The transcriptional regulator Lrp activates the expression of genes involved in tilivalline enterotoxin biosynthesis in Klebsiella oxytoca

Abstract

The toxigenic Klebsiella oxytoca strains secret the tilivalline enterotoxin, which causes antibiotic-associated hemorrhagic colitis. The tilivalline is a non-ribosomal peptide synthesized by enzymes encoded in two divergent operons clustered in a pathogenicity island. The transcriptional regulator Lrp (leucine-responsive regulatory protein) controls the expression of several bacterial genes involved in virulence. In this work, we determined the transcriptional expression of aroX and npsA, the first genes of each tilivalline biosynthetic operon in K. oxytoca MIT 09-7231 wild-type and its derivatives {Delta}lrp mutant and complemented strains. The results show that Lrp directly activates the transcription of both aroX and npsA genes by binding to the intergenic regulatory region in a leucine-dependent manner. Furthermore, the lack of Lrp significantly diminished the cytotoxicity of K. oxytoca on HeLa cells due to tilivalline reduced production. Altogether, our data highlight Lrp as a new regulator by which cytotoxin-producing K. oxytoca strains control the expression of genes involved in the biosynthesis of their main virulence factor. IMPORTANCETilivalline is an enterotoxin that is a hallmark for the cytotoxin-producing K. oxytoca strains, which cause antibiotic-associated hemorrhagic colitis. The biosynthesis of tilivalline is driven by enzymes encoded by the aroX- and NRPS-operons. In this study, we discovered that the transcriptional regulator Lrp directly activates expression of the aroX- and NRPS-operons and, in turn, tilivalline biosynthesis. Our results underscore a molecular mechanism by which tilivalline production by toxigenic K. oxytoca strains is regulated and shed further light on developing strategies to prevent the intestinal illness caused by this enteric pathogen.