Gram-negative bacteria that replicate in the cytosol of mammalian macrophages can activate a signaling pathway leading to caspase-1 cleavage and secretion of interleukin 1β, a powerful host response factor. Ipaf, a cytosolic pattern-recognition receptor in the family of nucleotide-binding oligomerization domain–leucine-rich repeat proteins, is critical in such a response to salmonella infection, but the mechanism of how Ipaf is activated by the bacterium remains poorly understood. Here we demonstrate that salmonella strains either lacking flagellin or expressing mutant flagellin were deficient in activation of caspase-1 and in interleukin 1β secretion, although transcription factor NF-κB–dependent production of interleukin 6 or the chemokine MCP-1 was unimpaired. Delivery of flagellin to the macrophage cytosol induced Ipaf-dependent activation of caspase-1 that was independent of Toll-like receptor 5, required for recognition of extracellular flagellin. In macrophages made tolerant by previous exposure to lipopolysaccharide, which abrogates activation of NF-κB and mitogen-activated protein kinases, salmonella infection still activated caspase-1. Thus, detection of flagellin through Ipaf induces caspase-1 activation independently of Toll-like receptor 5 in salmonella-infected and lipopolysaccharide-tolerized macrophages.
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We thank C. McDonald for critical review of the manuscript. Supported by the National Institutes of Health (R01 AI064748 and R01 AI063331; and 5/T32/HL007517 to T.-D.K.), Fondazione Italiana Ricerca sul Cancro (L.F.) and Fondation pour la Recherche Medicale (M.B.-M.).
The authors declare no competing financial interests.
Infection of macrophages with wild-type and mutant salmonella. (PDF 732 kb)
Generation of Ipaf KO mice. (PDF 1125 kb)
Detection of IL-1β (p17 subunit) in culture supernatants after infection with salmonella. (PDF 749 kb)
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