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.
Bibliographical noteFunding Information:
The authors would like to thank Dr. Jeff Gralnick (University of Minnesota), Dr. Fitnat Yildiz (UC Santa Cruz), and Dr. Don Court (NIH) for sharing of strains. The authors thank Dr. William Navarre (University of Toronto) for sharing purified H-NS and EMSA technical assistance. Data analysis was carried out using tools available through the Minnesota Supercomputing Institute at the University of Minnesota. The primary author, KSL, was supported by a fellowship from the United States Department of Agriculture National Institute of Food and Agriculture grant no. 2013-67011-21276 . TJJ was supported through funding from the University of Minnesota College of Veterinary Medicine Signature Programs.
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