REV-ERBα agonists reduce protein O-GlcNAcylation. REV-ERBα agonists reduce protein O-GlcNAcylation. (A) Protein O-GlcNAcylation, OGT, and REV-ERBα levels after treatment of HepG2 cells with the synthetic REV-ERBα agonist GSK4112 (10 µM). (B) REV-ERBα repressive activity determined by a transactivation assay using a Bmal-luc reporter gene, expression vectors coding for either wild-type or mutated REV-ERBα (WT, H602F) or empty vector (empty) transfected into HEK293 cells treated or not with 10 µM GSK4112. (C) O-GlcNAcylation, OGT, and REV-ERBα protein levels in HEK293 whole-cell extracts after transfection with REV-ERBα WT or H602F and treatment or not with 10 µM GSK4112 at 25 mM Glc. Results are expressed as mean ± SEM, and values were compared by a two-way ANOVA followed by a Tukey post hoc test, ****P < 0.0001. Wes, Simple Western (ProteinSimple). Alexandre Berthier et al. PNAS 2018;115:47:E11033-E11042 ©2018 by National Academy of Sciences