Insulin-regulated release from the endosomal recycling compartment is regulated by budding of specialized vesicles

In several cell types, specific membrane proteins are retained intracellula rly and rapidly redistributed to the surface in response to stimulation. In fat and muscle, the GLUT4 glucose transporter is dynamically retained beca use it is rapidly internalized and slowly recycled to the plasma membrane. Insulin increases the recycling of GLUT4, resulting in a net translocation to the surface. We have shown that fibroblasts also have an insulin-regulat ed recycling mechanism. Here we show that GLUT4 is retained within the tran sferrin receptor-containing general endosomal recycling compartment in Chin ese hamster ovary (CHO) cells rather than being segregated to a specialized , GLUT4-recycling compartment. With the use of total internal reflection mi croscopy, we demonstrate that the TR and GLUT4 are transported from the per icentriolar recycling compartment in separate vesicles. These data provide the first functional evidence for the formation of distinct classes of vesi cles from the recycling compartment. We propose that GLUT4 is dynamically r etained within the endosomal recycling compartment in CHO cells because it is concentrated in vesicles that form more slowly than those that transport TR. In 3T3-L1 adipocytes, cells that naturally express GLUT4, we find that GLUT4 is partially segregated to a separate compartment that is inaccessib le to the TR. We present a model for the formation of this specialized comp artment in fat cells, based on the general mechanism described in CHO cells , which may explain the increased retention of GLUT4 and its insulin-induce d translocation in fat cells.