The release of nano-sized membrane vesicles is a conserved phenomenon amongst Gram-negative bacteria. These bi-layered structures, known as outer membrane vesicles (OMVs), are adept at entering eukaryotic cells, facilitating delivery of bacterial factors into host cells. Amongst a range of bacterial proteins and glycolipids, OMVs have been shown to contain DNA. Despite its putative importance in both bacterial and immune functions, very little is known about the nature of OMV-associated DNA, its interactions with host cells and the immune system.
This study investigates the mechanisms by which OMV-associated DNA may play a role in inflammation. To this end, we characterised the form and sequence of OMV associated DNA, demonstrated its uptake into eukaryotic cells and investigated the innate immune molecules involved in sensing OMV-associated DNA. To characterise the form and sequence of OMV-associated DNA, we used Ion Torrent semiconductor sequencing and electron microscopy to show that OMVs are associated with bacterial genomic DNA that is predominantly bound to the OMV surface. Using immunofluorescence, we have shown for the first time the uptake of OMV-associated DNA into eukaryotic cells. Finally, we used in vitro systems to determine the response of the innate DNA sensors, endosomal Toll-like receptor 9 and the cytosolic DNA-sensing inflammasome absent in melanoma 2 (AIM2), to OMV-associated DNA. While TLR9 does not appear to play a significant role, our preliminary results indicate that OMVs induce an inflammasome response. Our future experiments aim to elucidate whether this inflammasome response is due to the DNA-sensing inflammasome, AIM2.
Collectively, our findings suggest that OMVs are associated with bacterial DNA which is carried as cargo into eukaryotic cells, whereby it may activate the cytosolic DNA-sensing inflammasome, AIM2. This study may shed light on the mechanism of action of OMV-based vaccines. Furthermore, it has the potential to lead to novel treatment targets against Gram negative bacterial disease progression.