Host-specific incompatibility by 9-bp direct repeats indicates a role in the maintenance of broad-host-range plasmid...

Broad-host-range incompatibility group P (IncP) plasmids RK2 and R751 have 9-bp direct repeats (DR) of unknown function located between their kilC and kilE loci. The nucleotide sequences of the 9-bp repeats are different for RK2 (an IncPalpha group plasmid) and R751 (IncPbeta group), but both DR regions are organized similarly, including an 11-bp spacer with identical 5'-CGCCA-3' cores and an adjacent binding site for KorB, a known partition protein and transcriptional repressor. The occurrence of similarly arranged DR elements with different repeat sequences is suggestive of an important plasmid-specific function for the DR regions. Here we show that the cloned RK2 DR region in trans to RK2 exhibits a host-specific incompatibility phenotype, in which RK2 is destabilized in Pseudomonas aeruginosa but not in Escherichia coli. Incompatibility was not dependent on the adjacent KorB-binding site. Deletion of the kilE locus, which is required for stable maintenance in P. aeruginosa, did not abolish DR-mediated incompatibility. Precise deletion of DR from RK2 had no effect on maintenance but eliminated sensitivity to DR in trans, showing that incompatibility requires DR to be present on both plasmids. These results raise the possibility that the DR region may be involved in a plasmid maintenance system for P. aeruginosa that is independent of the known stability functions on RK2.