Abstract
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-γ) 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 β-catenin regulates NKT cell differentiation and functionally biases NKT cell responses toward IL-4, at the expense of IFN-γ production. β-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 β-catenin activity shape liver NKT cell functions in vivo in response to the glycolipid antigen, α-galactosylceramide (α-GalCer) using a mouse model. Pharmacologic targeting of β-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-γ responses. By contrast, within several hours of α-GalCer challenge, myeloid-specific Wls deficiency, as well as pharmacologic targeting of Wnt release using the small molecule inhibitor IWP-2 impaired α-GalCer-induced IFN-γ responses, independent of β-catenin activity. These data suggest that myeloid cell-derived Wnt ligands drive early Wnt/β-catenin signaling that curbs IFN-γ responses, but that, subsequently, Wnt ligands sustain IFN-γ expression independent of β-catenin activity. Our analyses in ICG001-treated mice confirmed a role for β-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 α-GalCer is not driven by released Wnt ligands. Collectively, these data reveal complex temporal roles of Wnt ligands and β-catenin signaling in the regulation of liver NKT cell activation, and highlight Wnt-dependent and -independent contributions of β-catenin to NKT cell functions.
Highlights
Natural killer T (NKT) cells are an important population of hepatic lymphocytes in both humans and mice [1]
Studies on contributions of β-catenin activity and Wnt ligands to NKT cell functions in vivo have been hampered by the impact of genetic modulation of β-catenin expression on NKT cell development, as well as the limited availability of animal models suitable to assess Wnt functions in the adult organism in vivo
Through pharmacologic and conditional genetic targeting of β-catenin activity and key factors that drive Wnt release, collectively the data presented in this study support the conclusions that in response to the NKT cell antigen α-GalCer: (i) Wnt/β-catenin signaling in the liver environment regulates interferon gamma (IFN-γ) responses; (ii) active suppression of hepatic Wnt/β-catenin signaling shortly after α-GalCer exposure biases cytokine responses toward IFN-γ production; (iii) β-catenin activity independent of antigen-presenting cell (APC)-derived Wnt ligands contributes to IL-4 expression by NKT cells; (iv) Wnt ligands that act via β-catenin-independent signaling contribute to the perpetuation of IFN-γ responses within several hours post challenge; and (v) myeloid-derived Wnt ligands are, in part, responsible for these regulatory effects on NKT cells
Summary
Natural killer T (NKT) cells are an important population of hepatic lymphocytes in both humans and mice [1]. Hepatic Wnt proteins are central regulators of cell proliferation, differentiation, and functionality during liver injury, repair, regeneration, as well as homeostasis [6, 7] Their roles are complex and often context dependent. Stimulation of β-catenin-overexpressing NKT cells in vitro and in vivo resulted in decreased IFN-γ expression, and increased IL-4, IL-13, and IL-17 production, consistent with the effects on the development of NKT cell subsets in these mice [20]. These observations strongly suggest key roles for β-catenin and its interaction partners, TCF and LEF, in the development and functions of NKT cells. We employed pharmacologic and genetic perturbation of Wnt production and β-catenin to investigate how β-catenin activity and Wnt ligands shape NKT cell cytokine responses in vivo using the model antigen, α-galactosylceramide (α-GalCer)
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