Pseudomonas aeruginosa LasI/RhlI quorum sensing system controls protease-mediated autoaggregation behavior, cell envelope characteristics and extracellular proteome responses

Quorum sensing (QS) is an intercellular communication mechanism employed by the opportunistic bacteria <i>Pseudomonas aeruginosa</i> to regulate processes beneficial to the longevity of the community and related to its pathogenicity. The LasI/Rhl circuits of the quorum-sensing (QS) network operate through <i>N</i>-acyl-L-homoserine lactones (AHL) and appear at the top in the QS hierarchy. In natural habitats and host environments, bacteria exist and transit between different modes of lifestyle: planktonic single cells, suspended multicellular aggregates, and surface-attached biofilm communities. Using <i>P. aeruginosa</i> PA14 as a model, we determined the contribution of the master regulator LasI/RhlI QS system to multicellular community autoaggregation in liquid, ultrastructure and fitness characteristics of the cell envelope, and extracellular proteome responses, employing phenotypic assays, light imaging, high-resolution transmission electron microscopy and quantitative mass spectrometry-based proteomics and bioinformatics. Wild-type bacteria with a functional QS system were more effective in the protease-mediated autoaggregation than the <i>lasI-/rhlI-</i> mutant, lacking the production of AHL molecules and associated virulence traits. AHL-dependent communication impacted cell envelope characteristics, including ultrastuctural curvature and tolerance to membrane-damaging and antimicrobial agents. Moreover, the LasI/RhlI QS system perturbed the extracellular abundance of a total of 545 extracellular proteins during late exponential and early stationary growth phases. We allocated most of these differentially expressed proteins to the following large functional groups: metabolism; transcription and translation; transport and secretion systems; cell envelope integrity; redox processes; invasiveness and toxicity; and motility. Remarkably, approximately 95% of the extracellular proteome was upregulated in the <i>lasI-/rhlI-</i> mutant compared with the wild type, and these levels were restored to wild type-status when AHL was added. We observed a crucial contribution of the LasI/RhlI QS system to the protease-mediated community autoaggregation in <i>P. aeruginosa</i> PA14. Mechanistically, this was accompanied-through a sophisticated and multifactorial process-by differential expression of an array of components in the secreted proteome involved in both pathogenicity-specific and global readjustments in the homeostasis within the population. By fine tuning the LasI/RhlI system, <i>Pseudomonas</i> can regulate its pathogenic potential and long-term survival in different hosts and habitats.