Examination of E. faecalis toxin-antitoxin (TA) toxin Fst function utilizing a pheromone-inducible expression vector with tight repression and large dynamic range. Enterococcus faecalis

Tools for regulated gene expression in E. faecalis are extremely limited. In this study we describe the construction of an expression vector for E. faecalis, designated pCIE, utilizing the PQ pheromone-responsive promoter of plasmid pCF10. We demonstrate that this promoter is tightly repressed, responds to nanogram quantities of the peptide pheromone, and has a large dynamic range. To demonstrate its utility, the promoter was used to control expression of the toxic peptides of two par family toxin-antitoxin loci present in E. faecalis, parpAD1 of the pAD1 plasmid and parEF0409 located on the E. faecalis chromosome. Results demonstrated differences in the modes of regulation of toxin expression and in the effects of toxins of these two related systems. We anticipate that this vector will be useful for further investigation of par TA system function as well as the regulated expression of other genes in E. faecalis. Overall design: Examination of cellular effects of toxin FstEF0409 at two different levels compared to unexposed cells and comparison of the effects of the two related toxins FstEF0409 and FstpAD1. Average of two biological replicates.