Figure 4: Type I IFNs regulate innate IFNγ production by CD8⁺ T cells.

(a,b) Splenocytes from WT and IRF9^−/− mice were cultured for 16 h with or without IL-12+IL-18. Brefeldin A (5 μg ml⁻¹) was added for the last 3 h of culture. (a) Dot plots of IFNγ and Eomes expression among total CD8⁺ T cells from WT and IRF9^−/− mice are illustrated. (b) Frequency of IFNγ⁺ cells among total and memory CD44⁺ CD8⁺ T cells in WT and IRF9^−/− mice. Data shown are representative of at least three independent experiments. (c) WT and IRF9^−/− mice were infected with Listeria monocytogenes 1 day before killing. Splenocytes were isolated and cultured for 4 h in presence of Brefeldin A before intranuclear staining and analysis by flow cytometry. Total CD8⁺ T cells, CD44⁺CD8⁺ T cells and NK cells were analysed for IFNγ expression. (d,e) Mixed bone marrow chimera experiments. WT Thy1.1⁺ and IRF9^−/− Thy1.2⁺ (d) or WT CD45.1 and IFNAR^−/− CD45.2 (e) myeloid progenitors were transferred into the irradiated RAG2^−/− host. After 3 months, the frequency of CD44⁺CD49d^lo CD8⁺ T cells among WT and IRF9^−/− or IFNAR^−/− CD8⁺ T cells was analysed by flow cytometry. Eomes expression levels are represented among WT and IRF9^−/− or IFNAR^−/− VM CD8⁺ T cells. Right panel: splenocytes from the chimeras were cultured for 16 h with IL-12+IL-18. Brefeldin A (5 μg ml⁻¹) was added for the last 3 h of culture. The frequency of IFNγ⁺ cells among WT and IRF9^−/− or IFNAR^−/− total CD8⁺ T cells are shown. For all graphs, each dot represents an individual mouse. Bars represent mean±s.e.m. *P<0.05, **P<0.01 and ***P<0.001, NS: not significant (non-parametric Mann–Whitney or paired Wilcoxon test for mixed bone marrow chimera experiments).