MUTATION OF THE 2 CARBOXYL-TERMINAL TYROSINES IN THE INSULIN-RECEPTORRESULTS IN ENHANCED ACTIVATION OF MITOGEN-ACTIVATED PROTEIN-KINASE

Activation of mitogen-activated protein (MAP) kinase represents an imp ortant mechanism in hormonal regulation. To clarify the role of MAP ki nase activation in insulin action, we compared the activation of the e nzyme in Rat-1 cells transfected with Wild-type (Hire) and mutant insu lin receptors in which the 2 carboxyl-terminal tyrosines were substitu ted with phenylalanine (Y/F2). Expression of the Y/F2 mutant receptor enhanced the responsiveness of MAP kinase to insulin. Moreover, the in sulin responsiveness of the activator of this enzyme, MAP kinase kinas e, was also increased in these cells. To explore the early signaling e vents that might account for this increase in responsiveness, we evalu ated the tyrosine phosphorylation of the insulin receptor substrate, I RS-1, and its subsequent association with phosphatidylinositol (PI)-3 kinase. In both cell types, insulin led to a dose dependent increase i n the association of tyrosine phosphorylated IRS-1 with the SH2 domain of the p85 regulatory subunit of PI-3 kinase, and also increased the amount of PI kinase activity detected in anti-IRS-1 immunoprecipitates . The effect of insulin was significantly greater in Y/F2 cells, as de termined in both assays. In previous studies, cells bearing this recep tor mutant exhibited an identical metabolic response but enhanced mito genic response to insulin when compared with wild-type receptor. These data provide further evidence for divergence of the mitogenic and met abolic signaling pathways at or near the insulin receptor.