Insulin resistance of glycogen synthase mediated by o-linked N-acetylglucosamine.
We have investigated the mechanism by which high concentrations of glucose inhibit insulin stimulation of glycogen synthase. In NIH-3T3-L1 adipocytes cultured in low glucose (LG; 2.5 mm), the half-maximal activation concentration (A(0.5)) of glucose 6-phosphate was 162 +/- 15 microm. Exposure to either high glucose (HG; 20 mm) or glucosamine (GlcN; 10 mm) increased the A(0.5) to 558 +/- 61 or 612 +/- 34 microm. Insulin treatment with LG reduced the A(0.5) to 96 +/- 10 microm, but cells cultured with HG or GlcN were insulin-resistant (A(0.5) = 287 +/- 27 or 561 +/- 77 microm). Insulin resistance was not explained by increased phosphorylation of synthase. In fact, culture with GlcN decreased phosphorylation to 61% of the levels seen in cells cultured in LG. Hexosamine flux and subsequent enzymatic protein O-glycosylation have been postulated to mediate nutrient sensing and insulin resistance. Glycogen synthase is modified by O-linked N-acetylglucosamine, and the level of glycosylation increased in cells treated with HG or GlcN. Treatment of synthase in vitro with protein phosphatase 1 increased basal synthase activity from cells cultured in LG to 54% of total activity but was less effective with synthase from cells cultured in HG or GlcN, increasing basal activity to only 13 or 16%. After enzymatic removal of O-GlcNAc, however, subsequent digestion with phosphatase increased basal activity to over 73% for LG, HG, and GlcN. We conclude that O-GlcNAc modification of glycogen synthase results in the retention of the enzyme in a glucose 6-phosphate-dependent state and contributes to the reduced activation of the enzyme in insulin resistance.
Parker
Glendon J
GJ
Veterans Affairs Medical Center and Division of Endocrinology, University of Utah School of Medicine, Salt Lake City 84132, USA.
Lund
Kelli C
KC
Taylor
Rodrick P
RP
McClain
Donald A
DA
D016475:3T3 Cells; D000117:Acetylglucosamine / Q000378:metabolism*; D000818:Animals; D002478:Cells, Cultured; D004789:Enzyme Activation; D005944:Glucosamine / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D006006:Glycogen Synthase / Q000378:metabolism*; D006031:Glycosylation; D007333:Insulin Resistance; D051379:Mice; D010766:Phosphorylation
D006006:Glycogen Synthase; D005947:Glucose; D005944:Glucosamine; D000117:Acetylglucosamine