DIFFERENTIAL ACTIVATION OF MITOGEN-ACTIVATED PROTEIN-KINASE BY INSULIN AND EPIDERMAL GROWTH-FACTOR IN 3T3-L1 ADIPOCYTES - A POSSIBLE INVOLVEMENT OF PI3-KINASE IN THE ACTIVATION OF THE MAP KINASE BY INSULIN

Mitogen-activated protein (MAP) kinase plays crucial roles in cell gro wth and differentiation. It has recently been shown that the MAP kinas e cascade in growth factor signaling diverges and cross-talks with oth er signaling pathways. In the present study, we examined the effects o f wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase (P I3-kinase), on the activation of Pas, Raf-1 kinase, and MAP kinase by insulin and epidermal growth factor (EGF). The effect of LY294002, a s tructurally distinct PI3-kinase inhibitor, on the activation of Raf-1 kinase by both ligands was also examined. In 3T3-L1 adipocytes, 25 nmo l/l wortmannin inhibited the insulin-induced activation of Raf-1 kinas e to the basal level, whereas the same dose of wortmannin had little e ffect on the EGF-induced activation of Raf-1 kinase. One hundred mu mo l/l LY294002 blocked insulin-induced activation of Raf-1 kinase withou t affecting EGF-induced activation of this kinase. Twenty-five nmol/l wortmannin inhibited the insulin-induced activation of MAP kinase to t he basal level with no effect on the EGF-induced activation of this ki nase. But the same dose of wortmannin did not affect the formation of guanosine 5'-triphosphate (GTP)bound Pas stimulated by either ligand. In KB cells, results similar to those in 3T3-L1 adipocytes were obtain ed. In contrast, in Chinese hamster ovary cells overexpressing the hum an insulin receptor (CHO-HIR cells), neither wortmannin nor LP294002 i nhibited the insulin-induced activation of Raf-1 kinase, and wortmanni n had little effect on the activation of MAP kinase by insulin. These results indicate that 1) PI3-kinase or wortmannin-sensitive molecules are involved in the interaction between activated Pas and Raf-1 kinase in the insulin signaling in 3T3-L1 adipocytes, 2) the involvement of PI3-kinase or wortmannin-sensitive molecules in the insulin-induced ac tivation of MAP kinase appears to be cell-type specific, and 3) differ ential mechanisms to activate Raf-1 kinase and MAP kinase by insulin a nd EGF exist.