title: Inducible nitric-oxide synthase and nitric oxide donor decrease insulin receptor substrate-2 protein expression by promoting proteasome-dependent degradation in pancreatic beta-cells: involvement of glycogen synthase kinase-3beta.
abstract: Insulin receptor substrate-2 (IRS-2) plays a critical role in the survival and function of pancreatic β-cells. Gene disruption of IRS-2 results in failure of the β-cell compensatory mechanism and diabetes. Nonetheless, the regulation of IRS-2 protein expression in β-cells remains largely unknown. Inducible nitric-oxide synthase (iNOS), a major mediator of inflammation, has been implicated in β-cell damage in type 1 and type 2 diabetes. The effects of iNOS on IRS-2 expression have not yet been investigated in β-cells. Here, we show that iNOS and NO donor decreased IRS-2 protein expression in INS-1/832 insulinoma cells and mouse islets, whereas IRS-2 mRNA levels were not altered. Interleukin-1β (IL-1β), alone or in combination with interferon-γ (IFN-γ), reduced IRS-2 protein expression in an iNOS-dependent manner without altering IRS-2 mRNA levels. Proteasome inhibitors, MG132 and lactacystin, blocked the NO donor-induced reduction in IRS-2 protein expression. Treatment with NO donor led to activation of glycogen synthase kinase-3β (GSK-3β) and c-Jun N-terminal kinase (JNK/SAPK) in β-cells. Inhibition of GSK-3β by pharmacological inhibitors or siRNA-mediated knockdown significantly prevented NO donor-induced reduction in IRS-2 expression in β-cells. In contrast, a JNK inhibitor, SP600125, did not effectively block reduced IRS-2 expression in NO donor-treated β-cells. These data indicate that iNOS-derived NO reduces IRS-2 expression by promoting protein degradation, at least in part, through a GSK-3β-dependent mechanism. Our findings suggest that iNOS-mediated decreased IRS-2 expression may contribute to the progression and/or exacerbation of β-cell failure in diabetes.
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Tanioka
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Tamura
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Makiko
authors.initials: M
authors.lastname: Fukaya
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Shohei
authors.initials: S
authors.lastname: Shinozaki
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Ji
authors.initials: J
authors.lastname: Mao
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Minhye
authors.initials: M
authors.lastname: Kim
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and.
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Shimizu
authors.affiliation: Metabolic Signal Research Center, Institute for Molecular and Cellular Regulation, Gunma University, Gunma 371-8512, Japan.
authors.forename: Tadahiro
authors.initials: T
authors.lastname: Kitamura
authors.affiliation: Department of Anesthesia, Critical Care, and Pain Medicine, Massachusetts General Hospital, Shriners Hospitals for Children, Harvard Medical School, Charlestown, Massachusetts 02129 and. Electronic address: mkaneki@helix.mgh.harvard.edu.
authors.forename: Masao
authors.initials: M
authors.lastname: Kaneki
mesh_terms: D000111:Acetylcysteine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D000873:Anthracenes / Q000494:pharmacology; D045744:Cell Line, Tumor; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D055504:Insulin Receptor Substrate Proteins / Q000096:biosynthesis* / Q000235:genetics; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000378:metabolism*; D007371:Interferon-gamma / Q000235:genetics / Q000378:metabolism; D053583:Interleukin-1beta / Q000235:genetics / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D051379:Mice; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000235:genetics / Q000378:metabolism*; D061988:Proteasome Inhibitors; D051381:Rats
chemical_list: D000873:Anthracenes; D015853:Cysteine Proteinase Inhibitors; C526297:IRS2 protein, human; D055504:Insulin Receptor Substrate Proteins; D053583:Interleukin-1beta; C526298:Irs2 protein, mouse; C526299:Irs2 protein, rat; D007976:Leupeptins; D020030:Nitric Oxide Donors; D061988:Proteasome Inhibitors; C067713:lactacystin; C432165:pyrazolanthrone; D007371:Interferon-gamma; C496319:NOS2 protein, human; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; C496318:Nos2 protein, rat; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; C000605508:Gsk3b protein, rat; D048031:JNK Mitogen-Activated Protein Kinases; D038362:Glycogen Synthase Kinase 3; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; D000111:Acetylcysteine