Abstract GLUT4 function is dependent on cellular sorting and trafficking mechanisms that result in its intracellular sequestration in the basal state, and in its rapid redistribution to the plasma membrane in response to insulin stimulation. Structure–function studies utilizing chimeras of GLUT1 and GLUT4 have demonstrated that the 30-amino-acid COOH-terminus of GLUT4 is necessary and sufficient for its sequestration in intracellular membrane compartments in CHO, COS, L6 muscle cells, and 3T3-L1 adipocytes. When expressed in CHO, COS or other cells that do not contain endogenous GLUT4, intracellular retention is entirely dependent on a double leucine motif contained within the GLUT4 COOH-terminus, which directs both rapid endocytosis and intracellular retention of expressed transporters. However, in 3T3-L1 adipocytes, and additional motif or motifs in the 30-amino-acid COOH-terminus of GLUT4 may also operate to confer intracellular sequestration and insulin-mediated redistribution. Kinetic analyses of GLUT4 internalization and recycling indicate that insulin enhances the exocytic rate constant of GLUT4, and causes a small decrease in its internalization rate. Recent data suggest that phosphatidylinositol 3-kinases and their 3 phosphoinositide products may be required for GLUT4 exocytosis, perhaps through the regulation of components involved in membrane budding, fusion or movement. New findings support the hypothesis that insulin causes targeting of complexes containing insulin receptor substrate-1 and phosphatidylinositol 3-kinase to the specialized GLUT4 sequestration compartment, providing a possible mechanism for insulin action.