Phosphorylation of Ser307 in Insulin Receptor Substrate-1 Blocks Interactions with the Insulin Receptor and Inhibits Insulin Action

Abstract

Serine phosphorylation of insulin receptor substrate-1 (IRS-1) inhibits insulin signal transduction in a variety of cell backgrounds, which might contribute to peripheral insulin resistance. However, because of the large number of potential phosphorylation sites, the mechanism of inhibition has been difficult to determine. One serine residue located near the phosphotyrosine-binding (PTB) domain in IRS-1 (Ser307in rat IRS-1 or Ser312 in human IRS-1) is phosphorylated via several mechanisms, including insulin-stimulated kinases or stress-activated kinases like JNK1. During a yeast tri-hybrid assay, phosphorylation of Ser307 by JNK1 disrupted the interaction between the catalytic domain of the insulin receptor and the PTB domain of IRS-1. In 32D myeloid progenitor cells, phosphorylation of Ser307 inhibited insulin stimulation of the phosphatidylinositol 3-kinase and MAPK cascades. These results suggest that inhibition of PTB domain function in IRS-1 by phosphorylation of Ser307 (Ser312 in human IRS-1) might be a general mechanism to regulate insulin signaling. Serine phosphorylation of insulin receptor substrate-1 (IRS-1) inhibits insulin signal transduction in a variety of cell backgrounds, which might contribute to peripheral insulin resistance. However, because of the large number of potential phosphorylation sites, the mechanism of inhibition has been difficult to determine. One serine residue located near the phosphotyrosine-binding (PTB) domain in IRS-1 (Ser307in rat IRS-1 or Ser312 in human IRS-1) is phosphorylated via several mechanisms, including insulin-stimulated kinases or stress-activated kinases like JNK1. During a yeast tri-hybrid assay, phosphorylation of Ser307 by JNK1 disrupted the interaction between the catalytic domain of the insulin receptor and the PTB domain of IRS-1. In 32D myeloid progenitor cells, phosphorylation of Ser307 inhibited insulin stimulation of the phosphatidylinositol 3-kinase and MAPK cascades. These results suggest that inhibition of PTB domain function in IRS-1 by phosphorylation of Ser307 (Ser312 in human IRS-1) might be a general mechanism to regulate insulin signaling. insulin receptor substrate tumor necrosis factor c-Jun N-terminal kinase phosphotyrosine-binding mitogen-activated protein kinase insulin-like growth factor-1 glutathioneS-transferase JNK-interacting protein phosphatidylinositol 3-kinase insulin receptor synthetic dextrose extracellular signal-regulated kinase pleckstrin homology MAPK/ERK kinase The insulin signaling system plays an important role in many physiological processes, including carbohydrate and fat metabolism, reproduction, cellular growth, and survival (1White M.F. Myers M.G. DeGroot L.J. Jameson J.L. Endocrinology, Vol. 1., The Molecular Basis of Insulin Action. W. B. Saunders, Philadelphia2001Google Scholar). Acute insulin resistance is mediated, at least in part, by the action of pro-inflammatory cytokines that are produced during infection, physical trauma, or cancer (2Loddick S.A. Rothwell N.J. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 9449-9451Google Scholar, 3Hotamisligil G.S. Peraldi P. Budvari A. Ellis R.W. White M.F. Spiegelman B.M. Science. 1996; 271: 665-668Google Scholar, 4Yoshikawa T. Noguchi Y. Satoh S. Biochem. Biophys. Res. Commun. 