Negative Regulation of FOXP3 Expression by c-Rel O-GlcNAcylation.

O-GlcNAcylation is a reversible posttranslational protein modification that regulates fundamental cellular processes including immune responses and autoimmunity. Previously, we showed that hyperglycemia increases O-GlcNAcylation of the transcription factor, Nuclear Factor kappaB (NF-κB) c-Rel at serine residue 350 and enhances the transcription of the c-Rel-dependent proautoimmune cytokines interleukin-2 (IL-2), interferon gamma (IFNG), and granulocyte macrophage colony stimulating factor (GM-CSF) in T cells. c-Rel also plays a critical role in the transcriptional regulation of forkhead box P3 (FOXP3), the master transcription factor that governs development and function of Treg cells. Here we show that the regulatory effect of c-Rel O-GlcNAcylation is gene-dependent, and in contrast to its role in enhancing the expression of proautoimmune cytokines, it suppresses the expression of FOXP3. Hyperglycemia-induced O-GlcNAcylation-dependent suppression of FOXP3 expression was found in vivo in two mouse models of autoimmune diabetes; streptozotocin (STZ)-induced diabetes and spontaneous diabetes in non-obese diabetic (NOD) mice. Mechanistically, we show that both hyperglycemia-induced and chemically enhanced cellular O-GlcNAcylation decreases c-Rel binding at the FOXP3 promoter and negatively regulates FOXP3 expression. Mutation of the O-GlcNAcylation site in c-Rel, (serine 350 to alanine), augments T cell receptor-induced FOXP3 expression and resists the O-GlcNAcylation-dependent repression of FOXP3 expression. This study reveals c-Rel S350 O-GlcNAcylation as a novel molecular mechanism inversely regulating immunosuppressive FOXP3 expression and proautoimmune gene expression in autoimmune diabetes with potential therapeutic implications.