Elevation of global O-GlcNAc levels in 3T3-L1 adipocytes by selective inhibition of O-GlcNAcase does not induce insulin resistance.
The O-GlcNAc post-translational modification is considered to act as a sensor of nutrient flux through the hexosamine biosynthetic pathway. A cornerstone of this hypothesis is that global elevation of protein O-GlcNAc levels, typically induced with the non-selective O-GlcNAcase inhibitor PUGNAc (O-(2-acetamido-2-deoxy-D-glycopyranosylidene) amino-N-phenylcarbamate), causes insulin resistance in adipocytes. Here we address the potential link between elevated O-GlcNAc and insulin resistance by using a potent and selective inhibitor of O-GlcNAcase (NButGT (1,2-dideoxy-2'-propyl-alpha-D-glucopyranoso-[2,1-D]-Delta 2'-thiazoline), 1200-fold selectivity). A comparison of the structures of a bacterial homologue of O-GlcNAcase in complex with PUGNAc or NButGT reveals that these inhibitors bind to the same region of the active site, underscoring the competitive nature of their inhibition of O-GlcNAcase and the molecular basis of selectivity. Treating 3T3-L1 adipocytes with NButGT induces rapid increases in global O-GlcNAc levels, but strikingly, NButGT treatment does not replicate the insulin desensitizing effects of the non-selective O-GlcNAcase inhibitor PUGNAc. Consistent with these observations, NButGT also does not recapitulate the impaired insulin-mediated phosphorylation of Akt that is induced by treatment with PUGNAc. Collectively, these results suggest that increases in global levels of O-GlcNAc-modified proteins of cultured adipocytes do not, on their own, cause insulin resistance.
Macauley
Matthew S
MS
Department of Chemistry, Simon Fraser University, Burnaby, British Columbia V5A 1S6, Canada.
Bubb
Abigail K
AK
Martinez-Fleites
Carlos
C
Davies
Gideon J
GJ
Vocadlo
David J
DJ
D041721:3T3-L1 Cells; D000117:Acetylglucosamine / Q000378:metabolism*; D000118:Acetylglucosaminidase / Q000378:metabolism*; D017667:Adipocytes / Q000378:metabolism; D000818:Animals; D019086:Bridged Bicyclo Compounds, Heterocyclic / Q000378:metabolism*; D020134:Catalytic Domain; D004305:Dose-Response Relationship, Drug; D007333:Insulin Resistance; D007700:Kinetics; D051379:Mice; D008954:Models, Biological; D008956:Models, Chemical; D008968:Molecular Conformation; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational
C555050:1,2-dideoxy-2'-propylglucopyranoso(2,1-d)-delta 2'-thiazoline; D019086:Bridged Bicyclo Compounds, Heterocyclic; D000118:Acetylglucosaminidase; D000117:Acetylglucosamine