1999; 256: 676-681Google Scholar). Chronic insulin resistance is an inevitable consequence of genetic variation that is exacerbated by aging and obesity and contributes to multiple disorders, including glucose intolerance, hyperlipidemia, hypertension and cardiovascular mortality, infertility and polycystic ovarian syndrome, and type II diabetes (5Maezono K. Osman A. Patti M.E. Cusi K. Pendergrass M. DeFronzo R.A. Mandarino L. Diabetes. 1998; 47 (abstr.): 333AGoogle Scholar,6Kahn B.B. Flier J.S. J. Clin. Invest. 2000; 106: 473-481Google Scholar). Insulin resistance alone might not cause diabetes if pancreatic β-cells secrete enough insulin to compensate for reduced sensitivity; however, type II diabetes eventually develops, possibly because hyperinsulinemia itself exacerbates the pre-existing resistance until β-cells eventually fail to compensate (7Kahn B.B. Cell. 1998; 92: 593-596Google Scholar). Understanding the molecular basis of insulin resistance will provide a rational basis for treatment of many related disorders. The insulin signaling system is complex, and a common mechanism to explain the occurrence of acute and chronic insulin resistance is difficult to identify. Mutations in the insulin receptor are an obvious source of lifelong insulin resistance, but they occur rarely and are not the common cause of type II diabetes (8Hani E.H. Suaud L. Boutin P. Chevre J.C. Durand E. Philippi A. Demenais F. Vionnet N. Furuta H. Velho G. Bell G.I. Laine B. Froguel P. J. Clin. Invest. 1998; 101: 521-526Google Scholar, 9Carboni J.M. Yan N. Cox A.D. Bustelo X. Graham S.M. Lynch M.J. Weinmann R. Seizinger B.R. Der C.J. Barbacid M. Manne V. Oncogene. 1995; 10: 1905-1913Google Scholar, 10Vaxillaire M. Rouard M. Yamagata K. Oda N. Kaisaki P.J. Boriraj V.V. Chevre J.C. Boccio V. Cox R.D. Lathrop G.M. Dussoix P. Philippe J. Timsit J. Charpentier G. Velho G. Bell G.I. Froguel P. Hum. Mol. Genet. 1997; 6: 583-586Google Scholar, 11Comb D.G. Roseman S. J. Biol. Chem. 1958; 232: 807-827Google Scholar). Generally, insulin resistance is a consequence of dysregulated insulin signaling that arises from various sources. Nonspecific or regulated degradation of elements in the insulin signaling pathway might cause insulin resistance (12Sun X.J. Goldberg J.L. Qiao L.Y. Mitchell J.J. Diabetes. 1999; 48: 1359-1364Google Scholar); elevated activity or expression of protein or lipid phosphatases, including PTP1B, SHIP2, and pTen, directly inhibits insulin signals (13Ishihara H. Sasaoka T. Hori H. Wada T. Hirai H. Haruta T. Langlois W.J. Kobayashi M. Biochem. Biophys. Res. Commun. 1999; 260: 265-272Google Scholar, 14Clement S. Krause U. Desmedt F. Tanti J.-F. Behrends J. Pesesse X. Sasaki T. Penninger J. Doherty M. Malaisse W. Dumont J.E. Le Maechand-Brustel Y. Erneux C. Hue L. Schurmans S. Nature. 2001; 409: 92-97Google Scholar). Covalent modification of the IRS1 proteins by serine phosphorylation is implicated in insulin resistance associated with obesity and trauma. Serine phosphorylation of IRS-1 is known to be promoted by elevated circulating levels of several metabolites, including free fatty acids, diacylglycerol, fatty acyl-CoAs, ceramides, and glucose (15Shulman G.I. J. Clin. Invest. 2000; 106: 171-176Google Scholar). Moreover, adipose-derived cytokines like TNF-α also stimulate serine/threonine phosphorylation of IRS-1, which inhibits signaling (16Peraldi P. Hotamisligil G.S. Buurman W.A. White M.F. Spiegelman B.M. J. Biol. Chem. 1996; 271: 13018-13022Google Scholar). One of the branches of the TNF-α signaling pathway involves activation of JNK (17Yuasa T. Ohno S. Kehrl J.H. Kyriakis J.M. J. Biol. Chem. 1998; 273: 22681-22692Google Scholar, 18Kuan C.Y. Yang D.D. Samanta Roy D.R. Davis R.J. Rakic P. Flavell R.A. Neuron. 