Fig. 1. Differential ICOS and CD122 expression in MAIT1 and MAIT17 effector cells.

a–i Thymocytes from 8–10 weeks old WT mice were enriched for MAIT cells with 5-OP-RU-loaded MR1-Tet/magnetic beads and then stained with anti-TCRβ and other lineage antibodies (CD19, PBS-57-loaded CD1d-Tet, B220, CD11b, CD11c, F4/80, Ter119, Gr1, and TCRγδ). a Representative FACS plots showing gating strategies for MAIT cells and defining MAIT cell stages. b ICOS expression in stages 1–3 MAIT cells. c CD122 and ICOS expression, as well as T-bet and RORγt expression in stages 1–3 MAIT cells. d T-bet and RORγt expression in CD122⁺ICOS^low and CD122^-ICOS⁺ MAIT cells. e CD122 vs T-bet and ICOS vs RORγt staining of stage 3 thymic MAIT cells. f Representative FACS plots showing IFN-γ and IL-17A staining in MAIT cells after PMA plus ionomycin stimulation. Scatter plots show percentages of IFN-γ and IL-17A positive cells with the indicated MAIT subsets. g tSNE analysis of scRNAseq data from WT thymic MAIT cells. h Overlaid histograms show IRF4, cMAF, and BATF levels in thymic MAIT1, MAIT17, and non-MAIT TCRβ⁺ T cells. i Percentages of CD122⁺T-bet⁺ MAIT1 and ICOS⁺RORγt⁺ MAIT17 cells in different organs. j, k Analyses of Tbx21^f/f-Cd4Cre and control mice. j Representative FACS plots showing CD122 vs ICOS and T-bet vs RORγt staining in gated liver MAIT cells from WT and Tbx21^f/f-Cd4Cre mice. k Percentages of CD122⁺ICOS^low, CD122⁻ICOS⁺, RORγt⁻T-bet⁺, and RORγt⁺T-bet⁻ cells within control (red circle) and T-bet deficient (blue square) live MAIT cells. Each connection line indicates one pair of control and T-betKO mice examined in one experiment. Data shown were representative of or pooled from at least four experiments. Statistical significance is determined by two-tail unpaired (f, i) and pairwise (k) Student’s t test. P values of less than 0.05 are shown. All scatter and bar blots in this manuscript are presented as mean ± SEM. Source data for all graphs are provided as a Source Data file.