title: The level of insulin receptor tyrosine kinase activity modulates the activities of phosphatidylinositol 3-kinase, microtubule-associated protein, and S6 kinases.
abstract: The role of insulin receptor tyrosine kinase activity in stimulation of intracellular enzymes linked to insulin action [phosphatidylinositol 3-kinase (PtdIns 3-kinase), microtubule-associated protein (MAP) kinase, and S6 kinases] was studied in Chinese hamster ovary cells which overexpress wild type human insulin receptors, receptors with reduced kinase activity due to substitution of Phe for Tyr1146 (single-Phe), Tyr1150,1151 (double-Phe), and Tyr1146,1150,1151 (triple-Phe), or kinase-inactive receptors with a substitution of Ala for Lys1018 in the ATP binding site (A1018). We have previously shown that receptor autophosphorylation and kinase activity of these mutants were reduced by approximately 50, 65, 85, and 100%, respectively. Glycogen and DNA synthesis parallel the level of receptor autophosphorylation and kinase activity; however, receptor serine and threonine phosphorylation was independent of receptor tyrosine kinase activity and receptor internalization was completely dependent on maximal receptor kinase activity. Overexpression of the wild type insulin receptor increased both maximal insulin receptor substrate-1-associated and total insulin-stimulated PtdIns 3-kinase activity, as well as S6 and MAP kinase activities 2.0- to 3.6-fold. In addition there was a leftward shift of the dose-response curves for PtdIns 3-kinase and S6 kinases by approximately 10-fold. Expression of the single- and double-Phe mutant receptors also enhanced maximal PtdIns 3-kinase activity, but had no effect on insulin sensitivity, whereas expression of either the triple-Phe or kinase-inactive receptors did not enhance insulin stimulation or increase insulin sensitivity as compared to the control cells. When comparing the mutant and wild type receptors, differences in insulin sensitivity were least for insulin-stimulated MAP kinase and greatest for S6 kinase; with the latter there was greater than a 1000-fold difference in insulin sensitivity when cells that overexpress wild type vs. kinase-inactive insulin receptors were compared. Thus, the level of insulin receptor tyrosine autophosphorylation and kinase activity regulate both maximal activation and insulin sensitivity of these intracellular kinases in the insulin action pathway which may lead to glycogen and/or DNA synthesis. The differential sensitivity of these enzymes to changes in receptor activation suggests that they may be differently coupled to the receptor kinase.
authors.affiliation: Joslin Diabetes Center, Department of Medicine, Brigham and Women's Hospital, Boston, Massachusetts 02215.
authors.forename: P A
authors.initials: PA
authors.lastname: Wilden
authors.forename: C R
authors.initials: CR
authors.lastname: Kahn
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004789:Enzyme Activation; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D017354:Point Mutation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D019893:Ribosomal Protein S6 Kinases; D014162:Transfection
chemical_list: D007328:Insulin; D008869:Microtubule-Associated Proteins; D019869:Phosphatidylinositol 3-Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D017346:Protein Serine-Threonine Kinases; D011972:Receptor, Insulin; D011994:Recombinant Proteins; D019893:Ribosomal Protein S6 Kinases