Figure 1: ASC is critical for caspase-1 autoproteolysis by known inflammasomes.

(a,b) BALB/c and B6 macrophages were treated with PA (10 μg) and/or wild-type LF (10 μg) and the enzymatically inactive LF_E687C mutant (LFmt) for 3 h, after which lysates were immunoblotted for caspase-1 (a) and culture supernatants analysed for LDH release (b). Data are shown as mean±s.d. from a single representative experiment of three experiments, with each condition performed in triplicate. (c–e) Lysates of B6, ASC^−/−, B6^Nlrp1b+ and B6^Nlrp1b+ASC^−/− BMDMs were immunoblotted for caspase-1 after cells have been primed with 5 μg ml⁻¹ LPS for 3 h and subsequently stimulated with 20 μM nigericin for 60 min (c), pretreated with 5 μg ml⁻¹ LPS (3 h) and subsequently transfected with 1 μg dsDNA for 24 h (d) or infected with S. typhimurium (m.o.i. 10) for 3 h (e). (f,g) B6, B6^Nlrp1b+, B6^Nlrp1b+ASC^+/− and B6^Nlrp1b+ASC^−/− BMDMs were left untreated or pretreated with 5 μg ml⁻¹ LPS for 3 h prior to being exposed to different concentrations of LeTx (f, 10 μg PA+10 μg LF) or (g, 500 ng PA+250 ng LF) for another 3 h. Cell lysates were immunoblotted for caspase-1. Data are representative of results from three experiments.