Deletion of ripA Alleviates Suppression of the Inflammasome and MAPK by Francisella tularensis

Abstract

Francisella tularensis is a facultative intracellular pathogen and potential biothreat agent. Evasion of the immune response contributes to the extraordinary virulence of this organism although the mechanism is unclear. Whereas wild-type strains induced low levels of cytokines, an F. tularensis ripA deletion mutant (LVSΔripA) provoked significant release of IL-1β, IL-18, and TNF-α by resting macrophages. IL-1β and IL-18 secretion was dependent on inflammasome components pyrin-caspase recruitment domain/apoptotic speck-containing protein with a caspase recruitment domain and caspase-1, and the TLR/IL-1R signaling molecule MyD88 was required for inflammatory cytokine synthesis. Complementation of LVSΔripA with a plasmid encoding ripA restored immune evasion. Similar findings were observed in a human monocytic line. The presence of ripA nearly eliminated activation of MAPKs including ERK1/2, JNK, and p38, and pharmacologic inhibitors of these three MAPKs reduced cytokine induction by LVSΔripA. Animals infected with LVSΔripA mounted a stronger IL-1β and TNF-α response than that of mice infected with wild-type live vaccine strain. This analysis revealed novel immune evasive mechanisms of F. tularensis.

Repression of Inflammasome by Francisella tularensis during Early Stages of Infection

“…However, as reported in previous studies, we did not observe this NLRP3 dependence at 24 h post-infection either in F. tularensis LVS or the FTL_0325 mutant-infected macrophages. Similar to several previous reports, our results also demonstrate an AIM2-dependent but NLRP3-independent inflammasome activation at 16 -24 h post-infection in F. tularensis LVS-infected macrophages (34,35,48).…”

“…A phenotype similar to the FTL_0325 has also been reported for the ⌬ripA and mviN mutant of F. tularensis LVS. RipA suppresses the MAPK signaling pathway to down-regulate proIL-1␤ synthesis and IL-1␤ production and that activation of IL-1␤ in ⌬ripA mutant is mediated by an AIM2-dependent inflammasome (34). However, Peng et al (36) reported that the hyperinflammatory and hypercytotoxic nature of the ⌬ripA mutant was due to altered membrane integrity and intramacrophage lysis of the mutant, resulting in the release of excessive bacterial DNA and activation of AIM2 inflammasome.…”

“…Similarly, Mycobacterium tuberculosis prevents inflammasome activation and, consequently, IL-1␤ secretion by expressing a zinc protease Zmp1 (32), whereas expression of an exoenzyme, ExoU, by Pseudomonas aeruginosa results in the evasion of inflammasome activation in the infected host cells (33). The Francisella-encoded factors MviN and RipA were initially described as required for suppression of AIM2-mediated inflammasome responses (34,35). However, a later study identified that the activation of AIM2 inflammasome in ripA or mviN mutants was due to enhanced intramacrophage lysis of the mutant bacteria leading to the release of DNA into the host cell cytosol and subsequent activation of AIM2 inflammasome (36).…”

Identification of a Novel Francisella tularensis Factor Required for Intramacrophage Survival and Subversion of Innate Immune Response

“…Yersinia, Shigella, and Bacillus anthracis inhibit MAPK signaling to suppress inflammatory response that permits bacterial growth and dissemination (55)(56)(57)(58). It has been reported that F. tularensis LVS suppresses MAPK signaling via intracellular growth locus C and RipA (16,19). This study reports that the FTL_0325/FTT0831c proteins of both the F. tularensis LVS and SchuS4 suppress proinflammatory cytokine response by inhibiting NF-B signaling.…”

“…Moreover, it has been demonstrated that FopC suppresses IFN-␥ signaling and thus interferes with generation of an effective innate immune response (59). Several other structural components of both the LVS and F. novicida such as MviN, a lipid II filppase (61), RipA, a cytoplasmic membrane protein (19), or the LPS O-antigen (62) and capsular components have been shown to be required for virulence, suppression of proinflammatory cytokines, and cell death in infected macrophages. Results from the present study indicate that FTL_0325 serves to play a role different than these structural proteins.…”