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

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 ( l eucine-responsive r egulatory p rotein) 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 Δ 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. IMPORTANCE Tilivalline 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.