Dataset Information

Salmonella Flagellin Activates NAIP/NLRC4 and Canonical NLRP3 Inflammasomes in Human Macrophages.

ABSTRACT: Infection of human macrophages with Salmonella enterica serovar Typhimurium (S. Typhimurium) leads to inflammasome activation. Inflammasomes are multiprotein complexes facilitating caspase-1 activation and subsequent gasdermin D-mediated cell death and IL-1? and IL-18 cytokine release. The NAIP/NLRC4 inflammasome is activated by multiple bacterial protein ligands, including flagellin from the flagellum and the needle protein PrgI from the S. Typhimurium type III secretion system. In this study, we show that transfected ultrapure flagellin from S Typhimurium induced cell death and cytokine secretion in THP-1 cells and primary human monocyte-derived macrophages. In THP-1 cells, NAIP/NLRC4 and NLRP3 played redundant roles in inflammasome activation during infection with S. Typhimurium. Knockout of NAIP or NLRC4 in THP-1 cells revealed that flagellin, but not PrgI, now activated the NLRP3 inflammasome through a reactive oxygen species- and/or cathepsin-dependent mechanism that was independent of caspase-4/5 activity. In conclusion, our data suggest that NLRP3 can be activated by flagellin to act as a "safety net" to maintain inflammasome activation under conditions of suboptimal NAIP/NLRC4 activation, as observed in THP-1 cells, possibly explaining the redundant role of NLRP3 and NAIP/NLRC4 during S. Typhimurium infection.

SUBMITTER: Gram AM

PROVIDER: S-EPMC7812056 | biostudies-literature | 2021 Feb

REPOSITORIES: biostudies-literature

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Salmonella Flagellin Activates NAIP/NLRC4 and Canonical NLRP3 Inflammasomes in Human Macrophages.

Gram Anna M AM Wright John A JA Pickering Robert J RJ Lam Nathaniel L NL Booty Lee M LM Webster Steve J SJ Bryant Clare E CE

Journal of immunology (Baltimore, Md. : 1950) 20201230 3

Infection of human macrophages with Salmonella enterica serovar Typhimurium (S. Typhimurium) leads to inflammasome activation. Inflammasomes are multiprotein complexes facilitating caspase-1 activation and subsequent gasdermin D-mediated cell death and IL-1β and IL-18 cytokine release. The NAIP/NLRC4 inflammasome is activated by multiple bacterial protein ligands, including flagellin from the flagellum and the needle protein PrgI from the S. Typhimurium type III secretion sy ...[more]

PMID: 33380493

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Project description:BackgroundA recombinant fusion protein combining the adjuvant and TLR5-ligand flagellin with the major birch pollen allergen Bet v 1 (rFlaA:Betv1) has been suggested to prevent the manifestation of birch allergy. Noteworthy, rFlaA:Betv1 induced both pro- and anti-inflammatory responses which were differentially regulated. However, the mechanism by which flagellin fusion proteins modulate allergen-specific immune responses, especially the mechanisms underlying IL-1β secretion and their contribution to the overall immune responses remains elusive.ObjectiveTo investigate the mechanisms underlying the production of IL-1β from rFlaA:Betv1 stimulated macrophages.MethodsMacrophages were derived from mouse peritoneal-, human buffy-coat-, and PMA-differentiated THP-1 (wild type or lacking either ASC, NLRP3, or NLRC4) cells. Macrophages were stimulated with non-modified rFlaA:Betv1, mutant variants lacking either the flagellin DC0 domain or a sequence motif formerly described to mediate TLR5-activation, and respective controls in the presence or absence of inhibitors interfering with MAPK- and NFκB-signaling. Cytokine secretion was analyzed by ELISA and intracellular signaling by Western Blot. To study the contribution of IL-1β to the overall immune responses, IL1R-deficient mouse peritoneal macrophages were used.ResultsrFlaA:Betv1 consistently activated all types of investigated macrophages, inducing higher IL-1β secretion compared with the equimolar mixture of both proteins. rFlaA:Betv1-induced activation of THP-1 macrophages was shown to be independent of either the TLR5-activating sequence motif or the flagellin DC0 domain but depended on both NLRP3- and NLRC4-inflammasomes. In addition, NFκB and SAP/JNK MAP kinases regulated rFlaA:Betv1-induced inflammasome activation and cytokine secretion by modulating pro-Caspase-1- and pro-IL-1β-expression in THP-1 macrophages. Finally, lack of IL-1β positive feedback via the IL1R strongly diminished the rFlaA:Betv1-induced secretion of IL-1β, IL-6, and TNF-α from peritoneal macrophages.ConclusionThe mechanisms contributing to rFlaA:Betv1-induced IL-1β secretion from macrophages were shown to be complex, involving both NLRC4- and NLRP3-inflammsomes, as well as NFκB- and SAP/JNK MAP kinase-signaling. Better understanding the mechanisms regulating the activation of immune cells by novel therapeutic candidates like the rFlaA:Betv1 fusion protein will allow us to further improve and develop new treatment strategies when using flagellin as an adjuvant.

Dataset Information

Salmonella Flagellin Activates NAIP/NLRC4 and Canonical NLRP3 Inflammasomes in Human Macrophages.

Publications

<i>Salmonella</i> Flagellin Activates NAIP/NLRC4 and Canonical NLRP3 Inflammasomes in Human Macrophages.

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