Trafficking kinetics of the insulin-regulated membrane aminopeptidase in 3T3-L1 adipocytes

In fat and muscle cells insulin causes the marked translocation of the glucose transporter GLUT4 from its intracellular location to the plasma membrane, We and others have discovered an insulin-regulated membrane aminopeptidase (designated IRAP) that colocalizes with intracellular GLUT4 and also translocates markedly in response to insulin, This study describes the trafficking kinetics of IRAP in 3T3-L1 adipocytes. By means of a surface biotinylation method, the half-time for the increase in IRAP at the plasma membrane in response to insulin was found to be 2 min. The increase was completely blocked by the phosphatidylinositol 3-kinase inhibitor, wortmannin. In insulin-treated cells, biotinylated IRAP, initially at the plasma membrane, equilibrated with the intracellular pool with a half-time of 2 min, Thus, IRAP continuously recycles, Finally, vesicles isolated from the intracellular membranes with antibodies against IRAP and GLUT4 showed the same protein composition. In conjunction with results in the literature, these findings indicate that IRAP and GLUT4 traffic through the same intracellular compartments. (C) 1997 Academic Press.