Kinetic Control of Quorum Sensing in Pseudomonas aeruginosa by Multidrug Efflux Pumps

Abstract

Pseudomonas aeruginosa is an important human pathogen, the physiology and virulence of which are under the control of quorum sensing signals. These signals often have dual roles, functioning as toxins to some cells and as oxidative-stress protectors for their producer cells. Hence, their internal and external concentrations should be tightly controlled. In this study, we analyzed the interplay between the multidrug efflux transporters MexEF-OprN and MexG/HI-OpmD in quorum sensing of P. aeruginosa. We found that the two transporters have overlapping substrate specificities but different efficiencies. When overproduced, both MexEF-OprN and MexG/HI-OpmD provide clinical levels of resistance to diverse fluoroquinolones and protect P. aeruginosa against toxic phenazines. However, this similarity is enabled by synergistic interactions with the outer membrane. In hyperporinated cells, MexG/HI-OpmD is saturated by much lower concentrations of fluoroquinolones but is more efficient than MexEF-OprN in efflux of phenazines. Unlike MexEF-OprN, mutational inactivation of MexG/HI-OpmD reduces the levels of pyocyanin and makes P. aeruginosa cells hypersusceptible to phenazines. Our results further show that MexG binds pyocyanin, physically associates with MexHI and represses the activity of the transporter, revealing a negative regulatory role of this protein. We conclude that differences in kinetic properties of transporters are critical to maintain proper intra- and extra- cellular concentrations of phenazines and other signaling molecules, and that MexG/HI-OpmD controls the steady-state in the synthesis and secretion of phenazines.

Lactonase Specificity Is Key to Quorum Quenching in Pseudomonas aeruginosa

“…Several proteins involved in oxidative stress response and antibiotic sensitivity exhibited a modified expression level. The abundance of three proteins (PA14_18690, OpdB, and OprG) were increased and seven others (PA14_47550, PA14_18330/50, ObgE, MexG, MexH, and OmpD) were reduced with both treatments as compared to the control (Aendekerk, 2005;Breidenstein et al, 2008;Dotsch et al, 2009;Balasubramanian et al, 2013;Verstraeten et al, 2015;Sakhtah et al, 2016;Chevalier et al, 2017;Gaviard et al, 2018;Wolloscheck et al, 2018). Several other protein levels showed differences between the two lactonase treatments.…”

Der Naturstoff Elegaphenon verstärkt antibiotische Effekte gegen Pseudomonas aeruginosa

“…[19] Im Gegensatz dazu wurden keine ausgeprägten Unterschiede in Pyocyanin-Enzymen beobachtet, was darauf hindeutet, dass die geringeren extrazellulären Mengen dieses Toxins von vermindertem MexGHIvermitteltem Tr ansport herrühren. Die Proteome von mit ELP5 behandelten P. aeruginosa PAO1 und der PW9844 AlgP-Transposonmutanten wurden über LC-MS/MS analysiert und zeigten eine verblüffende allgemeine ¾hnlichkeit der regulierten Proteine (Abbildungen S9, S10).…”

“…Interessanterweise waren in die Pyoverdinbiosynthese involvierte Proteine sowie MexGHI-OpmD,ein bekannter Phenazintransporter, in beiden Proben stark herunterreguliert. [19]…”

The Natural Product Elegaphenone Potentiates Antibiotic Effects against Pseudomonas aeruginosa

“…Unlike the study in E. faecalis, EF-Tucould not be detected as significantly enriched. [19] By contrast, no pronounced changes for pyocyanin biosynthesis enzymes were observed, suggesting that the lower extracellular levels of this toxin may stem from reduced MexGHI-mediated transport. [17] Recombinant AlgP was also labeled by the probe,suggesting direct binding ( Figure S7).…”

“…Interestingly,p yoverdine biosynthesis proteins as well as MexGHI-OpmD,ak nown transporter of phenazines,w ere strongly downregulated in both samples. [19] By contrast, no pronounced changes for pyocyanin biosynthesis enzymes were observed, suggesting that the lower extracellular levels of this toxin may stem from reduced MexGHI-mediated transport. Taken together, our proteomic profiles confirm as trong link between compound treatment and impairment of AlgP function.…”