NFAT1 regulates the acute phase of IFN-γ production in T-helper cells.

(A) Reduced IFN-γ production by freshly isolated NFAT1^−/− IL-4^−/− T-helper cells in response to stimulation with anti-CD3 but not IL-12/IL-18. NFAT1^+/+IL-4^−/− (▪) and NFAT1^−/−IL-4^−/− (▨) CD4⁺ T cells were stimulated either with plate-bound anti-CD3 (left panel) or a combination of IL-12 and IL-18 (right panel) for 48 hours, after which IFN-γ was analyzed in the supernatants of the cells. Shown is a representative of 3 (left panel) or 5 (right panel) independent experiments. (B) The selective NFAT inhibitor GFP-VIVIT suppresses IFN-γ production by a murine T-cell clone. Cl.7W2 cells were transfected with plasmids encoding either GFP or GFP-VIVIT. The next day the cells were either left unstimulated or were stimulated for 5 hours with PMA and ionomycin. IFN-γ production was analyzed at a single-cell level by intracellular cytokine staining. Dot blots from 1 of 3 similar experiments are shown. Thresholds for IFN-γ production were set using the unstimulated control. Note that each panel shows transfected (GFP-positive) and nontransfected (GFP-negative) cells of the same sample.