Male and female Foxp3^YFP-Cre and Kdm6b^ΔTreg mice were analyzed at steady state or after PR8 influenza challenge on day 0 (2 μL/g body weight). (A–C) Lung Tregs from single-cell suspensions at steady state were analyzed for frequency of FOXP3⁺ Tregs among CD4⁺ lymphocytes (n = 11 mice/strain, ≥2 experiments) (A), FOXP3 expression (gMFI) (B), and Treg immunophenotype (n = 5–6 mice/strain, ≥2 experiments) (C). (D) Total lung cell counts from enzymatic digests at steady state (n = 11–12 mice/strain, ≥2 experiments). (E) Percent body weight change after PR8 influenza infection relative to baseline (n = 34–35 mice/strain/time point, pooled from ≥3 experiments). (F) Histopathology scores from H&E-stained lung sections in Foxp3^YFP-Cre (closed circles) or Kdm6b^ΔTreg mice (open circles) at steady state, day 9, or day 15 after PR8 influenza infection (n = 6–10 mice/strain/time point, ≥2 experiments). (G) Survival after PR8 influenza challenge (n = 40–41 mice/strain, ≥2 experiments). (H–N) Analyses were performed on day 15 after PR8 influenza infection. (H) Total lung cell counts. (I) Numbers of lymphocyte and NK subsets assessed using established gating strategies (38, 53) (n = 8–10 mice/strain, ≥2 experiments). (J) Lung Tregs quantified separately from (I). (K) Frequency of CD44⁺CD62L^– cells among CD4⁺, CD8⁺, or CD4⁺FOXP3⁺ T cells (n = 4–5 mice/strain). (L–N) (L) CD4⁺FOXP3⁺ cells from lung or spleen analyzed for FOXP3 expression (gMFI; n = 3–5 mice/strain), (M) proliferation (n = 6–10 mice/strain), and (N) lung Treg immunophenotyping (n = 4–5 mice/strain). Data are given as the mean ± SEM. Statistical tests: 2-way ANOVA with Holm-Šidák correction (C, F, I, and K–N), unpaired 2-tailed t test (A, B, D, H, and J), and log-rank test (G). *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001.