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Cytoplasmic flagellin activates caspase-1 and secretion of interleukin 1β via Ipaf

Abstract

Macrophages respond to Salmonella typhimurium infection via Ipaf, a NACHT–leucine-rich repeat family member that activates caspase-1 and secretion of interleukin 1β. However, the specific microbial salmonella-derived agonist responsible for activating Ipaf is unknown. We show here that cytosolic bacterial flagellin activated caspase-1 through Ipaf but was independent of Toll-like receptor 5, a known flagellin sensor. Stimulation of the Ipaf pathway in macrophages after infection required a functional salmonella pathogenicity island 1 type III secretion system but not the flagellar type III secretion system; furthermore, Ipaf activation could be recapitulated by the introduction of purified flagellin directly into the cytoplasm. These observations raise the possibility that the salmonella pathogenicity island 1 type III secretion system cannot completely exclude 'promiscuous' secretion of flagellin and that the host capitalizes on this 'error' by activating a potent host-defense pathway.

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Figure 1: Salmonella expressing flagellin activate caspase-1 in BMMs independently of bacterial motility or intact hook–basal body structure.
Figure 2: SPI1 TTSS–dependent, flagellin-independent IL-1β secretion occurs at a high MOI.
Figure 3: Flagellar mutants interact normally with cultured cells.
Figure 4: Cytoplasmic flagellin stimulates IL-1β secretion.
Figure 5: BMMs respond to flagellin by secreting IL-1β independently of TLR5.
Figure 6: Ipaf sensitizes J774A.1 cells to S. typhimurium.
Figure 7: Ipaf is required for the response to cytoplasmic flagellin.

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Acknowledgements

We thank V. Dixit and Genentech for Ipaf- and ASC-deficient mice; S. Akira for TLR5- and MyD88-deficient mice; G. Chilcott, H. Bonnifield, O. Nanassy, J. Karlinsey and K. Hughes for flagellar mutant strains and advice regarding flagellar biology; C. Carlson, M. Ohl and K. Smith for contributions and discussions; H. Bonnifield, C. Carlson and members of the Aderem lab for critical review of this manuscript; and E. Andersen-Nissen for providing flagellin and ovalbumin protein.

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Correspondence to Alan Aderem.

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Supplementary information

Supplementary Fig. 1

Schematic of the structures synthesized by flagellar mutants. (PDF 41 kb)

Supplementary Fig. 2

Mutant strains over expressing flagellin display increased cytotoxicity. (PDF 28 kb)

Supplementary Fig. 3

S. typhimurium repress flagellin transcription through PhoP/PhoQ. (PDF 23 kb)

Supplementary Fig. 4

Ipaf sensitizes macrophages to salmonella infection. (PDF 40 kb)

Supplementary Table 1

Strain list: Chromosomal insertion elements and plasmids. (PDF 27 kb)

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Miao, E., Alpuche-Aranda, C., Dors, M. et al. Cytoplasmic flagellin activates caspase-1 and secretion of interleukin 1β via Ipaf. Nat Immunol 7, 569–575 (2006). https://doi.org/10.1038/ni1344

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