title: PUGNAc induces protein ubiquitination in C2C12 myotube cells.
abstract: O-linked β-N-acetylglucosaminylation (O-GlcNAcylation) regulates many cellular processes including the cell cycle, cell signaling, and protein trafficking. Dysregulation of O-GlcNAcylation may be involved in the development of insulin resistance and type 2 diabetes. Therefore, it is necessary to identify cellular proteins that are induced by elevated O-GlcNAcylation. Here, using adenosine 5'-triphosphate affinity chromatography, we employed a proteomic approach in order to identify differentially expressed proteins in response to treatment with the O-GlcNAcase inhibitor, O-(2-acetamido-2-deoxy-d-glucopyranosylidene)amino-N-phenylcarbamate (PUGNAc), in mouse C2C12 myotube cells. Among 205 selected genes, we identified 68 nucleotide-binding proteins, 14 proteins that have adenosinetriphosphatase activity, and 10 proteins with ligase activity. Upregulation of proteins, including ubiquitin-activating enzyme E1, proteasome subunit 20S, cullin-associated NEDD8-dissociated protein 1, ezrin, and downregulation of the protein nucleoside diphosphate kinase B, were confirmed by western blot analysis. In particular, we found that the protein ubiquitination level in C2C12 cells was increased by PUGNAc treatment. This is the first report of quantitative proteomic profiles of myotube cells after treatment with PUGNAc, and our results demonstrate the potential to enhance understanding of the relationship between insulin resistance, O-GlcNAc, and PUGNAc in the future.
authors.affiliation: Department of Molecular Medicine, Kyungpook National University School of Medicine, Daegu, Republic of Korea.|Cell and Matrix Research Institute, Kyungpook National University School of Medicine, Daegu, Republic of Korea.
authors.forename: Ja-Hye
authors.initials: JH
authors.lastname: Park
authors.affiliation: Department of Molecular Medicine, Kyungpook National University School of Medicine, Daegu, Republic of Korea.|Cell and Matrix Research Institute, Kyungpook National University School of Medicine, Daegu, Republic of Korea.
authors.forename: Jeong-Eun
authors.initials: JE
authors.lastname: Lee
authors.affiliation: Department of Molecular Medicine, Kyungpook National University School of Medicine, Daegu, Republic of Korea.|Cell and Matrix Research Institute, Kyungpook National University School of Medicine, Daegu, Republic of Korea.
authors.forename: Pyong-Gon
authors.initials: PG
authors.lastname: Moon
authors.affiliation: Department of Molecular Medicine, Kyungpook National University School of Medicine, Daegu, Republic of Korea.|Cell and Matrix Research Institute, Kyungpook National University School of Medicine, Daegu, Republic of Korea.
authors.forename: Moon-Chang
authors.initials: MC
authors.lastname: Baek
mesh_terms: D000117:Acetylglucosamine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D005960:Glucosides / Q000378:metabolism; D051379:Mice; D058977:Molecular Sequence Annotation; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000378:metabolism*; D010091:Oximes / Q000494:pharmacology*; D048448:Phenylcarbamates / Q000494:pharmacology*; D011485:Protein Binding / Q000187:drug effects; D020543:Proteome / Q000378:metabolism; D040901:Proteomics; D013194:Staining and Labeling; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination / Q000187:drug effects*
chemical_list: D005960:Glucosides; D010091:Oximes; D048448:Phenylcarbamates; D020543:Proteome; D025801:Ubiquitin; C068836:N-acetylglucosaminono-1,5-lactone O-(phenylcarbamoyl)oxime; D000255:Adenosine Triphosphate; D000117:Acetylglucosamine
keywords: ATP-affinity chromatography; C2C12 myotube cell; O-GlcNAc; PUGNAc; proteomics; ubiquitination