Abstract
Bacteria-released components can modulate host innate immune response in the absence of direct host cell–bacteria interaction. In particular, bacteria-derived outer membrane vesicles (OMVs) were recently shown to activate host caspase-11-mediated non-canonical inflammasome pathway via deliverance of OMV-bound lipopolysaccharide. However, further precise understanding of innate immune-modulation by bacterial OMVs remains elusive. Here, we present evidence that flagellated bacteria-released OMVs can trigger NLRC4 canonical inflammasome activation via flagellin delivery to the cytoplasm of host cells. Salmonella typhimurium-derived OMVs caused a robust NLRC4-mediated caspase-1 activation and interleukin-1β secretion in macrophages in an endocytosis-dependent, but guanylate-binding protein-independent manner. Notably, OMV-associated flagellin is crucial for Salmonella OMV-induced inflammasome response. Flagellated Pseudomonas aeruginosa-released OMVs consistently promoted robust NLRC4 inflammasome activation, while non-flagellated Escherichia coli-released OMVs induced NLRC4-independent non-canonical inflammasome activation leading to NLRP3-mediated interleukin-1β secretion. Flagellin-deficient Salmonella OMVs caused a weak interleukin-1β production in a NLRP3-dependent manner. These findings indicate that Salmonella OMV triggers NLRC4 inflammasome activation via OMV-associated flagellin in addition to a mild induction of non-canonical inflammasome signaling via OMV-bound lipopolysaccharide. Intriguingly, flagellated Salmonella-derived OMVs induced more rapid inflammasome response than flagellin-deficient Salmonella OMV and non-flagellated Escherichia coli-derived OMVs. Supporting these in vitro results, Nlrc4-deficient mice showed significantly reduced interleukin-1β production after intraperitoneal challenge with Salmonella-released OMVs. Taken together, our results here propose that NLRC4 inflammasome machinery is a rapid sensor of bacterial OMV-bound flagellin as a host defense mechanism against bacterial pathogen infection.
Highlights
Host immune system attempts to detect invasion of bacterial pathogens via pattern-recognition receptors on tissue-resident sentinel cells, such as macrophages [1]
Given that flagellin may be the main stimulus for Salmonellareleased outer membrane vesicles (OMVs)-induced NLRC4 inflammasome activation, we examined the roles of OMVs from non-flagellated bacteria, such as E. coli strains BL21 and DH5a
E. coli OMVs promoted stronger gasdermin D cleavage (Figure 6D), as determined by the presence of cleaved GSDMD (NT) in the cytosol, and pyroptotic cell death (Figure 6E) than S. typhimurium OMVs at 8 htreatment to macrophages. These results propose that NLRC4 inflammasome machinery is a critical sensor for rapid detection of flagellated bacteria-released OMVs as a host defense mechanism, whereas non-flagellated bacteria-induced caspase-11 noncanonical inflammasome signaling rather contributes to pyroptosis in response to bacterial OMV-delivered LPS
Summary
Host immune system attempts to detect invasion of bacterial pathogens via pattern-recognition receptors on tissue-resident sentinel cells, such as macrophages [1]. Recognition of bacterial ligand by these receptors promptly results in the production of proinflammatory cytokines, which trigger host inflammatory responses by recruiting more leukocytes from the bloodstream to clear the invading pathogens [2]. Among the proinflammatory cytokines produced during the initial stage of bacterial infection, interleukin (IL)-1b has a crucial role in initiation of the inflammatory process [3]. This complex consists mostly of sensor protein, adaptor protein, and procaspase-1, leading to the generation of active caspase-1 [5]. Caspase-1 processes inactive pro-IL-1b into an active form and facilitates leaderless IL-1b secretion into the extracellular space
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