Fig.1: Ezh2 is required for CD8⁺ T cells to control tumor growth and promote memory precursor formation

. a WT and Ezh2 ^−/− naive Pmel-1 cells (1 × 10⁶, Thy1.1⁺) were transferred into sublethally irradiated (5 Gy) B6 mice (Thy1.2⁺) that had pre-established B16 melanoma, followed by treatment with IL-2 (1 × 10⁵ IU per injection, i.p., twice a day) and gp100-pulsed DCs (gp100-DCs, 1 × 10⁶ per mouse, i.p.) for 3 days. Tumor size was monitored over time. b–f WT and Ezh2 ^−/− T_N Pmel-1 cells (1 × 10⁶, Thy1.1⁺) were transferred into sublethally irradiated non-tumor-bearing B6 mice, followed by immunization with IL-2 and gp100-DCs for 3d. b Donor T cells were collected from the spleen 4 days, 7 days, and 35 days after adoptive transfer. Plots and graphs show the frequency and numbers of donor T cells. c Percentage of IFN-γ-producing donor T cells in the spleen. d Donor T cells were collected from the spleen of WT and Ezh2 ^−/− Pmel-1 cell primary recipients 42 days after transfer, and separately transferred into sublethally irradiated secondary non-tumor-bearing B6 mice (4 × 10⁴ cells per mouse), followed by treatment with IL-2 and gp100-DCs at 42 days, 43 days, and 44 days. By 49 days, donor T cells were collected from the spleen of the secondary mice. Plots and graph show the percentage of donor Pmel-1 cells. e Donor T cells derived from these secondary mice were activated with anti-CD3 Ab for 5 hrs to measure their production of IFN-γ. Graph shows the number of IFN-γ⁺ Pmel-1 cells in the spleen. f Donor T cells collected at 49d from the secondary mice were cultured ex vivo with IL-7 + IL-15 in the presence or absence of gp100 for additional 5 days. Plots and graphs show the percentage of donor T cells in cultures. *p < 0.05, **p < 0.01, and ***p < 0.001 (two-tailed unpaired t test). The data are representatives of three independent experiments with n = 5 mice per group in each (a; mean ± SD), or two experiments (b–f, n = 3–6 mice per group in each, mean ± SD)