Alphavirus-induced p38 activity correlates with NLRP1 activation. (A and B) HEK^NLRP1+ASC, HEK^NLRP3+ASC (A), or N/TERT-1^C1C-EGFP (B) cells were infected with SFV (A: MOI 1/B: MOI 5), EEEV (MOI 1/5), VEEV (MOI 1/50), CHIKV (MOI 1/50), MAYV (MOI 25/50), ONNV (MOI 25/50), BFV (MOI 25/50), or RRV (MOI 25/50) for 20 h in the absence or presence of 20 µM SB. Infected cells were stained with antibodies for dsRNA and infection and speck assembly in infected cells were quantified by flow cytometry as in Fig. 4. (C and D) HEK^NLRP1+ASC cells were infected with the indicated viruses as in A. Control cells were infected with VSV at an MOI of five or treated with 15 µM Aniso. Infected cells were stained with antibodies for dsRNA or VSV G, and P-p38 in infected cells (total cells in case of uninfected samples) was quantified by flow cytometry. Average fractions of P-p38–positive cells from three independent experiments (C) and representative histograms (D) are displayed. N/TERT-1 cells were infected in the presence of 100 µM VX for all flow cytometry experiments. Data represents average values (with individual data points) from three independent experiments ± SEM.