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SLO and Steady: Role of Streptolysin O during Inflammasome Activation in Group A Streptococcus‐infected THP‐1 Macrophages
Author(s) -
Alcantara Cheldon Ann T.,
Okumura Cheryl Y.M.
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.04716
Subject(s) - inflammasome , streptolysin , secretion , streptococcus pyogenes , microbiology and biotechnology , aim2 , phagocytosis , inflammation , cytosol , proinflammatory cytokine , intracellular , thp1 cell line , bacteria , chemistry , biology , immunology , biochemistry , enzyme , staphylococcus aureus , bacterial protein , cell culture , genetics
Group A Streptococcus (GAS; Streptococcus pyogenes ) is a Gram‐positive bacterium that infects millions of people globally each year, with a number of cases developing into serious infections such as septicemia, necrotizing fasciitis, and streptococcal toxic shock syndrome. Upon phagocytosis of GAS, intracellular detection of the bacteria triggers a localized inflammatory response by activation of the NLRP3 inflammasome, a cytosolic multimeric protein complex that produces pro‐inflammatory cytokines. A two‐signal system is required for NLRP3 activation which 1) induces transcription of the oligomer components and pro‐IL‐1β, and 2) detects cellular damage to elicit cleavage of pro‐IL‐1β to mature IL‐1β and subsequent extracellular release. Although several GAS proteins have been reported to trigger NLRP3 inflammasome activation, the mechanisms by which bacteria cause inflammation are generally unknown. We have shown that proteins up to 40kDa can escape GAS‐infected phagosomes via Streptolysin O (SLO)‐induced pores, which suggests that either a host phagolysosomal protein or a secreted bacterial protein is detected by cytosolic pathogen recognition receptors (PRRs) to trigger NLRP3‐induced inflammation. To test this, we first infected THP‐1 macrophages with WT or heat‐killed (HK) GAS to establish if a secreted protein activates the inflammasome by monitoring IL‐1β secretion using enzyme‐linked immunosorbent assay (ELISA). We found that HK GAS did not stimulate IL‐ Iβ release, indicating that secretion of bacterial components are necessary for inflammasome activation. To determine essential proteins for GAS‐induced inflammation, we infected THP‐1 macrophages with various secretory protein knockout strains and found that only strains lacking SLO displayed significantly decreased IL‐1β release, confirming that pore formation is necessary for inflammasome activation. Using recombinant SLO (rSLO), we found that the protein itself does not stimulate IL‐1β release. However, when rSLO is coupled with HK GAS, IL‐1β release occurs, indicating that host phagolysosomal proteins may be the trigger for inflammasome activation. The information gained through our investigation provides insight into how GAS and possibly other pathogens or agents that damage the phagosome activate the inflammasome, which may contribute to the development of more effective therapies for infections that cause life‐threatening inflammation. Support or Funding Information American Heart Association (17GRNT3410851), Occidental College Undergraduate Research Center