INSULIN INCREASES THE ASSOCIATION OF AKT-2 WITH GLUT4-CONTAINING VESICLES

Expression of a constitutively active, membrane-associated Akt-1 (PKB alpha) construct in 3T3L1 adipocytes was shown to induce glucose uptak e in the absence of insulin by stimulating Glut4 translocation to the plasma membrane (Kohn, A.D., Summers, S.A., Birnbaum, M.J., and Roth, R.A. (1996) J. Biol. Chem. 271, 31372-31378). However, in rat fat cell the vast majority of Akt-1 is cytosolic and shows no re-distribution to the plasma membrane in response to insulin. On the other hand, litt le work has been done with other Akt family members such as Akt-2 (PKB beta) or Akt-3 (PKB gamma). In this report, an analysis of the subcel lular distribution of Akt-2 in rat adipocytes shows that Akt-2 is pres ent in significant amounts in various membrane compartments, as well a s in the cytosol, and the former include the light microsomes where Gl ut4 is present in the basal state. The distribution of Akt-2 in restin g adipocytes was found to substantially overlap with that of Glut4 whe n light microsomes were subfractionated by a sucrose velocity gradient indicating possible co-localization. We confirmed co-localization of Akt-2 and Glut4 in the basal state by immunopurification of Glut4 vesi cles, which exhibited a 5.5-fold increase in Akt-2 in response to insu lin relative to the amount of Glut4. These results are consistent with the possibility that Akt-2 may be involved in Glut4 vesicle transloca tion.