1999; 22: 667-676Google Scholar, 19Rincon M. Whitmarsh A. Yang D.D. Weiss L. Derijard B. Jayaraj P. Davis R.J. Flavell R.A. J. Exp. Med. 1998; 188: 1817-1830Google Scholar). JNK phosphorylates numerous cellular proteins, including IRS-1, IRS-2, Shc, and Gab-1 (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google Scholar). Previous work has revealed that the major JNK phosphorylation site in rat IRS-1 is located at Ser307 (Ser312 in human IRS-1), which is located on the C-terminal side of the phosphotyrosine-binding (PTB) domain (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google Scholar). In this report, a yeast tri-hybrid assay revealed that JNK1 phosphorylation of Ser307 inhibits the interaction between IRS-1 and the insulin receptor, providing a rational mechanism to explain, at least in part, the insulin resistance that occurs during trauma and obesity. Phospho-specific MAPK, control MAPK, and phosphoro-specific Akt antibodies were purchased from New England Biolabs Inc. Control anti-Akt and anti-JNK1 antibodies were purchased from Santa Cruz Biotechnology. Anti-phosphotyrosine antibodies were purchased from Transduction Laboratories. Antibodies against IRS-1, IRS-2, and p85 have been described (21Sun X.J. Wang L.M. Zhang Y. Yenush L. Myers Jr., M.G. Glasheen E.M. Lane W.S. Pierce J.H. White M.F. Nature. 1995; 377: 173-177Google Scholar, 22Pons S. Asano T. Glasheen E.M. Miralpeix M. Zhang Y. Fisher T.L. Myers Jr., M.G. Sun X.J. White M.F. Mol. Cell. Biol. 1995; 15: 4453-4465Google Scholar). Antibodies directed against phosphorylated Ser612 in IRS-1 were purchased from BIOSOURCE. Rabbit polyclonal serum directed against phosphorylated Ser307 was generated using a synthetic peptide designed to contain phosphorylated Ser307 and surrounding amino acids (Boston Biomolecules). Insulin was purchased from Roche Molecular Biochemicals. IGF-1 was a gift from Lilly. TNF was purchased from R&D Systems. IRS-1 tyrosine phosphorylation site mutants have been previously described (23Myers Jr., M.G. Zhang Y. Aldaz G.A.I. Grammer T.C. Glasheen E.M. Yenush L. Wang L.M. Sun X.J. Blenis J. Pierce J.H. White M.F. Mol. Cell. Biol. 1996; 16: 4147-4155Google Scholar, 24Myers Jr., M.G. Wang L.M. Sun X.J. Zhang Y. Yenush L. Schlessinger J. Pierce J.H. White M.F. Mol. Cell. Biol. 1994; 14: 3577-3587Google Scholar). Point mutants for Ser307 and in the JNK-binding domain of IRS-1 were generated using appropriate oligonucleotides with the Stratagene QuikChange site-directed mutagenesis method. JNK1 and GST-JIP JNK-binding domain constructs have been described (25Derijard B. Hibi M. Wu I.H. Barrett T. Su B. Deng T. Karin M. Davis R.J. Cell. 1994; 76: 1025-1037Google Scholar, 26Dickens M. Rogers J.S. Cavanagh J. Raitano A. Xia Z. Halpern J.R. Greenberg M.E. Sawyers C.L. Davis R.J. Science. 1997; 277: 693-696Google Scholar). 32D cells were maintained in RPMI 1640 medium supplemented with 10% fetal bovine serum, 5% WEHI conditioned medium (as a source of interleukin-3), and 5 mm histidinol and made quiescent by serum starvation for 4 h. 32D transfectants were generated by electroporation and selected in histidinol as previously described (27Yenush L. Zanella C. Uchida T. Bernal D. White M.F. Mol. Cell. Biol. 1998; 18: 6784-6794Google Scholar). HEK293 cells were maintained in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum and made quiescent by serum starvation for 12 h. 32D cells were lysed in 50 mm Tris (pH 7.4) containing 130 mm NaCl, 5 mm EDTA, 1.0% Nonidet P-40, 100 mm NaF, 50 mm β-glycerophosphate, 100 μm NaVO4, 1 mmphenylmethylsulfonyl fluoride, 5 μg/ml leupeptin, and 