A rapamycin-sensitive pathway down-regulates insulin signaling via phosphorylation and proteasomal degradation of insulin receptor substrate-1.
Insulin receptor substrate-1 (IRS-1) is a major substrate of the insulin receptor and acts as a docking protein for Src homology 2 domain containing signaling molecules that mediate many of the pleiotropic actions of insulin. Insulin stimulation elicits serine/threonine phosphorylation of IRS-1, which produces a mobility shift on SDS-PAGE, followed by degradation of IRS-1 after prolonged stimulation. We investigated the molecular mechanisms and the functional consequences of these phenomena in 3T3-L1 adipocytes. PI 3-kinase inhibitors or rapamycin, but not the MEK inhibitor, blocked both the insulin-induced electrophoretic mobility shift and degradation of IRS-1. Adenovirus-mediated expression of a membrane-targeted form of the p110 subunit of phosphatidylinositol (PI) 3-kinase (p110CAAX) induced a mobility shift and degradation of IRS-1, both of which were inhibited by rapamycin. Lactacystin, a specific proteasome inhibitor, inhibited insulin-induced degradation of IRS-1 without any effect on its electrophoretic mobility. Inhibition of the mobility shift did not significantly affect tyrosine phosphorylation of IRS-1 or downstream insulin signaling. In contrast, blockade of IRS-1 degradation resulted in sustained activation of Akt, p70 S6 kinase, and mitogen-activated protein (MAP) kinase during prolonged insulin treatment. These results indicate that insulin-induced serine/threonine phosphorylation and degradation of IRS-1 are mediated by a rapamycin-sensitive pathway, which is downstream of PI 3-kinase and independent of ras/MAP kinase. The pathway leads to degradation of IRS-1 by the proteasome, which plays a major role in down-regulation of certain insulin actions during prolonged stimulation.
Haruta
T
T
First Department of Medicine, Toyama Medical and Pharmaceutical University Japan. tharuta-tym@umin.ac.jp
Uno
T
T
Kawahara
J
J
Takano
A
A
Egawa
K
K
Sharma
P M
PM
Olefsky
J M
JM
Kobayashi
M
M
D016475:3T3 Cells; D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000256:Adenoviridae / Q000235:genetics; D017667:Adipocytes / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D003847:Deoxyglucose / Q000378:metabolism; D004622:Embryo, Mammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015870:Gene Expression; D006801:Humans; D007328:Insulin / Q000494:pharmacology*; D055504:Insulin Receptor Substrate Proteins; D007668:Kidney; D051379:Mice; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D046988:Proteasome Endopeptidase Complex; D015398:Signal Transduction / Q000187:drug effects*; D020123:Sirolimus / Q000494:pharmacology*; D014162:Transfection
D000111:Acetylcysteine; D003546:Cysteine Endopeptidases; D003847:Deoxyglucose; D004791:Enzyme Inhibitors; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D009097:Multienzyme Complexes; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010750:Phosphoproteins; D046988:Proteasome Endopeptidase Complex; D020123:Sirolimus; C526292:IRS1 protein, human; C526295:Irs1 protein, mouse; C067713:lactacystin