Insulin Mobilizes Peripheral Endosomal GLUT4 Compartments to the Plasma Membrane

Abstract

Insulin-stimulated GLUT4 redistribution to the plasma membrane (PM) plays essential roles in glucose homeostasis in mammals. Besides GLUT4 storage vesicles (GSVs) being released by Rab10 activation, endosomal GLUT4 compartments located at the cell periphery are also suggested to contribute to insulin-stimulated GLUT4 accumulation at the PM. Peripheral endosomal GLUT4 compartments are well suited to be studies with total internal reflection fluorescence (TIRF) microscopy. However, visualizing these GLUT4-positive structures after insulin is hindered by the fact that a large amount of GLUT4 is present on the PM after insulin stimulation, obscuring all intracellular GLUT4 compartments.In this study, a new GLUT4 probe, GLUT4-exoGFP, is developed. In combination with Bromophenol Blue (BPB), GLUT4-exoGFP can be used to specifically visualize intracellular GLUT4 compartments, without interference from GLUT4 at the PM. Employing this probe we studied the mobilization of GLUT4 from peripheral GLUT4 compartments. Our results indicate that Rab4A-positive endosomes serve as an important reservoir of GLUT4 molecules that are delivered to the PM after insulin stimulation.