Sphingosine induction of thePseudomonas aeruginosahemolytic phospholipase C/sphingomyelinase, PlcH

Abstract

AbstractThe hemolytic phospholipase C, PlcH, is an important virulence factor forPseudomonas aeruginosa. PlcH preferentially hydrolyzes sphingomyelin and phosphatidylcholine and this hydrolysis activity can drives tissue damage, inflammation, and interferes with the oxidative burst of immune cells. Among other contributors, transcription ofplcHwas previously shown to be induced by phosphate starvation via PhoB and by the choline metabolite, glycine betaine, via GbdR. Here, we show that sphingosine can induceplcHtranscription and resultant secreted PlcH enzyme activity. This induction is dependent on the sphingosine-sensing transcription regulator SphR. The SphR induction ofplcHoccurs from the promoter for the gene upstream ofplcHthat encodes the neutral ceramidase, CerN, and transcriptional read-through of thecerNtranscription terminator. Evidence for these conclusions come from mutation of the SphR binding site in thecerNpromoter, mutation of thecerNterminator, enhancement ofcerNtermination by adding therrnBterminator, and RT-PCR showing that the intergenic region betweencerNandplcHis made as RNA during sphingosine, but not choline, induction. We also observe that, like glycine betaine induction, sphingosine induction ofplcHis under catabolite repression control, which likely explains why such induction was not seen in other studies using sphingosine in rich media. The addition of sphingosine as a novel inducer for PlcH points to regulation ofplcHtranscription as a site for integration of multiple host-derived signals.