Extended Data Fig. 6: Epithelial sensing of IFN-γ promotes eATPase production from IE-T cells.

(A-I) Ifngr1^fl/fl and Ifngr^fl/fl Vil^Cre mice were infected with Citrobacter rodentium. (A and B) RT-PCR analyses for indicated factors in colon epithelial tissue fraction (n = 4 for d0, d6 per group and n = 8 for Ifngr1^fl/fl and n = 6 for Ifngr1^fl/fl Vil^Cre for d12). (C, E-I) flow cytometric (n = 3 for d0, n = 5 for d6 and d12 per group) and (D) immunofluorescence evaluation of CD39 expression, representative of two independent experiments and (G) RT-PCR analyses of Entpd5 expression from the purified IE-T cells and IECs (n = 6 per group). (I) Ifngr1^fl/fl and Ifngr1^fl/fl Vil^Cre mice were infected with ova⁺ Citrobacter and the ova-specific CD8αβ⁺ and CD4⁺ IE-T cells were evaluated for CD39 expression and (J) comparison of GM-CSF and CD39 producing CD4⁺ colonic IE-T cells in Ifngr1^fl/fl mice at day 12 p.i. (K-M) Ifngr1^fl/fl and Ifngr1^fl/fl Vil^Cre mice were infected with Citrobacter rodentium and treated with 250ug isotype or CD8β-depleting antibodies every 2 days after the infection and sacrificed on day 12 p.i. (n = 8 per group). (K) FACS plot for CD8⁺ IE-T cells from mice given isotype or CD8β-depleting antibodies. (L) Colon explants were analyzed for indicated analytes and (M) colon length measurement. (N) Ifngr1^fl/fl, Ifngr1^fl/fl Vil^CreERT2 and Ifngr1^fl/fl Vil^CreERT2 R26^LSLIrf1 mice were infected with Citrobacter rodentium on day 0, injected with tamoxifen at days 3–6 post-infection and analyzed on day 12 for CD39 expression from CD8αβ⁺ and CD4⁺ IE-T cells (n = 5 per group). The horizontal bar represents the median, and each symbol represents an individual mouse. Data were analyzed by (A, B) by Brown-Forsythe and Welch’s ANOVA followed by Dunnett’s post-test or (E-I) Mann-Whitney U test (two-sided) or (L-N) Kruskal-Wallis test followed by Dunn’s post-test. Scale bar = 100 um.

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