Supportive and inhibitory elements of a putative PrfA-dependent promoter in Listeria monocytogenes

Elements essential for PrfA-dependent transcription were analysed on two promoters of Listeria monocytogenes, the PrfA-dependent promoter of the phospholipase gene plcA (PplcA) and a putative promoter of the aroA gene (ParoA2) which contains a similar PrfA-binding site and a similar -10 box as PplcA but does not function as PrfA-dependent promoter. We constructed a series of hybrid plcA-aroA promoters by exchanging corresponding sequence elements of these two 'promoters'. The results showed that the two critical elements of PrfA-dependent promoters, the PrfA-box and the -10 box, can be functionally exchanged as long as the distance in between is maintained to 22 or 23 bp. However, the interspace sequence and the sequence downstream of the -10 box of ParoA2 were strongly inhibitory for PrfA-dependent transcription. A detailed analysis of these two sequences revealed that the RNA polymerase binding site being part of the actual in vivo and in vitro used aroA promoter (ParoA1) and a sequence immediately downstream of the putative -10 site, possibly blocking the formation of the open complex, were responsible for the inhibition of PrfA-dependent transcription from ParoA2. Taking into consideration the lessons learned from this study we were able to construct a functional PrfA-dependent aroA promoter.