Temporal regulation of natural Killer T cell interferon gamma responses by β-catenin-Dependent and -Independent Wnt signaling

Natural killer T (NKT) cells are prominent innate-like lymphocytes in the liver with critical roles in immune responses during infection, cancer, and autoimmunity. Interferon gamma (IFN-alpha) and IL-4 are key cytokines rapidly produced by NKT cells upon recognition of glycolipid antigens presented by antigen-presenting cells (APCs). It has previously been reported that the transcriptional coactivator beta-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-alpha production. beta-Catenin is not only a central effector of Wnt signaling but also contributes to other signaling networks. It is currently unknown whether Wnt ligands regulate NKT cell functions. We thus investigated how Wnt ligands and beta-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, alpha-galactosylceramide (alpha-GalCer) using a mouse model. Pharmacologic targeting of beta-catenin activity with ICG001, as well as myeloid-specific genetic ablation of Wntless (Wls), to specifically target Wnt protein release by APCs, enhanced early IFN-alpha responses. By contrast, within several hours of alpha-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired alpha-GalCer-induced IFN-alpha responses, independent of beta-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/beta-catenin signaling that curbs IFN-alpha responses, but that, subsequently, Wnt ligands sustain IFN-alpha expression independent of beta-catenin activity. Our analyses in ICG001-treated mice confirmed a role for beta-catenin activity in driving early IL-4 responses by liver NKT cells. However, neither pharmacologic nor genetic perturbation of Wnt production affected the IL-4 response, suggesting that IL-4 production by NKT cells in response to alpha-GalCer is not driven by released Wnt ligands. Collectively, these data reveal complex temporal roles of Wnt ligands and beta-catenin signaling in the regulation of liver NKT cell activation, and highlight Wnt-dependent and -independent contributions of beta-catenin to NKT cell functions.