title: Phosphorylation of PTP1B at Ser(50) by Akt impairs its ability to dephosphorylate the insulin receptor.
abstract: PTP1B is a protein tyrosine phosphatase that negatively regulates insulin sensitivity by dephosphorylating the insulin receptor. Akt is a ser/thr kinase effector of insulin signaling that phosphorylates substrates at the consensus motif RXRXXS/T. Interestingly, PTP1B contains this motif (RYRDVS(50)), and wild-type PTP1B (but not mutants with substitutions for Ser(50)) was significantly phosphorylated by Akt in vitro. To determine whether PTP1B is a substrate for Akt in intact cells, NIH-3T3(IR) cells transfected with either wild-type PTP1B or PTP1B-S50A were labeled with [(32)P]-orthophosphate. Insulin stimulation caused a significant increase in phosphorylation of wild-type PTP1B that could be blocked by pretreatment of cells with wortmannin or cotransfection of a dominant inhibitory Akt mutant. Similar results were observed with endogenous PTP1B in untransfected HepG2 cells. Cotransfection of constitutively active Akt caused robust phosphorylation of wild-type PTP1B both in the absence and presence of insulin. By contrast, PTP1B-S50A did not undergo phosphorylation in response to insulin. We tested the functional significance of phosphorylation at Ser(50) by evaluating insulin receptor autophosphorylation in transfected Cos-7 cells. Insulin treatment caused robust receptor autophosphorylation that could be substantially reduced by coexpression of wild-type PTP1B. Similar results were obtained with coexpression of PTP1B-S50A. However, under the same conditions, PTP1B-S50D had an impaired ability to dephosphorylate the insulin receptor. Moreover, cotransfection of constitutively active Akt significantly inhibited the ability of wild-type PTP1B, but not PTP1B-S50A, to dephosphorylate the insulin receptor. We conclude that PTP1B is a novel substrate for Akt and that phosphorylation of PTP1B by Akt at Ser(50) may negatively modulate its phosphatase activity creating a positive feedback mechanism for insulin signaling.
authors.affiliation: Cardiology Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: L V
authors.initials: LV
authors.lastname: Ravichandran
authors.forename: H
authors.initials: H
authors.lastname: Chen
authors.forename: Y
authors.initials: Y
authors.lastname: Li
authors.forename: M J
authors.initials: MJ
authors.lastname: Quon
mesh_terms: D016475:3T3 Cells; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D016384:Consensus Sequence; D005246:Feedback; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D017346:Protein Serine-Threonine Kinases; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D011972:Receptor, Insulin / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection
chemical_list: D007328:Insulin; D011518:Proto-Oncogene Proteins; D010768:Phosphoserine; D011972:Receptor, Insulin; C494918:AKT1 protein, human; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; C516607:PTPN1 protein, human; D054562:Protein Tyrosine Phosphatase, Non-Receptor Type 1; D017027:Protein Tyrosine Phosphatases; C516609:Ptpn1 protein, mouse