Expression of adhesive pili and the collagenbinding adhesin Ace is activated by ArgR family transcription factors in Enterococcus faecalis

It was shown previously that the disruption of the ahrC gene encoding a predicted ArgR family transcription factor results in a severe defect in biofilm formation in vitro, as well as a significant attenuation of virulence of Enterococcus faecalis strain OG1RF in multiple experimental infection models. Using transcriptome sequencing (RNA-seq), we observed ahrC-dependent changes in the expression of more than 20 genes. AhrC-repressed genes included predicted determinants of arginine catabolism and several other metabolic genes and predicted transporters, while AhrC-activated genes included determinants involved in the production of surface protein adhesins. Most notably, the structural and regulatory genes of the ebp locus encoding adhesive pili were positively regulated, as well as the ace gene, encoding a collagen-binding adhesin. Using lacZ transcription reporter fusions, we determined that ahrC and a second argR transcription factor gene, argR2, both function to activate the expression of ebpR, which directly activates the transcription of the pilus structural genes. Our data suggest that in the wild-type E. faecalis, the low levels of EbpR limit the expression of pili and that biofilm biomass is also limited by the amount of pili expressed by the bacteria. The expression of ace is similarly enhanced by AhrC and ArgR2, but ace expression is not dependent on EbpR. Our results demonstrate the existence of novel regulatory cascades controlled by a pair of ArgR family transcription factors that might function as a heteromeric protein complex.