Chemical Inhibition of Kynureninase Reduces Pseudomonas aeruginosa Quorum Sensing and Virulence Factor Expression

Abstract

The opportunistic pathogen Pseudomonas aeruginosa utilizes multiple quorum sensing (QS) pathways to coordinate an arsenal of virulence factors. We previously identified several cysteine-based compounds inspired by natural products from the plant Petiveria alliacea which are capable of antagonizing multiple QS circuits as well as reducing P. aeruginosa biofilm formation. To understand the global effects of such compounds on virulence factor production and elucidate their mechanism of action, RNA-seq transcriptomic analysis was performed on P. aeruginosa PAO1 exposed to S-phenyl-l-cysteine sulfoxide, the most potent inhibitor from the prior study. Exposure to this inhibitor down-regulated expression of several QS-regulated virulence operons (e.g., phenazine biosynthesis, type VI secretion systems). Interestingly, many genes that were differentially regulated pertain to the related metabolic pathways that yield precursors of pyochelin, tricarboxylic acid cycle intermediates, phenazines, and Pseudomonas quinolone signal (PQS). Activation of the MexT-regulon was also indicated, including the multidrug efflux pump encoded by mexEF-oprN, which has previously been shown to inhibit QS and pathogenicity. Deeper investigation of the metabolites involved in these systems revealed that S-phenyl-l-cysteine sulfoxide has structural similarity to kynurenine, a precursor of anthranilate, which is critical for P. aeruginosa virulence. By supplementing exogenous anthranilate, the QS-inhibitory effect was reversed. Finally, it was shown that S-phenyl-l-cysteine sulfoxide competitively inhibits P. aeruginosa kynureninase (KynU) activity in vitro and reduces PQS production in vivo. The kynurenine pathway has been implicated in P. aeruginosa QS and virulence factor expression; however, this is the first study to show that targeted inhibition of KynU affects P. aeruginosa gene expression and QS, suggesting a potential antivirulence strategy.

The aliphatic amidase AmiE is involved in regulation of Pseudomonas aeruginosa virulence

“…In parallel it is interesting to note that retrieving AmiE’s in silico functional interactions from STRING database42, led to predict interactions of AmiE with PA14_63110, as well as with the guanidinobutyrase (GbuA), a mutant of which was shown to be strongly affected in PQS production43. Remarkably, AmiE over-production induced also a decrease in PQS synthesis at least through a lower accumulation of i) its biosynthetic enzymes PqsB, PqsC and PqsD, and ii) the kynunerinase KinU (Supplementary Table S1) that is involved in the degradation of kynurenine leading to the production of the PQS precursor anthranilate44. Taken together, these data suggest strong relationships between AmiE and QS signals production, thus linking metabolism and QS in P. aeruginosa , as previously suggested43.…”

Abnormal kynurenine pathway of tryptophan catabolism in cardiovascular diseases

“…have increased Kyn and KA levels in the brain, blood, liver, and kidney, associating with a decreased locomotor activity [157]. Recently, another structural analogue of Kyn, S-phenyl-L-cysteine sulfoxide, was shown to competitively inhibit P. aeruginosa KYNU activity in vitro, suggesting a potential antivirulence strategy [159]. Since KYNU directly controls 3-HAA formation, which regulates cardiovascular cells inflammation and apoptosis, it would be interesting to exam the effect of KYNU inhibition on CVD initiation and development.…”

Cinnamaldehyde Induces Expression of Efflux Pumps and Multidrug Resistance in Pseudomonas aeruginosa

“…oils activate AcrAB-TolC in Escherichia coli (14,15). The S-phenyl-L-cysteine sulfoxide from the tropical plant Petiveria alliacea reduces the virulence of P. aeruginosa by inhibiting kynureninase KynU, but it stimulates MexEF-OprN expression (16).…”