title: Hexosamine biosynthesis impairs insulin action via a cholesterolgenic response.
abstract: Plasma membrane cholesterol accumulation has been implicated in cellular insulin resistance. Given the role of the hexosamine biosynthesis pathway (HBP) as a sensor of nutrient excess, coupled to its involvement in the development of insulin resistance, we delineated whether excess glucose flux through this pathway provokes a cholesterolgenic response induced by hyperinsulinemia. Exposing 3T3-L1 adipocytes to physiologically relevant doses of hyperinsulinemia (250pM-5000pM) induced a dose-dependent gain in the mRNA/protein levels of 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase (HMGR). These elevations were associated with elevated plasma membrane cholesterol. Mechanistically, hyperinsulinemia increased glucose flux through the HBP and O-linked β-N-acetylglucosamine (O-GlcNAc) modification of specificity protein 1 (Sp1), known to activate cholesterolgenic gene products such as the sterol response element-binding protein (SREBP1) and HMGR. Chromatin immunoprecipitation demonstrated that increased O-GlcNAc modification of Sp1 resulted in a higher binding affinity of Sp1 to the promoter regions of SREBP1 and HMGR. Luciferase assays confirmed that HMGR promoter activity was elevated under these conditions and that inhibition of the HBP with 6-diazo-5-oxo-l-norleucine (DON) prevented hyperinsulinemia-induced activation of the HMGR promoter. In addition, both DON and the Sp1 DNA-binding inhibitor mithramycin prevented the hyperinsulinemia-induced increases in HMGR mRNA/protein and plasma membrane cholesterol. In these mithramycin-treated cells, both cortical filamentous actin structure and insulin-stimulated glucose transport were restored. Together, these data suggest a novel mechanism whereby increased HBP activity increases Sp1 transcriptional activation of a cholesterolgenic program, thereby elevating plasma membrane cholesterol and compromising cytoskeletal structure essential for insulin action.
authors: Departments of Cellular and Integrative Physiology, Indiana UniversitySchool of Medicine, Indianapolis, IN 46202, USA.
authors: Brent A
authors: BA
authors: Penque
authors: April M
authors: AM
authors: Hoggatt
authors: B Paul
authors: BP
authors: Herring
authors: Jeffrey S
authors: JS
authors: Elmendorf
mesh_terms: D041721:3T3-L1 Cells; D000818:Animals; D053898:Biosynthetic Pathways / Q000187:drug effects; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002784:Cholesterol / Q000096:biosynthesis*; D004247:DNA / Q000378:metabolism; D003980:Diazooxonorleucine / Q000494:pharmacology; D005947:Glucose / Q000378:metabolism; D006031:Glycosylation / Q000187:drug effects; D006595:Hexosamines / Q000096:biosynthesis*; D006801:Humans; D006903:Hydroxymethylglutaryl CoA Reductases / Q000235:genetics; D006946:Hyperinsulinism / Q000503:physiopathology; D007328:Insulin / Q000494:pharmacology*; D007333:Insulin Resistance; D051379:Mice; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding / Q000187:drug effects / Q000235:genetics; D021381:Protein Transport / Q000187:drug effects; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051780:Sterol Regulatory Element Binding Protein 1 / Q000378:metabolism; D015533:Transcriptional Activation / Q000187:drug effects / Q000235:genetics
chemical_list: D006595:Hexosamines; D007328:Insulin; D016329:Sp1 Transcription Factor; C497388:Srebf1 protein, mouse; D051780:Sterol Regulatory Element Binding Protein 1; D003980:Diazooxonorleucine; D004247:DNA; D002784:Cholesterol; D006903:Hydroxymethylglutaryl CoA Reductases; D005947:Glucose