The replication and stable-inheritance functions of IncP-9 plasmid pM3 The GenBank accession number for the sequence reported in this paper is AF078924.

Abstract

Little is known of the transfer and maintenance machinery of the IncP-9 plasmids, which are found in Pseudomonas spp. and include both degradative and resistance plasmids. One such plasmid, pM3, which confers resistance to streptomycin and tetracycline, was found repeatedly in Pseudomonas species from numerous locations in Belarus. pM3 has a broad host range, but is unable to replicate in enterobacteria at 37 SC and above. A mini derivative, pMT2, was constructed by partial PstI digestion and ligation with a fragment encoding Km R . The complete sequence of pMT2 was determined. Analysis of its 8526 bp of pM3 DNA revealed several ORFs whose predicted polypeptide products were found to have similarity to previously analysed proteins involved in plasmid replication (rep gene), transfer (mpf ; mating-pair formation gene) and stable maintenance (par, mrs genes). The organization of these genes showed similarity to several plasmid systems including the Ti and pSYM plasmids as well as IncP-1 plasmids. Subcloning narrowed down the region required for replication, and identified the putative rep gene and putative par promoter region as able to express incompatibility. rep deletion mutants were lost from the cell line, and expression of the rep gene was shown to be controlled by negative autoregulation. A pMT2 derivative with an insertion between the rep and par genes showed very weak, if any, ability to replicate autonomously, suggesting that plasmid maintenance may depend on a close interaction of rep and par functions.

Diversity of IncP-9 plasmids of Pseudomonas

“…The PCR products were sequenced ( oriV and rep sequences of pFKY1 were provided by M. Tsuda, Tohoku University) and multiple sequence alignments were created using UWGCG and clustal _ x software (excluding the primer sequences). The rep – oriV sequence from Bordetella bronchiseptica plasmid pBBR1 (co-ordinates 1850–2635), which is distantly related to IncP-9 plasmids but compatible with pMT2 (Greated et al , 2000), was used as an outgroup. The resulting neighbour-joining rep and oriV IncP-9 phylogenies (Fig.…”

Genomic and Functional Analysis of the IncP-9 Naphthalene-Catabolic Plasmid NAH7 and Its Transposon Tn 4655 Suggests Catabolic Gene Spread by a Tyrosine Recombinase

“…This closer phylogenetic relationship of NAH7 to pDTG1 than to pWW0 was also clear from the comparison of the Rep proteins of the three plasmids: 96% and 78% amino acid identities between NAH7 and pDTG1 and between NAH7 and pWW0, respectively. As is the case in other IncP-9 plasmids, such as pDTG1, pWW0, and pM3 (10,18,19), the putative origins of plasmid transfer (oriT) and replication (oriV) were situated between traA and traD and between rep and rsv, respectively ( Fig. 1A and the table in the supplemental material).…”

Identification and Characterization of the Conjugal Transfer Region of the pCg1 plasmid from Naphthalene-Degrading Pseudomonas putida Cg1

“…2 and Table 3, there is considerable similarity in both the order and sequence of many open reading frames in the putative transfer regions in pCg1, pDTG1, pWW0 (TOL plasmid in P. putida; IncP-9 incompatibility group [15]), and pM3 (IncP-9 incompatibility group; found in 16 isolates of P. putida species originating from farmland soil in Belarus, from industrial site soil in Belarus and Azerbaijan, and from sewage from a pharmaceutical plant in Belarus [16]). Nine of the 10 genes in the mpf cluster in pDTG1/pCg1 have their closest homologue (65 to 87% identity) in pWW0.…”