IFN regulatory factor (IRF)-2(-/-) mice are significantly more resistant to LPS challenge than wild-type littermates, and this was correlated with increased numbers of apoptotic Kupffer cells. To assess the generality of this observation, and to understand the role of IRF-2 in apoptosis, responses of peritoneal macrophages from IRF-2(+/+) and IRF-2(-/-) mice to apoptotic stimuli, including the fungal metabolite, gliotoxin, were compared. IRF-2(-/-) macrophages exhibited a consistently higher incidence of apoptosis that failed to correlate with caspase-3/7 activity. Using microarray gene expression profiling of liver RNA samples derived from IRF-2(+/+) and IRF-2(-/-) mice treated with saline or LPS, we identified >40 genes that were significantly down-regulated in IRF-2(-/-) mice, including Stat3, which has been reported to regulate apoptosis. Compared with IRF-2(+/+) macrophages, STAT3alpha mRNA was up-regulated constitutively or after gliotoxin treatment of IRF-2(-/-) macrophages, whereas STAT3beta mRNA was down-regulated. Phospho-Y705-STAT3, phospho-S727-STAT1, and phospho-p38 protein levels were also significantly higher in IRF-2(-/-) than control macrophages. Activation of the STAT signaling pathway has been shown to elicit expression of CASP1 and apoptosis. IRF-2(-/-) macrophages exhibited increased basal and gliotoxin-induced caspase-1 mRNA expression and enhanced caspase-1 activity. Pharmacologic inhibition of STAT3 and caspase-1 abolished gliotoxin-induced apoptosis in IRF-2(-/-) macrophages. A novel IFN-stimulated response element, identified within the murine promoter of Casp1, was determined to be functional by EMSA and supershift analysis. Collectively, these data support the hypothesis that IRF-2 acts as a transcriptional repressor of Casp1, and that the absence of IRF-2 renders macrophages more sensitive to apoptotic stimuli in a caspase-1-dependent process.