ConclusionAlthough orexin receptor antagonists and agonists have been examined as drug candidates for various diseases such as narcolepsy, depression, and metabolic disorders, the direction of our research is distinct. Our study revealed that the HCRT gene was silenced by the hyperglycemia condition, and ManNAc and its derivatives were useful for restoring the orexin neurons. AcknowledgmentWe thank Drs. Koji Hayakawa, Osamu Kanie, and Yasuharu Sakamoto for their valuable contribution to this project.Creation of carbohydrate derived epigenetic controlling moleculesYukishige Ito, Kunio ShiotaFigure 3. Metabolism of ManNAc derivatives analyzed by Nano LC-MS COS-7 cells were incubated with Lac-Sph-BODIPY (fluorescent glycolipid substrate)-BSA complex and ManNAc, ManNFAc, or 5S-ManNAc. Total lipid extracts were analyzed by Nano LC-MS and the formation of GM3-BODIPY was detected by MS.orexin neurons to high glucose in culture caused irreversible silencing of the HCRT gene, which was characterized by H3/H4 hypoacetylation and hyper-O-GlcNAcylation. The DNA hypomethylation status, once established in orexin neurogenesis, was maintained in the HCRT-silenced orexin neurons, indicating that histone modifications, but not DNA methylation, were responsible for the HCRT silencing. Thus, the epigenetic status of the HCRT gene is unique to the hyperglycemia-induced silencing. Intriguingly, the treatment of ManNAc and its derivatives reactivated HCRT gene expression, while inhibitors SIRT1 and the OGT did not4).References1) Hirosawa M, Hayakawa K, Yoneda C, Arai D, Shiota H, Suzuki T, Tanaka S, Dohmae N, Shiota K, Novel O-GlcNAcylation on Ser 40 of canonical H2A isoforms specific to Viviparity: Sci Rep (6):31785, 2016. DOI: 10.1038/srep317852) Hayakawa K, Hirosawa M, Tani R, Yoneda C, Tanaka S, Shiota K. H2A O-GlcNAcylation at serine 40 functions genomic protection in association with acetylated H2AZ or γH2AX: Epigenetics & Chromatin (10): 51-64, 2017. DOI: 10.1186/s13072-017-0157-x3) Hayakawa K, Hirosawa M, Tabei Y, Arai D, Tanaka S, Murakami N, Yagi S, Shiota K. Epigenetic switching by the metabolism-sensing factors in the generation of orexin neurons from mouse embryonic stem cells. J Biol Chem (288): 17099-17110, 2013. DOI: 10.1074/jbc.M113.4558994) Hayakawa K, Sakamoto Y, Kanie O, Ohtake A, Daikoku S, Ito Y, Shiota K. Reactivation of hyperglycemia-induced hypocretin (HCRT) gene silencing by N-acetyl-D-mannosamine in the orexin neurons derived from human iPS cells. Epigenetics (12): 764–778, 2017. DOI: 10.1080/15592294.2017.1346775.5) Ohtake A, Daikoku S, Suzuki K, Ito Y, Kanie O. Analysis of the cellular dynamics of fluorescently tagged glycosphingolipids by using a nanoliquid chromatography-tandem mass spectrometry platform. Anal Chem (85): 8475-8482, 2013. DOI:10.1021/ac401632t52
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