Elevated O‐GlcNAc Exacerbates Pro‐Inflammatory Cytokine Secretion from CD4+ T cells

Abstract

Chronic inflammation is a feature of obesity and enhances the risk of atherosclerosis, cancer, diabetes, and autoimmunity. Specifically, pro-inflammatory TH17 CD4+ effector T cells are increased in metabolic diseases. However, a clear molecular mechanism linking metabolic changes with pro-inflammatory T cells is lacking. We hypothesize that elevated levels of O-linked β-N-acetylglucosamine (O-GlcNAc), a post-translational modification of nuclear and cytoplasmic proteins, promotes pro-inflammatory CD4+ T cell function. Since production of O-GlcNAc involves input from carbohydrate, amino acid, fatty acid, and nucleic acid metabolism, the modification acts as a sensor of a cell's nutritional status. To investigate the role of O-GlcNAc in a setting of metabolic disease, we analyzed O-GlcNAc levels in CD4+ T cells from mice fed a high fat and cholesterol “Western” diet. Naïve CD4+ T cells from obese mice have elevated O-GlcNAc levels compared to cells from mice fed standard chow. When polarized to a TH17 lineage, cells from obese mice secrete more IL-17A, the eponymous TH17 cytokine. Importantly, when naïve CD4+ T cells from lean and obese mice are polarized to a TH17 lineage in the presence of Thiamet-G (TMG, an inhibitor of the enzyme that removes O-GlcNAc (O-GlcNAcase, OGA)) the cells from lean mice secrete levels of IL-17 comparable to those from obese mice and IL-17 secretion was exacerbated in cells from obese mice. Transcript levels of IL-17 are similarly elevated in both lean and obese mice TH17 cells treated with TMG. RORγt (retinoic acid-related orphan receptor gamma) acts as the TH17 master transcription factor, orchestrating the TH17 differentiation program. Uniquely among the CD4+ effector T cell master transcription factors, RORγt is regulated by fatty acid ligands which promote or repress its activity. Acetyl CoA carboxylase 1 (ACC1), the rate limiting enzyme in fatty acid synthesis, is known to enhance generation of TH17 cells by producing ligands that increase the activity of RORγt. We discovered that ACC1 is O-GlcNAcylated. Furthermore, lipidomics analysis identified clear differences in the lipid environment of T cells isolated from lean and obese mice −/+ TMG. Thus, abnormally elevated O-GlcNAc levels may favor the generation of ligands that enhance RORγt activity at the IL-17 promoter. More directly implicating O-GlcNAc's regulation of RORγt, chromatin immunoprecipation of RORγt and O-GlcNAc transferase (OGT, the enzyme that modifies proteins with O-GlcNAc) at the IL-17 promoter and an enhancer, CNS2 (conserved noncoding sequence 2), is increased with TMG treatment. Collectively, our data suggest that elevated O-GlcNAc levels increase pro-inflammatory IL-17 secretion from TH17 cells through alteration of the lipidome and direct effects on the IL-17 promoter complex. Further study into the molecular mechanism of how O-GlcNAcylation promotes pro-inflammatory TH17 T cell function will provide insight into how nutritional excess worsens inflammation. Support or Funding Information This research was supported by NIH grant R01DK100595-01, NIH grant R01DK091277-03, COBRE P30GM122731, and a KUMC Institute of Reproductive Health and Regenerative Medicine pilot grant. MM is supported by a KUMC Biomedical Research Training Program grant. Elevated O-GlcNAc increases pro-inflammatory IL-17 cytokine secretion from TH17 cells This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.