MECHANISM OF DNA SEGREGATION IN PROKARYOTES - REPLICON PAIRING BY PARC OF PLASMID R1

Prokaryotic chromosomes and plasmids encode partitioning systems that are required for DNA segregation at cell division. The systems are tho ught to be functionally analogous to eukaryotic centromeres and to pla y a general role in DNA segregation. The parA system of plasmid R1 enc odes two proteins ParM and ParR, and a cis-acting centromere-like site denoted parC. The ParR protein binds to parC in vivo and in vitro. Th e ParM protein is an ATPase that interacts with ParR specifically boun d to parC. Using electron microscopy, we show here that parC mediates efficient pairing of plasmid molecules. The pairing requires binding o f ParR to parC and is stimulated by the ParM ATPase. The ParM mediated stimulation of plasmid pairing is dependent on ATP hydrolysis by ParM . Using a ligation kinetics assay, we find that ParR stimulates ligati on of parC-containing DNA fragments. The rate-of-ligation was increase d by wild type ParM protein but not by mutant ParM protein deficient i n the ATPase activity. Thus, two independent assays show that parC med iates pairing of plasmid molecules in vitro. These results are consist ent with the proposal that replicon pairing is part of the mechanism o f DNA segregation in prokaryotes.