Dysregulated hemolysin liberates bacterial outer membrane vesicles for cytosolic lipopolysaccharide sensing

PLoS Pathog. 2018 Aug 23;14(8):e1007240. doi: 10.1371/journal.ppat.1007240. eCollection 2018 Aug.

Abstract

Inflammatory caspase-11/4/5 recognize cytosolic LPS from invading Gram-negative bacteria and induce pyroptosis and cytokine release, forming rapid innate antibacterial defenses. Since extracellular or vacuole-constrained bacteria are thought to rarely access the cytoplasm, how their LPS are exposed to the cytosolic sensors is a critical event for pathogen recognition. Hemolysin is a pore-forming bacterial toxin, which was generally accepted to rupture cell membrane, leading to cell lysis. Whether and how hemolysin participates in non-canonical inflammasome signaling remains undiscovered. Here, we show that hemolysin-overexpressed enterobacteria triggered significantly increased caspase-4 activation in human intestinal epithelial cell lines. Hemolysin promoted LPS cytosolic delivery from extracellular bacteria through dynamin-dependent endocytosis. Further, we revealed that hemolysin was largely associated with bacterial outer membrane vesicles (OMVs) and induced rupture of OMV-containing vacuoles, subsequently increasing LPS exposure to the cytosolic sensor. Accordingly, overexpression of hemolysin promoted caspase-11 dependent IL-18 secretion and gut inflammation in mice, which was associated with restricting bacterial colonization in vivo. Together, our work reveals a concept that hemolysin promotes noncanonical inflammasome activation via liberating OMVs for cytosolic LPS sensing, which offers insights into innate immune surveillance of dysregulated hemolysin via caspase-11/4 in intestinal antibacterial defenses.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bacterial Outer Membrane Proteins / metabolism*
  • Caco-2 Cells
  • Caspases / genetics
  • Caspases / metabolism
  • Caspases, Initiator / genetics
  • Caspases, Initiator / metabolism
  • Cytosol / metabolism
  • Extracellular Vesicles / metabolism*
  • Gram-Negative Bacteria / immunology
  • Gram-Negative Bacteria / metabolism
  • Gram-Negative Bacteria / ultrastructure
  • HEK293 Cells
  • HT29 Cells
  • HeLa Cells
  • Hemolysin Proteins / genetics*
  • Hemolysin Proteins / metabolism
  • Humans
  • Immunity, Innate / genetics*
  • Inflammasomes / metabolism
  • Inflammation / immunology
  • Inflammation / metabolism
  • Lipopolysaccharides / metabolism*
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Transfection
  • Up-Regulation / genetics

Substances

  • Bacterial Outer Membrane Proteins
  • Hemolysin Proteins
  • Inflammasomes
  • Lipopolysaccharides
  • CASP4 protein, human
  • Caspases
  • Caspases, Initiator