Mutual control of the PecS/PecM couple, two proteins regulating virulence-factor synthesis in Erwinia chrysanthemi

The Erwinia chrysanthemi pecS mutant displays constitutive production of virulence factors, such as pectinases or cellulases. Complementation of the pecS mutation can be obtained in the presence of the pecS wild-type gene on a low-copy-number plasmid. Moreover, the resulting plasmid decreases the expression of a pecS::uidA chromosomal fusion, indicating the existence of an autoregulation mechanism. This negative autoregulation was confirmed and quantified by analysis of the pecS transcripts using primer-extension experiments. Band-shift assays and DNase I footprinting experiments demonstrated that the PecS protein could bind to the intergenic regulatory region, located between the pecS and pecM genes, with a relatively high affinity (apparent dissociation constant (K'[d]) close to 4nM). These PecS-binding sites overlap the pecS and pecM promoters. The comparison of these new PecS-binding sites with those previously characterized on the target genes confirms the absence of a consensus. This observation was in accordance with the results of the missing-contact experiments performed on the pecS-pecM intergenic regulatory region and the celZ operator. Concurrently, we demonstrated that the PecS protein negatively controls the expression of the divergently transcribed pecM gene located 400bp upstream from the pecS gene. By following the efficiency of pecS autoregulation in a double E. chrysanthemi pecM-pecS mutant, we established that the PecM protein potentiates PecS activity in vivo.