and Physiology, University of Dundee, UK.
Biochem Biophys Res Commun. 1995 Oct 4;215(1):321-8. doi: 10.1006/bbrc.1995.2469.
The role of Rab4, a small molecular weight GTP binding protein implicated in endosomal/plasma membrane (PM) recycling, in the translocation of the GLUT4 transporter in rat skeletal muscle was studied. Muscle membranes, prepared by subcellular fractionation of control and insulin treated rat skeletal muscle, were subjected to SDS/PAGE and immunoblot analyses. Insulin treatment caused an increase in GLUT4 in a plasma membrane (PM) enriched fraction from an intracellular membrane (IM) fraction. Immunoprecipitation of GLUT4 vesicles from the IM compartment revealed that Rab4 could be coprecipitated. Importantly, however, and unlike in adipocytes, immunoisolated GLUT4 vesicles from rat skeletal muscle contained no detectable immunoreactivity towards the transferrin receptor, suggesting that Rab4 was present in a GLUT4 IM pool that was not characteristic of early endosomes. The involvement of Rab4 in GLUT4 translocation was further supported by the finding that insulin treatment resulted in a significant (43%) reduction in Rab4 in the IM compartment. Our results suggest (i) that insulin induces the loss of both GLUT4 and Rab4 from the same IM compartment, (ii) that Rab4 may be involved in GLUT4 translocation based on its coprecipitation with the transporter from the insulin-sensitive pool and (iii) that Rab4 can be localized to intracellular membranes which appear not to be of early endosome origin.
研究了小分子质量GTP结合蛋白Rab4在内体/质膜（PM）循环中所起的作用，及其在大鼠骨骼肌中葡萄糖转运蛋白4（GLUT4）转运过程中的作用。通过对对照和胰岛素处理的大鼠骨骼肌进行亚细胞分级分离制备肌膜，然后进行SDS/PAGE和免疫印迹分析。胰岛素处理使来自细胞内膜（IM）部分的富含质膜（PM）的部分中GLUT4增加。从IM区室免疫沉淀GLUT4囊泡发现，Rab4可以共沉淀。然而，重要的是，与脂肪细胞不同，从大鼠骨骼肌免疫分离的GLUT4囊泡对转铁蛋白受体没有可检测到的免疫反应性，这表明Rab4存在于GLUT4的IM池中，而该池不是早期内体的特征。胰岛素处理导致IM区室中Rab4显著减少（43%），这一发现进一步支持了Rab4参与GLUT4转运。我们的结果表明：（i）胰岛素诱导同一IM区室中GLUT4和Rab4的丢失；（ii）基于Rab4与来自胰岛素敏感池的转运蛋白共沉淀，Rab4可能参与GLUT4转运；（iii）Rab4可以定位于似乎不是早期内体起源的细胞内膜。
