The inhibition of glycogen synthase kinase-3 by insulin or insulin-like growth factor 1 in the rat skeletal muscle cell line L6 is blocked by wortmannin, but not by rapamycin: evidence that wortmannin blocks activation of the mitogen-activated protein kinase pathway in L6 cells between Ras and Raf.
Glycogen synthase kinase-3 (GSK3) is inactivated in vitro by p70 S6 kinase or MAP kinase-activated protein kinase-1 beta (MAPKAP kinase-1 beta; also known as Rsk-2). Here we show that GSK3 isoforms are inhibited by 40% within minutes after stimulation of the rat skeletal-muscle cell line L6 with insulin-like growth factor-1 (IGF-1) or insulin. GSK3 was similarly inhibited in rabbit skeletal muscle after an intravenous injection of insulin. Inhibition resulted from increased phosphorylation of GSK3, probably at a serine/threonine residue(s), because it was reversed by incubation with protein phosphatase-2A. Rapamycin blocked the activation of p70 S6 kinase by IGF-1 in L6 cells, but had no effect on the inhibition of GSK3 or the activation of MAPKAP kinase-1 beta. In contrast, wortmannin, a potent inhibitor of PtdIns 3-kinase, prevented the inactivation of GSK3 and the activation of MAPKAP kinase-1 beta and p70 S6 kinase by IGF-1 or insulin. Wortmannin also blocked the activation of p74raf-1. MAP kinase kinase and p42 MAP kinase, but not the formation of GTP-Ras by IGF-1. The results suggest that the stimulation of glycogen synthase by insulin/IGF-1 in skeletal muscle involves the MAP-KAP kinase-1-catalysed inhibition of GSK3, as well as the previously described activation of the glycogen-associated form of protein phosphatase-1.
Cross
D A
DA
Department of Biochemistry, University of Dundee, Scotland, UK.
Alessi
D R
DR
Vandenheede
J R
JR
McDowell
H E
HE
Hundal
H S
HS
Cohen
P
P
D000595:Amino Acid Sequence; D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D000906:Antibodies; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors*; D002460:Cell Line; D004789:Enzyme Activation; D005260:Female; D006006:Glycogen Synthase / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3; D038341:Glycogen Synthase Kinases; D007166:Immunosuppressive Agents / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D007700:Kinetics; D008969:Molecular Sequence Data; D009132:Muscles / Q000201:enzymology*; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D011090:Polyenes / Q000494:pharmacology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019908:Proto-Oncogene Proteins c-raf; D011817:Rabbits; D051381:Rats; D020123:Sirolimus; D000077191:Wortmannin; D018631:ras Proteins / Q000378:metabolism*
D000730:Androstadienes; D000906:Antibodies; D007166:Immunosuppressive Agents; D007328:Insulin; D010455:Peptides; D011090:Polyenes; D011518:Proto-Oncogene Proteins; D007334:Insulin-Like Growth Factor I; D006006:Glycogen Synthase; D038341:Glycogen Synthase Kinases; D017346:Protein Serine-Threonine Kinases; D019908:Proto-Oncogene Proteins c-raf; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D018631:ras Proteins; D020123:Sirolimus; D000077191:Wortmannin