Figure 5. IRF1 cooperates with IRF2 to regulate non‐canonical inflammasome activation in PMA‐differentiated macrophages.

U937 cell lines were differentiated with PMA and primed with IFN‐γ.

• A
  Cell death was quantified by LDH assay 2 h after LPS electroporation.
• B
  Cells were primed with Pam3CSK4. IL‐1β levels were assessed by ELISA 4 h after LPS electroporation.
• C
  Cell death was quantified by measuring propidium iodide (PI) incorporation/fluorescence every 5 min after infection with F. novicida or (F) E. coli. Cell death was normalized using untreated and TX‐100‐treated cells. One real‐time cell death experiment (mean and SD of triplicate) representative of three independent experiments is shown. The areas under the curve (AUC) (normalized to the WT AUC) of three independent experiments are shown. Each point represents the mean of a triplicate of one experiment; the bar represents the mean (and SD) of the three independent experiments.
• (D–H)
  (D, G) IL‐1β and (E, H) TNF levels were assessed by ELISA 6 h after WT or ΔFPI mutant F. novicida (D, E) or E. coli (G, H) infection.

Data information: (A, B) Each dot corresponds to the mean of LDH triplicate of one experiment; the bar represents the mean ± SD of three independent experiments. (D–E, G–H) Each dot corresponds to the mean of biological triplicates of one experiment, the bar represents the mean ± SD of three (D, G, H) to four (E) independent experiments. (A–H) One‐way ANOVA with Dunnet's multiple comparisons test was performed. (A) WT vs. IRF1/2^KO **P = 0.0084, WT vs. CASP4^KO **P = 0.0019 (B) WT vs. IRF1^KO *P = 0.012, WT vs. IRF1/2^KO ***P = 0.0003, WT vs. CASP4^KO ***P < 0.0001 (C) (WT vs. IRF2^KO: P = 0.26, WT vs. IRF1^KO: **P = 0.0015, WT vs. IRF1/2^DKO: ***P = 0.0002, WT vs. CASP4^KO: **P = 0.0067). (D) (WT vs. IRF2^KO: ***P = 0.001, WT vs. IRF1^KO: ***P < 0.0001, WT vs. IRF1/2^DKO: ***P < 0.0001, WT vs. CASP4^KO: ***P = 0.0005). (E) (WT vs. IRF2^KO: P = 0.045, WT vs. IRF1^KO: *P = 0.19, WT vs. IRF1/2^DKO: P = 1, WT vs. CASP4^KO: P = 0.57). (G) (WT vs. IRF2^KO: *P = 0.016, WT vs. IRF1^KO: *P = 0.024, WT vs. IRF1/2^DKO: **P = 0.0032, WT vs. CASP4^KO: P = 0.57). (F) WT vs. IRF1^KO *P = 0.045, WT vs. IRF1/2^KO **P = 0.0017, WT vs. CASP4^KO *P = 0.0107 (H) (WT vs. IRF2^KO: P = 0.86, WT vs. IRF1^KO: P = 1, WT vs. IRF1/2^DKO: P = 0.79, WT vs. CASP4^KO: P = 1).Source data are available online for this figure.