INVOLVEMENT OF ARGR AND PEPA IN THE PAIRING OF COLE1 DIMER RESOLUTION SITES

Dimer formation and associated copy number depression is an important cause of multicopy plasmid instability. Natural multicopy plasmids employ site-specific recombination to convert dimers to monomers, thus maximizing the number of independently segregating molecules at cell division. Resolution of dimers of Escherichia coli plasmid ColE1 requires the plasmid cer site and at least four chromosome-encoded proteins: the XerC and XerD recombinases, and accessory factors ArgR and PepA. It has been suggested that ArgR has a role in the initial pairing of recombination sites and we describe here an attempt to detect this process in vivo. Our approach exploits a previous observation that a cer-like site known as the type II hybrid supports inter-molecular recombination and causes extensive multimerization of plasmids. We report that type-II-mediated multimerization can be repressed by a cer site in cis or in trans and propose that this is due to a physical interaction between the sites. If this hypothesis is correct, suppression of multimer formation provides an assay of site pairing. Our results demonstrate that the putative pairing interaction is independent of the topological relationship of the sites and that both PepA and ArgR are involved. Although most recombination-deficient mutant derivatives of ArgR are unable to pair recombination sites, we have found two (ArgR110 and ArgR115*) which retain pairing activity. The validity of the pairing hypothesis is discussed in the light of alternative explanations for our data.