Multiple FadD Acyl-CoA Synthetases Contribute to Differential Fatty Acid Degradation and Virulence in Pseudomonas aeruginosa

Abstract

A close interconnection between nutrient metabolism and virulence factor expression contributes to the pathophysiology of Pseudomonas aeruginosa as a successful pathogen. P. aeruginosa fatty acid (FA) degradation is complicated with multiple acyl-CoA synthetase homologs (FadDs) expressed in vivo in lung tissue during cystic fibrosis infections. The promoters of two genetically linked P. aeruginosa fadD genes (fadD1 and fadD2) were mapped and northern blot analysis indicated they could exist on two different transcripts. These FadDs contain ATP/AMP signature and FA-binding motifs highly homologous to those of the Escherichia coli FadD. Upon introduction into an E. coli fadD -/fadR - double mutant, both P. aeruginosa fadDs functionally complemented the E. coli fadD -/fadR - mutant, allowing degradation of different chain-length FAs. Chromosomal mutagenesis, growth analysis, induction studies, and determination of kinetic parameters suggested that FadD1 has a substrate preference for long-chain FAs while FadD2 prefers shorter-chain FAs. When compared to the wild type strain, the fadD2 mutant exhibited decreased production of lipase, protease, rhamnolipid and phospholipase, and retardation of both swimming and swarming motilities. Interestingly, fadD1 mutant showed only increased swarming motility. Growth analysis of the fadD mutants showed noticeable deficiencies in utilizing FAs and phosphatidylcholine (major components of lung surfactant) as the sole carbon source. This defect translated into decreased in vivo fitness of P. aeruginosa in a BALB/c mouse lung infection model, supporting the role of lipids as a significant nutrient source for this bacterium in vivo.

Identification of Genes Involved in Neisseria meningitidis Colonization

“…The V. cholerae fadD mutant was attenuated in an in vivo infant mouse cholera model due to downregulation of the major virulence genes in this mutant (38). Inactivation of fadD affects the virulence of several other bacterial pathogens, including the plant pathogen Sinorhizobium meliloti (39), Salmonella enterica serovar Typhimurium (40), and P. aeruginosa (41). Transcriptome or proteome profiling of a meningococcal fadD mutant could help to explain the colonization defect observed with this mutant.…”

Fatty Acid Biosynthesis in Pseudomonas aeruginosa Is Initiated by the FabY Class of β-Ketoacyl Acyl Carrier Protein Synthases

“…Fatty acid metabolism in especially complex in pseudomonads, with an unusually large genetic allocation to both the catabolism and anabolism of fatty acids (32,50,51). To further confirm the two activities of PA3286 observed in P. aeruginosa PAO1, to probe the substrate specificity and to determine whether PA3286 requires partner proteins for intercepting ␤-oxidation cycle intermediates, we introduced the PA3286 gene into the model E. coli strain BW25113.…”

“…The cis-3-decenoyl-ACP is then specifically extended by the FabB elongase to make unsaturated fatty acids (69). Intermediates from the FAS pathway also provide the ␤-hydroxyl myristoyl-ACP and lauroyl-/myristoyl-ACP needed for lipopolysaccharide biosynthesis (64).The divergence between mammalian and bacterial FAS pathways, coupled with the multifaceted role in both the viability and virulence of many pathogenic bacteria, makes fatty acid inhibition an attractive strategy for the development of new antimicrobial agents (8,32,37,92). Much effort has been devoted toward understanding bacterial FAS enzymes and cognate inhibitors, particularly for the initiating condensing enzyme FabH (9,44, 56,91).…”

“…The divergence between mammalian and bacterial FAS pathways, coupled with the multifaceted role in both the viability and virulence of many pathogenic bacteria, makes fatty acid inhibition an attractive strategy for the development of new antimicrobial agents (8,32,37,92). Much effort has been devoted toward understanding bacterial FAS enzymes and cognate inhibitors, particularly for the initiating condensing enzyme FabH (9,44,56,91).…”

FadD Is Required for Utilization of Endogenous Fatty Acids Released from Membrane Lipids

“…Swarming is a type of bacterial motility characterized by a rapid and coordinated population migration across a surface and involves a complex process of differentiation in which cells usually become hyperflagellated and elongated (19). Interestingly, fadD1 mutants of Pseudomonas aeruginosa also showed an increased swarming motility (25), and fadD mutants of Vibrio cholerae showed a repression of the major virulence genes as well as an increase in motility (38).…”