5 μg/ml aprotinin. Immunoprecipitations were performed for 2 h at 4 °C, followed by collection on protein A-Sepharose. Lysates and immunoprecipitates were resolved by SDS-PAGE and transferred to nitrocellulose, and proteins were detected by immunoblotting and either 125I-labeled protein A or enhanced chemiluminescence (Amersham-Pharmacia) and analysis by autoradiography or on a Molecular Dynamics PhosphorImager. HEK293 cells were lysed in 20 mm Tris (pH 7.4) containing 137 mm NaCl, 25 mm β-glycerophosphate, 2 mm sodium pyrophosphate, 2 mm EDTA, 1% 10% 1 mm fluoride, 5 μg/ml leupeptin, 5 μg/ml 2 mm and proteins containing of IRS-1 were made by the and using proteins were with cell for 2 h at 4 to the proteins were by transferred to nitrocellulose, and by with antibodies against JNK1. HEK293 cells were with either or using transfectants were made quiescent by serum starvation for 12 h and at h. stimulation with μg/ml and was with for 2 h at 4 °C, and were on was with peptide in kinase mm (pH 25 25 mm 100 μm sodium and at 4 were by of kinase and 50 μm to insulin receptor in a of 50 of kinase were at with and of SDS-PAGE and to nitrocellulose, phosphorylation of substrate proteins was by autoradiography and activity were performed on IRS-1 immunoprecipitates as previously described T. Myers Jr., M.G. White M.F. Mol. Cell. Biol. 2000; Scholar). The yeast system were purchased The yeast and the and constructs were a gift from A. was with constructs by the to were on the appropriate for at were and for 5 at to expression of proteins and to was as described with various and a containing various were on the appropriate for at were and The were and at for 5 to growth and to expression of IRS-1 of rat or human many potential serine phosphorylation that are to during insulin signaling. One of sites, Ser307 in rat IRS-1, was to be phosphorylated by Ser307 was to be phosphorylated in IRS-1 from cells and with or (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google Scholar, L. V. G.I. A. A. A. White M.F. J. Clin. Invest. 2001; Scholar). of Ser307 is because inhibits insulin-stimulated tyrosine phosphorylation of rat IRS-1. the role of Ser307 in insulin rat IRS-1 or a IRS-1 containing a was in 32D cells are myeloid progenitor cells that insulin and of IRS-1 and the insulin receptor many of the insulin signaling pathway in 32D cells T. Myers Jr., M.G. White M.F. Mol. Cell. Biol. 2000; Scholar, L.M. Myers Jr., M.G. Sun X.J. S.A. White M.F. Pierce J.H. Science. Scholar). of IRS-1 from cells with insulin or were by immunoblotting with antibodies against for antibodies with IRS-1, that phosphorylation were phosphorylated the or insulin phosphorylation of Ser307 and Ser612 was phosphorylated during stimulation previously (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google TNF-α either insulin or Ser307 is a common phosphorylation site for several signaling that was detected by immunoblotting with Insulin phosphorylation of Ser307 in cells was to insulin to to the expression of insulin receptor in Previous results that insulin-stimulated Ser307 phosphorylation L. V. G.I. A. A. A. White M.F. J. Clin. Invest. 