The transcriptional regulator Lrp activates the expression of genes involved in the biosynthesis of tilimycin and tilivalline enterotoxins in .

The toxigenic strains secrete tilymicin and tilivalline enterotoxins, which cause antibiotic-associated hemorrhagic colitis. Both enterotoxins are non-ribosomal peptides synthesized by enzymes encoded in two divergent operons clustered in a pathogenicity island. The transcriptional regulator Lrp (eucine-responsive egulatory rotein) controls the expression of several bacterial genes involved in virulence. In this work, we have uncovered novel findings that have significant implications. We determined the transcriptional expression of and , the first genes of each tilimycin (TM)/tilivalline (TV) biosynthetic operon in MIT 09-7231 wild-type and its derivatives Δ mutant and complemented strains. Our results suggest that Lrp directly activates the transcription of both and genes by binding to the intergenic regulatory region in a leucine-dependent manner. Furthermore, the lack of Lrp significantly diminished the cytotoxicity of on HeLa cells due to reduced production of TM and TV. Altogether, our data present a new perspective on the role of Lrp as a regulator in cytotoxin-producing strains and how it controls the expression of genes involved in the biosynthesis of their main virulence factors.IMPORTANCETilimycin (TM) and tilivalline (TV) are enterotoxins that are a hallmark for the cytotoxin-producing strains, which cause antibiotic-associated hemorrhagic colitis. The biosynthesis of TM and TV is driven by enzymes encoded by the - and NRPS-operons. In this study, we discovered that the transcriptional regulator Lrp plays a crucial role in activating the expression of the - and NRPS-operons, thereby initiating TM and TV biosynthesis. Our results underscore a molecular mechanism by which TM and TV production by toxigenic strains is regulated and shed further light on developing strategies to prevent the intestinal illness caused by this enteric pathogen.