Inflammasomes are cytosolic protein complexes that activate caspase-1, an inflammatory protease that converts pro-interleukin (IL)-1β and IL-18 into their secreted, bioactive forms 1; and mechanisms regulating inflammasome activation have attracted interest because aberrant caspase-1 activity is implicated in diverse inflammatory and neurodegenerative diseases 2. Saleh et al. 3, 4 reported that caspase-12 acts as a dominant-negative regulator of caspase-1 activation and that caspase-12 deletion in splenocytes enhanced secretion of IL-1β and IL-18. We sought to confirm and further explore the role of caspase-12 in inflammasome inhibition, however, analysis of three independently generated caspase-12−/− mouse strains has brought into question the notion that caspase-12 is a physiologic inhibitor of caspase-1 activation. In contrast to the findings of Saleh et al. 3, 4, we further show that reported caspase-12-deficient mice also lack caspase-11 expression, and this prevented non-canonical caspase-1 activation. Host-and pathogen-encoded CARD-and pyrin-only proteins inhibit inflammasome activation 5. Saleh et al. 3, 4 proposed caspase-12 as an additional dominant-negative regulator that dampens caspase-1 activation and inhibits secretion of IL-1β and IL-18. Using overexpression in HEK293T cells and recombinant rodent caspase-12, they found that it physically interacted with caspase-1, and inhibited caspase-1 protease activity 3, 4. Saleh et al. 3, 4 also showed that overexpressed caspase-12 inhibited ATP-induced secretion of IL-1β from LPS-primed, Pam3CSK4-primed and poly (I: C)-primed THP-1 cells. In addition, they proposed that caspase-12-deficient splenocytes secreted increased IL-1β levels when stimulated with LPS and ATP ex vivo 3. From these results, they suggested that caspase-12 represses IL-1β secretion by inhibiting caspase-1 activation. The therapeutic potential of caspase-1 inhibition prompted us to confirm and explore the role of caspase-12 in modulating inflammasome signalling. Myeloid cells are the prominent source of the caspase-1-dependent cytokines IL-1β and IL-18, and several inflammasomes that activate caspase-1 in macrophages have been described 2. Surprisingly however, LPS-primed bone-marrow-derived macrophages (BMDMs) from the caspase-12-deficient mice described by Saleh et al. 3, 4 (referred to hereafter as caspase-12−/− s) secreted normal levels of ATP-and nigericin-induced IL-1β and IL-18 (Fig. 1a, b). Caspase-1 maturation and pyroptosis induction by these Nlrp3 inflammasome-activating agents also were normal (Fig. 1c, d). We similarly failed to observe increased IL-1β and IL-18 secretion, caspase-1 maturation and pyroptosis induction by the Nlrc4 (Fig. 1e–h) and AIM2 (Fig. 1i–l) inflammasomes upon Salmonella enterica serovar Typhimurium (S. Typhimurium) infection and double-stranded DNA (dsDNA) transfection, respectively. Caspase-11 is dispensable for the inflammasome stimuli above, but it is required for Escherichia coli-and Citrobacter rodentium-induced caspase-1 activation and downstream inflammasome responses 6 (Fig. 1m–p). Remarkably, rather than being augmented by caspase-12 deletion, E. coli-and C. rodentium-induced IL-1β and IL-18 secretion, caspase-1 maturation and pyroptosis induction by the non-canonical Nlrp3 inflammasome were blunted in caspase-12−/− s macrophages (Fig. 1m–p). Saleh et al. 3 reported that caspase-12−/− s mice express normal levels of caspase-11 mRNA and protein, but unlike wild-type BMDMs, caspase-12−/− s BMDMs failed to express caspase-11 when stimulated with LPS or interferon-γ (IFN-γ) in our studies (Fig. 1q …