Expression of catalytically inactive Syp phosphatase in 3T3 cells blocks stimulation of mitogen-activated protein kinase by insulin.
To explore the role of the protein tyrosine phosphatase Syp in insulin signaling, a catalytically inert mutant Syp protein was expressed under an inducible promoter in cells transfected with the human insulin receptor. Expression of the mutant phosphatase significantly reduced the stimulation of mitogenesis by insulin, indicating that the mutation produced a dominant negative phenotype. Tyrosine phosphorylation of both the insulin receptor and its major substrates, Shc and insulin receptor substrate-1, were unaffected by the mutant phosphatase. However, both the insulin-dependent tyrosine phosphorylation and activation of mitogen-activated protein kinase were markedly attenuated. Expression of the mutant phosphatase allowed the detection of a 120-kDa protein phosphorylated in response to insulin that associated with the src homology (SH) 2 domains of the phosphatase, suggesting a possible regulatory role for this protein. These results indicate that the activity of Syp plays a critical part in the mitogenic actions of insulin.
Milarski
K L
KL
Department of Signal Transduction, Parke-Davis Pharmaceutical Research, Warner Lambert Co., Ann Arbor, Michigan 48105.
Saltiel
A R
AR
D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D017931:DNA Primers; D004789:Enzyme Activation; D007328:Insulin / Q000494:pharmacology*; D007329:Insulin Antagonists / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D019950:Mitogen-Activated Protein Kinase 1; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000096:biosynthesis / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism
D017931:DNA Primers; C526292:IRS1 protein, human; D007328:Insulin; D007329:Insulin Antagonists; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; C526295:Irs1 protein, mouse; D010750:Phosphoproteins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; C516724:PTPN11 protein, human; C509249:PTPN6 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse