Modification of RelA by O-linked N-acetylglucosamine links glucose metabolism to NF-κB acetylation and transcription

• Published in: Proceedings of the National Academy of Sciences - PNAS 2012-10, Vol.109 (42), p.16888-16893
• Main Authors: Allison, David F, Wamsley, J. Jacob, Kumar, Manish, Li, Duo, Gray, Lisa G, Hart, Gerald W, Jones, David R, Mayo, Marty W
• Format: Article
• Language: English
• Subjects: Acetylation; Acetylglucosamine - metabolism; Biological Sciences; biosynthesis; Cell lines; cell viability; Cells; chromatin; Chromatin Immunoprecipitation; Cultured cells; DNA, Complementary - genetics; Enzyme-Linked Immunosorbent Assay; etoposide; Etoposide - metabolism; Gene expression; Gene Expression Regulation - physiology; Genes; glucose; Glucose - metabolism; HEK293 Cells; Hexosamines - biosynthesis; Histones; Humans; Immunoblotting; Immunoprecipitation; Luciferases; mammals; Metabolic Networks and Pathways - genetics; Metabolic Networks and Pathways - physiology; mutants; N-acetylglucosamine; NF-kappa B - metabolism; Plasmids - genetics; precipitin tests; Real-Time Polymerase Chain Reaction; Stem cells; transcription factor NF-kappa B; Transcription Factor RelA - genetics; Transcription Factor RelA - metabolism; Transcription factors; Tumor Necrosis Factor-alpha - metabolism; tumor necrosis factors
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1208468109

The molecular mechanisms linking glucose metabolism with active transcription remain undercharacterized in mammalian cells. Using nuclear factor-κB (NF-κB) as a glucose-responsive transcription factor, we show that cells use the hexosamine biosynthesis pathway and O-linked β- N -acetylglucosamine (O -GlcNAc) transferase (OGT) to potentiate gene expression in response to tumor necrosis factor (TNF) or etoposide. Chromatin immunoprecipitation assays demonstrate that, upon induction, OGT localizes to NF-κB–regulated promoters to enhance RelA acetylation. Knockdown of OGT abolishes p300-mediated acetylation of RelA on K310, a posttranslational mark required for full NF-κB transcription. Mapping studies reveal T305 as an important residue required for attachment of the O -GlcNAc moiety on RelA. Furthermore, p300 fails to acetylate a full-length RelA(T305A) mutant, linking O- GlcNAc and acetylation events on NF-κB. Reconstitution of RelA null cells with the RelA(T305A) mutant illustrates the importance of this residue for NF-κB–dependent gene expression and cell survival. Our work provides evidence for a unique regulation where attachment of the O- GlcNAc moiety to RelA potentiates p300 acetylation and NF-κB transcription.