Fig. 4. Double knockout of Foxo3a and p53 fully protects cells from IL-3-induced apoptosis and prevents Puma induction.

a Ba/F3 p53^−/− Foxo3a^−/− single-cell clones (DKO #1 and DKO #2) and Ba/F3 expressing CRISPR/Cas9 constructs targeting Luciferase (crLUC) were deprived of IL-3 for 18 h and analyzed for apoptosis by Annexin V staining. Error bars represent 95% confidence intervals from four independent experiments (n = 4). Significance was tested by one-way ANOVA with post hoc Tukey’s multiple comparison test. ns = not significant. ***P < 0,001. b The same cells were treated with LY294002 (10 µM) for 18 h or left untreated. The cells were harvested after the treatment and subjected to western blotting. The protein levels were analyzed by the antibodies indicated. c The same cells were deprived of IL-3 for 18 h, harvested after the treatment and subjected to western blotting.