Cytosolic entry of LPS and activation of noncanonical inflammasome via bacterial outer membrane vesicles

Abstract

Abstract Sensing of lipopolysaccharide (LPS) in the cytosol triggers caspase-11 activation and is central to host defense against Gram-negative bacterial infections and to the pathogenesis of sepsis. Most Gram-negative bacteria that activate caspase-11 however are not cytosolic and the underlying mechanism by which LPS from these bacteria gains access to caspase-11 in the cytosol remains elusive. Here we identify outer membrane vesicles (OMV) produced by Gram-negative bacteria as a vehicle that delivers LPS into the cytosol triggering caspase-11-dependent effector responses in vitro and in vivo. OMV are internalized by macrophages and LPS is released into the cytosol from early and late endosomes. The use of hypovesiculating bacterial mutants, compromised in their ability to generate OMV, reveal the importance of OMV in mediating the cytosolic localization of LPS. Collectively, these findings demonstrate a critical role for OMV in enabling the cytosolic entry of LPS and consequently caspase-11 activation during Gram-negative bacterial infections. In broader terms, our data reveal a fundamental mechanism that serves as a conduit to deliver pathogen-associated molecular patterns (PAMP) to the cytosol to alert the immune system. Our data suggest that OMV with the cargo of immunostimulatory PAMP represent a cardinal sign of infection with not only viable but also multiplying bacteria consistent with the “patterns of pathogenesis” hypothesis.