The repAC replication system of the Rhizobium leguminosarum pRL7 plasmid is functional: implications regarding the origin and evolution of repABC plasmids.

The repABC replication/partitioning systems are commonly found in alpha-proteobacteria plasmids and in secondary chromosomes. All of the elements required for their replication and stable maintenance are encoded within a single transcription unit: the repABC operon. The repC gene encodes an initiator protein, while RepA, RepB and centromere-like sequence (parS) direct plasmid segregation. Strains containing two or more repABC plasmids are a common feature in some alpha proteobacteria groups, indicating that the repABC plasmid family embraces several incompatibility groups. Genes encoded within repABC operons are highly dynamic: each one possess its own distinctive phylogeny and homologous recombination events are common within these operons. Additionally, alpha-proteobacterial genomes contain repAB genes not associated with the ctRNA or with repC as well as plasmids whose replication depends on a ctRNA-repC module without the participation of repAB genes. Some alphaproteobacteria have repC genes clustered with other genes that are not involved in replication/partitioning functions. These atypical associations of genes could have an important role in the origin and diversification of new plasmids. Here we evaluated the functionality and possible evolutionary consequences of one of these atypical gene associations: the repAC genes present in the Rhizobium leguminosarum plasmid pRL7. The repAC genes are organized in an operon and they are capable of sustaining replication but in an unstable manner. RepC was essential for replication, and the origin of replication resides within its coding region. In contrast, RepA plays a minor role in the negative regulation of its own transcription.