Abstract
Delivery of cargo to target cells is fundamental to bacterial competitiveness. One important but poorly understood system, ubiquitous among Gram-negative organisms, involves packaging cargo into outer membrane vesicles (OMVs). These biological nanoparticles are involved in processes ranging from toxin delivery to cell-cell communication. Despite this, we know comparatively little about how OMVs are formed. Building upon the discovery that the Pseudomonas Quinolone Signal (PQS) stimulates OMV biogenesis in Pseudomonas aeruginosa, we proposed a model where PQS interacts with the outer membrane to induce curvature and ultimately OMV formation. Though this model is well supported in P. aeruginosa, it remained unclear whether other organisms produce similar compounds. Here we describe the development of a tightly controlled experimental system to test the interaction of bacterially-produced factors with target cells. Using this system, we show that multiple species respond to PQS by increasing OMV formation, that PQS accumulates in the induced vesicles, and that other bacteria secrete OMV-promoting factors. Analysis of induced vesicles indicates that recipient-mediated mechanisms exist to control vesicle size and that relatedness to the producer organism can dictate susceptibility to OMV-inducing compounds. This work provides evidence that small molecule induced OMV biogenesis is a widely conserved process and that cross-talk between systems may influence OMV production in neighboring bacteria.
Highlights
Outer Membrane Vesicles are small unilamellar structures released from Gram-negative bacteria that contain cargo such as virulence factors, enzymes, DNA, and communication signals[1,2,3,4]
We proposed that intercalation of Pseudomonas Quinolone Signal (PQS) into the outer leaflet of the P. aeruginosa outer membrane causes asymmetric leaflet expansion leading to membrane curvature and outer membrane vesicles (OMVs) biogenesis[23]
Since we showed that PQS induces OMV production in several strains of gammaproteobacteria and that PQS can be recovered in induced P. aeruginosa ΔpqsA OMVs, we investigated whether PQS induced vesicles from other recipient gammaproteobacteria contained PQS
Summary
Outer Membrane Vesicles are small unilamellar structures released from Gram-negative bacteria that contain cargo such as virulence factors, enzymes, DNA, and communication signals[1,2,3,4]. A different class of self-produced hydrophobic signaling molecule (related to the diffusible signal factor of Xanthomonas campestris) has been associated with both increased (in Stenotrophomonas maltophilia27) and decreased (in Xylella fastidiosa28) production of OMVs, though in both of these cases the authors described their findings in the context of quorum signaling rather than physical interaction with the membrane Observations such as these highlight the potential for SMI OMV biogenesis to be active across species and even to be involved in cross-species interactions. With this work we set out to develop a rigorous and controlled system to test the cross-species effects of a known OMV-inducing compound and to lay the groundwork for identification of similar factors produced by other species Using this system, we show that PQS is capable of OMV induction in other gammaproteobacteria under strict conditions, but that its effect is limited in more distantly related species. We identify cross-species (targeting P. aeruginosa) OMV-inducing activities in the supernatants of Escherichia coli and Klebsiella pneumoniae, supporting the hypothesis that SMI OMV biogenesis is active in multiple species
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