2001; Scholar). insulin-stimulated Ser307 phosphorylation tyrosine was in potential tyrosine phosphorylation and to various proteins, including and (23Myers Jr., M.G. Zhang Y. Aldaz G.A.I. Grammer T.C. Glasheen E.M. Yenush L. Wang L.M. Sun X.J. Blenis J. Pierce J.H. White M.F. Mol. Cell. Biol. 1996; 16: 4147-4155Google Scholar). not phosphorylated at Ser307 during insulin was phosphorylated during and TNF-α stimulation Ser307 phosphorylation was by site-directed mutagenesis that in including and the in not Moreover, of the in IRS-1 not insulin-stimulated Ser307 phosphorylation several signaling including insulin stimulation of the at Ser307 to or inhibition of IRS-1 signaling to the insulin During in cells and the PTB domain of IRS-1 to the phosphorylated in the of the insulin receptor A. J. Biol. Chem. 1995; Scholar, A. Mol. Cell. Biol. 1994; 14: Scholar, G. T. E. L. Lynch A. White M.F. M. J. J. Biol. Chem. 1995; Scholar, L. J. Myers Jr., M.G. White M.F. J. Biol. Chem. 1996; 271: Scholar). In cells, this interaction phosphorylation of IRS-1 during insulin Ser307 is near the PTB domain of IRS-1, phosphorylation of this residue during insulin or TNF-α stimulation might the interaction between the insulin receptor and IRS-1. phosphorylation of Ser307 inhibits between the insulin receptor and IRS-1, a yeast tri-hybrid assay was to the of JNK1 on the interaction between the insulin receptor and various IRS-1 constructs work revealed that the PTB domain the to IRS-1 in yeast A. Mol. Cell. Biol. 1994; 14: Scholar); however, to the tri-hybrid assay, the interaction between JNK1 and IRS-1 in yeast was IRS-1 homology between and and and and and and in the rat that might JNK1 (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google Scholar, J. Whitmarsh Cavanagh J. M. Davis R.J. Mol. Cell. Biol. 1999; Scholar). The of this in is for JNK interaction M. Rogers J.S. Cavanagh J. Raitano A. Xia Z. Halpern J.R. Greenberg M.E. Sawyers C.L. Davis R.J. Science. 1997; 277: 693-696Google Scholar). which with various constructs of IRS-1 were with JNK1 in the yeast IRS-1 with JNK1 as revealed by activity in yeast on an IRS-1 homology not yeast growth IRS-1 constructs or which contain the homology in the promoted growth and activity These results suggest that the homology in human IRS-1 between and to JNK1 in the yeast In that the homology in rat IRS-1 JNK1. proteins containing a of rat IRS-1 with or homology were with cell containing JNK1. JNK1 associated with the of IRS-1 However, a but not a the of rat IRS-1 to JNK1 cells the human insulin receptor and human IRS-1 constructs on and growth was not inhibited JNK1 or an was in yeast cells of JNK1 growth, that the interaction was inhibited of Ser312 for in human IRS-1 to Ser307 in rat IRS-1) growth that phosphorylation of Ser307 is for the of the Moreover, human IRS-1 of including Ser312 but the homology was to of the interaction The insulin receptor not with JNK1 in a yeast assay not and was not phosphorylated by JNK1 during in kinase using insulin receptor and JNK1 These results are with the that the interaction between the insulin receptor and IRS-1 is inhibited by phosphorylation of Ser307 during of JNK1 with the homology of IRS-1. was to phosphorylation of Ser307 in rat IRS-1 and to the on insulin-stimulated and MAPK cascades. Ser307 phosphorylation of IRS-1, with a 100 A and and that Ser307 was from the and The immunoblotting of by might with phosphorylation in IRS-1, as phosphorylation of IRS-1 was detected by immunoblotting with antibodies as previously described Jr., M.G. Zhang Y. Yenush L. Glasheen E.M. Grammer T.C. Wang L.M. Blenis J. Sun X.J. Pierce J.H. White M.F. Diabetes. 1995; (abstr.): Scholar). of cells for with or μg/ml inhibited insulin-stimulated tyrosine phosphorylation by and inhibition with μg/ml of on tyrosine phosphorylation of and inhibition at μg/ml The of not occur degradation of IRS-1 or inhibition of insulin receptor not However, inhibition of IRS-1 tyrosine phosphorylation a homology of the of the JNK-binding domain of to JNK interaction M. Rogers J.S. Cavanagh J. Raitano A. Xia Z. Halpern J.R. Greenberg M.E. Sawyers C.L. Davis R.J. Science. 