PHOSPHORYLATION OF GLUT4-CONTAINING LOW-DENSITY MICROSOME PROTEINS BYINSULIN

To understand a possible insulin mediated translocation mechanism of t he insulin dependent glucose transpoter (GLUT4) from the intracellular vesicles to the plasma membrane, we investigated the phosphorylation of membrane proteins in GLUT4-containing vesicles by the kinases which are related to the Pas signalling system. When 3T3-L1 adipocytes were treated with insulin, the increase of GLUT4 translocation from the in tracellular vesicle to the plasma membrane was observed by immunofluor escence microscopy. In the anti-Ras immunoprecipitates from the 3T3-L1 adipocytes, there was no change in the amount of GAP and ERK by insul in treatement. However, the tyrosine phosphorylation of the GAP and ER K was increased in the anti-Ras immunoprecipitates from the insulin tr eated cells. In case of MEK, the amount was increased by the insulin t reatement. Several components of the immunopurified LDM proteins (62, 46, 42, and 33.5 kDa) were phosphorylated when LDM was incubated with the anti-Ras immunoprecipitates. In addition, the amount of P-32-incor poration into each component was increased when LDM was phosphorylated with the insulin-treated anti-Ras imunoprecipitates. These results su ggest that the phosphorylation of LDM components by activated Ras-asso ciated kinases may have roles on the insulin-regulated translocation o f GLUT4 molecules from the intracellular vesicle to the plasma membran e.