Mutation of the two carboxyl-terminal tyrosines in the insulin receptor results in enhanced activation of mitogen-activated protein kinase.
Activation of mitogen-activated protein (MAP) kinase represents an important mechanism in hormonal regulation. To clarify the role of MAP kinase activation in insulin action, we compared the activation of the enzyme in Rat-1 cells transfected with wild-type (Hirc) and mutant insulin receptors in which the 2 carboxyl-terminal tyrosines were substituted with phenylalanine (Y/F2). Expression of the Y/F2 mutant receptor enhanced the responsiveness of MAP kinase to insulin. Moreover, the insulin responsiveness of the activator of this enzyme, MAP kinase kinase, was also increased in these cells. To explore the early signaling events that might account for this increase in responsiveness, we evaluated the tyrosine phosphorylation of the insulin receptor substrate, IRS-1, and its subsequent association with phosphatidylinositol (PI)-3 kinase. In both cell types, insulin led to a dose-dependent increase in 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 determined in both assays. In previous studies, cells bearing this receptor mutant exhibited an identical metabolic response but enhanced mitogenic response to insulin when compared with wild-type receptor. These data provide further evidence for divergence of the mitogenic and metabolic signaling pathways at or near the insulin receptor.
Pang
L
L
Department of Physiology, University of Michigan, School of Medicine, Ann Arbor 48109.
Milarski
K L
KL
Ohmichi
M
M
Takata
Y
Y
Olefsky
J M
JM
Saltiel
A R
AR
D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D005982:Glutathione Transferase / Q000378:metabolism; D055504:Insulin Receptor Substrate Proteins; D019950:Mitogen-Activated Protein Kinase 1; D009154:Mutation; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D011972:Receptor, Insulin / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014443:Tyrosine / Q000235:genetics*
D005982:Glutathione Transferase; D055504:Insulin Receptor Substrate Proteins; D019950:Mitogen-Activated Protein Kinase 1; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D017853:Phosphotransferases (Alcohol Group Acceptor); D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D011993:Recombinant Fusion Proteins; D014443:Tyrosine; C526296:Irs1 protein, rat