1997; 277: 693-696Google Scholar). J. of the homology by of the to a the of These results that the homology inhibition of insulin-stimulated tyrosine phosphorylation of in cell from and with the of for to stimulation with insulin for 5 were with antibodies IRS-1 JNK-binding domain insulin signals are the of in IRS-1 to the in various signaling proteins including and L. White M.F. 1997; Scholar). with the inhibition of insulin-stimulated tyrosine phosphorylation of IRS-1, inhibited the of p85 to IRS-1 in however, the of to p85 during insulin stimulation was not inhibited by In the inhibited insulin-stimulated activity associated with IRS-1, but on activity associated with during insulin stimulation Insulin the of with IRS-1 or Shc, which the phosphorylation of and as detected by immunoblotting with antibodies (1White M.F. Myers M.G. DeGroot L.J. Jameson J.L. Endocrinology, Vol. 1., The Molecular Basis of Insulin Action. W. B. Saunders, Philadelphia2001Google Scholar). In cells, analysis of the of on phosphorylation was by stimulation of phosphorylation by and insulin the pathway in the of IRS-1 or expression was phosphorylated during insulin stimulation of either IRS-1 or and inhibited insulin stimulation of phosphorylation inhibition at in phosphorylation was inhibited at the in cells These that phosphorylation of Ser307 inhibits results a general mechanism for the and of the IRS-1 of the insulin signaling pathway inhibition of PTB domain function by phosphorylation of Previous work that the interaction in yeast between the insulin receptor catalytic domain and IRS-1 is the of the phosphorylated in the insulin receptor to the PTB domain in IRS-1 A. J. Biol. Chem. 1995; Scholar, A. Mol. Cell. Biol. 1994; 14: Scholar, W. A. Mol. Cell. Biol. 1995; 15: Scholar). on this that of the between the insulin receptor and IRS-1 in yeast JNK1 occurs because phosphorylation of Ser307 PTB domain function G. T. E. L. Lynch A. White M.F. M. J. J. Biol. Chem. 1995; Scholar, L. J. Myers Jr., M.G. White M.F. J. Biol. Chem. 1996; 271: Scholar). of the control this including of the insulin receptor and IRS-1 in yeast a JNK1 of the insulin receptor and a human IRS-1 (Ser312 in yeast a JNK1 and of the insulin receptor and an IRS-1 the homology in yeast JNK1. the yeast tri-hybrid assay that insulin the suggest that inhibits IRS-1 tyrosine phosphorylation by at Previous work revealed that phosphorylation of IRS-1 on N-terminal the pleckstrin homology domain and the PTB domain L. J. Myers Jr., M.G. White M.F. J. Biol. Chem. 1996; 271: Scholar, S. H. J. Myers Jr., M.G. Yenush L. White M.F. J. Biol. Chem. 1997; Scholar). of the and PTB inhibits phosphorylation during insulin stimulation of cells, of either the or PTB domain tyrosine Ser307 phosphorylation inhibits PTB domain the domain might be for inhibition of IRS-1 tyrosine of IRS-1 to levels of insulin either the or PTB domain is in cells levels of insulin in cells with a number of Ser307 phosphorylation might a major Ser307 phosphorylation might be in cells with a number of L. J. Myers Jr., M.G. White M.F. J. Biol. Chem. 1996; 271: Scholar). the might be including degradation of IRS-1. is with the that serine phosphorylation of the insulin receptor or the proteins inhibits signal the potential of this the of phosphorylation and the have been difficult to identify. serine phosphorylation of IRS-1 is a common in insulin resistance and type II diabetes A. K. J. Clin. Invest. 