Phenylacetyl-CoA, not phenylacetic acid, attenuates CepIR-regulated virulence inBurkholderia cenocepacia

Abstract

AbstractDuring phenylalanine catabolism, phenylacetic acid (PAA) is converted to phenylacetyl-CoA (PAA-CoA) by a ligase, PaaK, and then epoxidized by a multicomponent monooxygenase, PaaABCDE, before further degradation to the TCA cycle. In the opportunistic pathogenBurkholderia cenocepacia,loss ofpaaABCDEattenuates virulence factor expression, which is under control of the LuxIR-like quorum sensing system, CepIR. To further investigate the link between CepIR-regulated virulence and PAA catabolism, we created knockout mutants of the first step of the pathway (PAA-CoA synthesis by PaaK) and characterized them in comparison to apaaABCDEmutant using liquid chromatography/mass spectrometry (LC-MS/MS) and virulence assays. We found that while loss of PaaABCDE decreased virulence, deletion of thepaaKgenes resulted in a more virulent phenotype than the wild type strain. Deletion of eitherpaaKorpaaABCDEled to higher levels of released PAA but no differences in internal accumulation, compared to wild type. While we found no evidence of directcepIRdownregulation by PAA-CoA or PAA, a low virulencecepRmutant reverted to a virulent phenotype upon removal of thepaaKgenes. On the other hand, removal ofpaaABCDEin thecepRmutant did not impact its attenuated phenotype. Together, our results suggest an indirect role for PAA-CoA in supressingB. cenocepaciaCepIR-activated virulence.ImportanceThe opportunistic pathogenBurkholderia cenocepaciauses a chemical signal process called quorum sensing (QS) to produce virulence factors. InB. cenocepacia, QS relies on the presence of the transcriptional regulator CepR, which upon binding QS signal molecules, activates virulence. In this work, we found that even in the absence of CepR,B. cenocepaciacan elicit a pathogenic response if phenylacetyl-CoA, an intermediate of the phenylacetic acid degradation pathway, is not produced. Instead, accumulation of phenylacetyl-CoA appears to attenuate pathogenicity. Therefore, we have discovered that it is possible to trigger virulence in the absence of CepR, challenging the classical view of activation of virulence by this QS mechanism. Our work provides new insight into the relationship between metabolism and virulence in opportunistic bacteria. We propose that, in the event that QS signaling molecules cannot accumulate to trigger a pathogenic response, a metabolic signal can still activate virulence inB. cenocepacia.