A Single Gene on the Staphylococcal Multiresistance Plasmid pSK1 Encodes a Novel Partitioning System

Abstract

The orf245 gene is located immediately upstream of, and divergently transcribed from, the replication initiation gene, rep, of the Staphylococcus aureus multiresistance plasmid pSK1, and related genes have been found in association with a range of evolutionarily distinct replication genes on plasmids from various gram-positive genera. orf245 has been shown previously to extend the segregational stability of a pSK1 minireplicon. Here we describe an investigation into the basis of orf245-mediated stabilization. orf245 was not found to influence transcription of pSK1 rep, indicating that it is not directly involved in plasmid replication. This was confirmed by demonstrating that orf245 is able to enhance the segregational stability of heterologous theta-and rolling-circle-replicating replicons, suggesting that it encodes a plasmid maintenance function. Evidence inconsistent with postsegregational killing and multimer resolution mechanisms was obtained; however, the intergenic region upstream of orf245 was found to mediate orf245-dependent incompatibility, as would be expected if it encodes a cis-acting centromere-like site. Taken together, these findings implicate active partitioning as the probable basis of the activity of orf245, which is therefore redesignated par. Since it is unrelated to any gene known to play a role in plasmid segregation, it seems likely that pSK1 par potentially represents the prototype of a novel class of active partitioning systems that are distinguished by their capacity to enhance plasmid segregational stability via a single protein-encoding gene.

Sequence analysis of the mobilizable lactococcal plasmid pGdh442 encoding glutamate dehydrogenase activity

“…Indeed, Foley et al (1996) have demonstrated the incompatibility of two plasmids with the same ori site by providing three and a half copies of the 22 bp sequence of the pCI305 ori site in trans on a compatible vector, which leads to the loss of pCI305 from the cell population. In addition, partitionmediated incompatibility occurs when two plasmids introduced into the same cell carry identical centromerelike sites, which are indistinguishable from each other during the partitioning process, as previously reported in the Staphylococcus pSK1 plasmid (Simpson et al, 2003). The incompatibility of pGdh442 with pSK08-type plasmids will probably prevent the transfer of pGdh442 in most strains capable of growing in milk, since out of 15 tested prtPcontaining strains, 12 contained the same replication and partition genes as pSK08, as revealed by PCR amplification with specific primers (Table 1) derived from the ori, repA, parA and parB sequences of pSK08 (data not shown).…”

Tubulin homolog TubZ in a phage-encoded partition system

“…Furthermore, the Par protein, responsible for the stabilization of plasmid pSK1 in Staphylococcus spp. (24), could be a TubY remote homolog because it shares both the HTH and coiled-coil motifs and 45% identity at the C-terminal tail with TubY, but this plasmid lacks TubZ-R homologs. but does not carry any essential genes for C. botulinum survival.…”

Toxin–antitoxin systems are ubiquitous and plasmid-encoded in vancomycin-resistant enterococci

“…The stability of the vancomycin-resistant plasmid in the transconjugant, S345RF, was studied in triplicate, as described, with some modifications (73). One hundred microliters of an overnight culture of the strain grown in BHI broth containing 10 g/ml vancomycin was used to inoculate 10 ml of fresh BHI broth containing 10 g/ml vancomycin and grown for 4 h. Dilutions of the culture were prepared in sterile H 2 O and plated onto nonselective BHI agar plates for viable counts.…”