Insulin regulation of protein traffic in rat adipose cells

Rat adipocytes were biotinylated with cell-impermeable reagents, sulfo-N-hy droxysuccinimide-biotin and sulfo-N-hydroxysuccinimide-S-S-biotin in the ab sence and presence of insulin. Biotinylated and nonbiotinylated populations of the insulin-like growth factor-II/mannose B-phosphate receptor, the tra nsferrin receptor, and insulin-responsive aminopeptidase were separated by adsorption to streptavidin-agarose to determine the percentage of the bioti nylated protein molecules versus their total amount in different subcellula r compartments. Results indicate that adipose cells possess at least two di stinct cell surface recycling pathways for insulin-like growth factor-II/ma nnose 6-phosphate receptor (MPR) and transferrin receptor (TfR): one which is mediated by glucose transporter isoform 4(Glu4)-vesicles and another tha t bypasses this compartment, Under basal conditions, the first pathway is n ot active, and cell surface recycling of TfR and, to a lesser extent, MPR p roceeds via the second pathway. Insulin dramatically stimulates recycling t hrough the first pathway and has little effect on the second. Within the Glut4-containing compartment, insulin has profoundly different e ffects on intracellular trafficking of insulin-responsive aminopeptidase on one hand and MPR and TfR on the other. After insulin administration, insul in-responsive aminopeptidase is redistributed from Glut4-containing vesicle s to the plasma membrane and stays there for at least 30 min with minimal d etectable internalization and recycling, whereas MPR and TfR rapidly shuttl e between Glut4 vesicles and the plasma membrane in such a way that after 3 0 min of insulin treatment, virtually every receptor molecule in this compa rtment completes at least one trafficking cycle to the cell surface. Thus, different recycling proteins, which compose Glut4-containing vesicles, are internalized into this compartment at their own distinctive rates.