Virulence adaptation of Pseudomonas aeruginosa phospholipase mutant with altered membrane phospholipid composition

Abstract

Membrane protein and phospholipid (PL) composition changes in response to environmental cues and during infections. Covalent modification and remodelling of the acyl chain length of PLs is an important bacterial adaptation mechanism. However, little is known about which bacterial pathways are regulated in response to altered PL composition. Here, we showed that P. aeruginosa phospholipase A, PlaF, which modulates membrane PL composition, is important for biofilm biogenesis, and we performed whole-cell quantitative proteomics of P. aeruginosa wild-type and {Delta}plaF biofilms to identify pathways regulated by PlaF. The results revealed profound alterations in the abundance of several two-component systems (TCSs), including accumulation of PprAB, which controls the transition to biofilm. Furthermore, a unique phosphorylation pattern of transcriptional regulators, transporters and metabolic enzymes, as well as differential production of seven proteases, in {Delta}plaF, indicate that PlaF-mediated virulence adaptation involves complex transcriptional and posttranscriptional regulation. Moreover, proteomics revealed the depletion of pyoverdine-mediated iron uptake pathway proteins in {Delta}plaF, which agrees with the decreased concentrations of extracellular pyoverdine and intracellular iron and is likely responsible for its prolonged lag growth phase, presumably due to reduced iron uptake. Conversely, the accumulation of proteins from alternative iron-uptake systems in {Delta}plaF suggests that PlaF may function as a switch between different iron-acquisition pathways. The observation that {Delta}plaF accumulates PL-acyl chain modifying and PL synthesis enzymes reveals novel insights into the role of PlaF for membrane PL homeostasis. Although the precise mechanism by which PlaF simultaneously affects multiple pathways remains to be elucidated, we suggest that PlaF-catalyses the degradation of PLs which then serve as a signal that is amplified by proteins of two-component, phosphorylation and proteolytic degradation systems to elicit the global adaptive response in P. aeruginosa.