Caspase-11 auto-processing at Asp₂₈₅ is essential for GSDMD processing, pyroptosis, and IL-1β release. (A–D) BMDMs were treated with 1 µg/ml Pam3CSK4 for 5 h. (A) Immunoblot of caspase-11, GSDMD, and actin in extract (ext) + supernatant (sup) from WT, Casp11^{Enz C254A/C254A}, and Casp11^{Prc D285A/D285A} BMDMs at 60 min after LPS electroporation or control. (B) Cell death measured by percent YOYO-1⁺ cells from live cell images taken every 30 min over a 16-h time course following LPS electroporation of BMDMs from two mice per genotype. (C) LDH, IL-1β, and IL-18 release from BMDMs stimulated with transfected LPS (5 µg/ml plus FuGENE HD), ATP (5 mM), or nigericin (10 µg/ml) after 16 h, 3 h, or 30 min, respectively. (D) LDH, IL-1β, and IL-18 release measured from supernatants of BMDMs infected with indicated strains of bacteria. Data are representative of at least two independent experiments (A and B). Data are presented as mean ± SD (n = 3) and representative of at least three independent experiments (C and D).