Characterization of Acr2, an H-NS-like protein encoded on A/C2-type plasmids

•Binding sites of Acr2 were defined and differentially expressed genes were identified in strains lacking Acr2.•Acr2 was found to bind multiple loci on the plasmid as well as sites on the host bacterial chromosome.•Acr2 shares a DNA binding motif with other H-NS-like homologs, critical for its function as a repressor of IncA/C conjugation.

Conjugation plays an important role in the horizontal movement of DNA between bacterial species and even genera. Large conjugative plasmids in Gram-negative bacteria are associated with multi-drug resistance and have been implicated in the spread of these phenotypes to pathogenic organisms. A/C plasmids often carry genes that confer resistance to multiple classes of antibiotics. Recently, transcription factors were characterized that regulate A/C conjugation. In this work, we expanded the regulon of the negative regulator Acr2. We developed an A/C variant, pARK01, by precise removal of resistance genes carried by the plasmid in order to make it more genetically tractable. Using pARK01, we conducted RNA-Seq and ChAP-Seq experiments to characterize the regulon of Acr2, an H-NS-like protein. We found that Acr2 binds several loci on the plasmid. We showed, in vitro, that Acr2 can bind specific promoter regions directly and identify key amino acids which are important for this binding. This study further characterizes Acr2 and suggests its role in modulating gene expression of multiple plasmid and chromosomal loci.