TGF-β signaling is continuously needed to inhibit differentiation of antigen-specific CD8 T cells during chronic LCMV infection. (A) Rosa26Cre-ERT2⁺ TGF-βRII^fl/fl (Tgfbr2^fl/fl) and Rosa26Cre-ERT2⁺ TGF-βRII^+/+ (WT) P14 cells were cotransferred into naive B6 mice with CD4 T cell depletion and were then infected with LCMV clone 13. Recipient mice were treated with TAM or vehicle for 5 consecutive days (D31–D35 p.i.) to induce the deletion of TGF-βRII. Donor cells were harvested from spleen 3 wk after TAM treatment. (B and C) Donor cells were harvested from spleen on D60 p.i. Representative FACS plots show the expression of Tim3 and TCF-1 (B) or Ki67 and TCF-1 (C) by Tgfbr2^fl/fl and WT P14 cells from mice treated with vehicle or TAM. (D–G) Plots show frequencies of TCF-1⁺ Tim3⁻ (D), Ki67⁺ (E), Tim3⁺ CD101⁻ (F), and CX3CR1⁺ TCF-1⁻ (G) cells among WT and Tgfbr2^fl/fl P14 cells in the spleen. In D–G, paired Student’s t test; *, P < 0.05. **, P < 0.01. Data in B–G are representative of two independent experiments (n = 5–6 each time point per experiment).