The TetR-Type MfsR Protein of the Integrative and Conjugative Element (ICE) ICE clc Controls both a Putative Efflux System and Initiation of ICE Transfer

Integrative and conjugating elements (ICE) are self-transferable DNAs widely present in bacterial genomes, which often carry a variety of auxiliary genes of potential adaptive benefit. One of the model ICE is ICEclc , an element originally found inPseudomonas knackmussii B13 and known for its propensity to provide its host with the capacity to metabolize chlorocatechols and 2-aminophenol. In this work, we studied the mechanism and target of regulation of MfsR, a TetR-type repressor previously found to exert global control on ICEclc horizontal transfer. By using a combination of ICEclc mutant and transcriptome analysis, gene reporter fusions, and DNA binding assays, we found that MfsR is a repressor of both its own expression and that of a gene cluster putatively coding for a major facilitator superfamily efflux system on ICEclc (namedmfsABC ). Phylogenetic analysis suggests thatmfsR was originally located immediately adjacent to the efflux pump genes but became displaced from its originalcis target DNA by a gene insertion. This resulted in divergence of the original bidirectional promoters into two separated individual regulatory units. Deletion ofmfsABC did not result in a strong phenotype, and despite screening a large number of compounds and conditions, we were unable to define the precise current function or target of the putative efflux pump. Our data reconstruct how the separation of an ancestormfsR-mfsABC system led to global control of ICEclc transfer by MfsR.