title: Mechanism of feedback regulation of insulin receptor substrate-1 phosphorylation in primary adipocytes.
abstract: Serine and threonine phosphorylation of IRS-1 (insulin receptor substrate-1) has been reported to decrease its ability to be tyrosine-phosphorylated by the insulin receptor. Insulin itself may negatively regulate tyrosine phosphorylation of IRS-1 through a PI3K (phosphoinositide 3-kinase)-dependent feedback pathway. In the present study, we examined the regulation and role of IRS-1 serine phosphorylation in the modulation of IRS-1 tyrosine phosphorylation in physiologically relevant cells, namely freshly isolated primary adipocytes. We show that insulin-stimulated phosphorylation of Ser312 and Ser616 in IRS-1 was relatively slow, with maximal phosphorylation achieved after 20 and 5 min respectively. The effect of insulin on phosphorylation of both these sites required the activation of PI3K and the MAPKs (mitogen-activated protein kinases) ERK1/2 (extracellular-signal-regulated kinase 1 and 2), but not the activation of mTOR (mammalian target of rapamycin)/p70S6 kinase, JNK (c-Jun N-terminal kinase) or p38MAPK. Although inhibition of PI3K and ERK1/2 both substantially decreased insulin-stimulated phosphorylation of Ser312 and Ser616, only wortmannin enhanced insulin-stimulated tyrosine phosphorylation of IRS-1. Furthermore, inhibition of mTOR/p70S6 kinase, JNK or p38MAPK had no effect on insulin-stimulated IRS-1 tyrosine phosphorylation. The differential effect of inhibition of ERK1/2 on insulin-stimulated IRS-1 phosphorylation of Ser312/Ser616 and tyrosine indicates that these events are independent of each other and that phosphorylation of Ser312/Ser616 is not responsible for the negative regulation of IRS-1 tyrosine phosphorylation mediated by PI3K in primary adipocytes.
authors.affiliation: Department of Biochemistry, School of Medical Sciences, University of Bristol, Bristol BS8 1TD, UK. i.hers@bris.ac.uk
authors.forename: Ingeborg
authors.initials: I
authors.lastname: Hers
authors.forename: Jeremy M
authors.initials: JM
authors.lastname: Tavaré
mesh_terms: D017667:Adipocytes / Q000378:metabolism*; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002070:Butadienes / Q000494:pharmacology; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D025461:Feedback, Physiological; D007328:Insulin / Q000502:physiology; D055504:Insulin Receptor Substrate Proteins; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D048670:MAP Kinase Kinase 4; D008297:Male; D020929:Mitogen-Activated Protein Kinase Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009570:Nitriles / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000037:antagonists & inhibitors / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases; D013997:Time Factors; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D002070:Butadienes; D048049:Extracellular Signal-Regulated MAP Kinases; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D048670:MAP Kinase Kinase 4; D020929:Mitogen-Activated Protein Kinase Kinases; D009570:Nitriles; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D011494:Protein Kinases; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D058570:TOR Serine-Threonine Kinases; D000077191:Wortmannin; C526296:Irs1 protein, rat; C113580:U 0126; C546845:mTOR protein, rat