1999; Scholar). IRS-1 inhibits insulin-stimulated of the insulin receptor, glucose and insulin-stimulated G.S. Peraldi P. Budvari A. Ellis R.W. White M.F. Spiegelman B.M. Science. 1996; 271: 665-668Google Scholar, K. R.A. J. Biol. Chem. 1997; Scholar, J.E. F. R.A. Mol. 1994; Scholar, J. K. R.A. J. Biol. Chem. 1999; Scholar, E. J. Biol. Chem. 1996; 271: Scholar, J.-F. T. E. Le Y. J. Biol. Chem. 1994; Scholar, P. 1995; Scholar, J.M. J.E. J. Biol. Chem. Scholar, G. B. M. E. B. A. L. H. A. H. J. Clin. Invest. 1996; Scholar, K. R. D. A. E. H. Y. J. Biol. Chem. 1997; Scholar). the JNK phosphorylation site at IRS-1 serine/threonine in for many protein including kinase protein protein kinase MAPK, and protein kinase (21Sun X.J. Wang L.M. Zhang Y. Yenush L. Myers Jr., M.G. Glasheen E.M. Lane W.S. Pierce J.H. White M.F. Nature. 1995; 377: 173-177Google Scholar, K. R.A. J. Biol. Chem. 1997; Scholar, J. K. R.A. J. Biol. Chem. 1999; Scholar, X.J. J.M. E. White M.F. Nature. Scholar, A. K. R.A. Biochem. Biophys. Res. Commun. 1998; Scholar, K. R.A. 1997; Scholar). suggest that serine phosphorylation of IRS-1 inhibits to with the insulin receptor and to as a substrate for tyrosine phosphorylation G.S. Peraldi P. Budvari A. Ellis R.W. White M.F. Spiegelman B.M. Science. 1996; 271: 665-668Google Scholar, K. R.A. J. Biol. Chem. 1997; Scholar, E. J. Biol. Chem. 1996; 271: Scholar, K. R. D. A. E. H. Y. J. Biol. Chem. 1997; Scholar, K. R.A. 1997; Scholar, H. R. R. A. J. Biol. Chem. 1995; Scholar). the of serine/threonine of signal inhibition might a molecular basis for insulin resistance that the of type II Ser307 phosphorylation general inhibition of IRS-1 as revealed by reduced activation of the and MAPK cascades. not occur inhibition of insulin receptor but is with reduced between the insulin receptor and IRS-1. of p85 or with IRS-1 on of tyrosine phosphorylation that are by to amino acids in the These results are with the general inhibition of tyrosine phosphorylation during inhibition of PTB domain function by Ser307 general inhibition is in to the inhibition of p85 at tyrosine phosphorylation directly to previously serine L. Myers Jr., M.G. White M.F. E. 1998; Scholar, E. J. Biol. Chem. 1996; 271: Scholar, K. R.A. 1997; Scholar). least kinases phosphorylation of including a kinase JNK and an kinase that is inhibited by and that JNK might be the common that Ser307 phosphorylation of various an JNK1 to IRS-1 (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google and the JNK-binding of IRS-1 is for the inhibition of insulin-stimulated tyrosine phosphorylation of IRS-1 by However, with potential physiological of insulin resistance not this and TNF-α stimulate JNK and Ser307 the kinase inhibits with on JNK activity L. V. G.I. A. A. A. White M.F. J. Clin. Invest. 2001; Scholar). insulin JNK in cells, this pathway is not inhibited by that a is in to at least kinases phosphorylation of These kinases might the common to to the JNK-binding domain in IRS-1, be In potential of chronic insulin resistance, as TNF-α and to of IRS-1 that are phosphorylated at Ser307 and that to the insulin Chronic might also IRS-1 for degradation or to to the insulin proteins, IRS-2, might be to serine a homology a residue to Ser307 not in (20Aguirre V. Uchida T. Yenush L. Davis R.J. White M.F. J. Biol. Chem. 2000; 275: 9047-9054Google Scholar). is during TNF-α or which inhibits insulin-stimulated tyrosine IRS-1 is to insulin in serine inhibition might peripheral insulin resistance and of the phosphorylation in that insulin-stimulated tyrosine phosphorylation and the kinase to is an important for for for and and for