Evidence that insulin causes translocation of glucose transport activity to the plasma membrane from an intracellular storage site.

Abstract

The glucose transport activity of fat cells was assayed in a cell-free system. The activity was solubilized and incorporated into egg-lecithin liposomes. The carrier-mediated glucose transport activity was estimated by subtracting the cytochalasin B-insensitive component from the total glucose uptake activity of the modified liposomes. When a crude microsomal preparation from fat cells was fractionated by sucrose density gradient centrifugation, two transport activities (peaks A and B) were separated. Peak A coincided with the peak of 5'-nucleotidase, a marker of the plasma membrane. Peak B appeared to coincide with the peak of UDPGal:N-acetylglucosamine galactosyltransferase, a marker of the Golgi apparatus. Peak A was considerably smaller than peak B under basal conditions. When cells were exposed to 1 nM insulin for 5 min before homogenization, the height of peak A increased whereas that of peak B decreased. Insulin had no significant effect on the galactosyltransferase activity. The Km values of glucose transport facilitated by the activities in peaks A and B were both approximately 10-15 mM. These results imply that insulin facilitates translocation of the transport activity from an intracellular storage site to the plasma membrane.