The IncP-6 Plasmid Rms149 Consists of a Small Mobilizable Backbone with Multiple Large Insertions

Abstract

Plasmid Rms149, the archetype of Pseudomonas plasmid incompatibility group IncP-6, was identified in Pseudomonas aeruginosa as an agent conferring resistance to streptomycin, sulfanilamide, gentamicin, and carbenicillin in 1975. It has been classed as a broad-host-range plasmid due to its ability to replicate in both Escherichia coli (where it is designated IncG) and Pseudomonas species, although both species are ␥-proteobacteria. To provide reference information on this Inc group, we have determined the complete sequence of Rms149 and found that, although the genome comprises 57,121 bp, it is essentially a small mobilizable plasmid carrying multiple mobile elements, which make up 79% (>45 kb) of its genome. A replicon has been identified which encodes a single polypeptide with moderate identity to other replication proteins. The region encoding this protein can replicate in Pseudomonas putida and E. coli. This sequence is directly downstream of a putative partitioning region highly similar to that of pRA2. A functional IncQ-type mobilization region is also present. Thus, the backbone appears to be a novel combination of modules already identified in other plasmid systems. Analysis of the segments that fall outside this core of stable inheritance and transfer functions show that this plasmid has been subject to multiple insertion events and that the plasmid appears to carry a considerable load of DNA that no longer should be phenotypically advantageous. The plasmid therefore functions not just as a vehicle for spread of selective traits but also as a store for DNA that is not currently under selection.The key features of a plasmid are the ability to replicate autonomously and to be maintained in a cell lineage without a high rate of segregational loss. An optional but frequently encountered property is the ability to transfer or to be mobilized from one bacterium to another. From these properties it follows that genes that become associated with a plasmid may spread rapidly from one genetic background to another. Some plasmids appear to consist of nothing more than functions that confer the above core abilities, and these have been termed cryptic plasmids.To succeed as such a selfish element, a plasmid must be stable and confer minimal burden on its host, or at least overcome competitive losses by horizontal transfer (42). Conversely, a plasmid carrying genes that promote the growth of its host, relative to competitor bacteria, will benefit by increased propagation. However, since maintenance of a plasmid is generally found to place a metabolic or phenotypic load on the cell, universally useful genes will be selectively reacquired by the chromosome over evolutionary time. Therefore, successful plasmids typically carry payload genes, favorable in some environments but not others (12), or genes with a transient plasmid association that are spreading through a microbial community. In passing through different strains or species, plasmids are exposed to different genetic contexts and provide the opportunity for comb...

Comparative Biology of Two Natural Variants of the IncQ-2 Family Plasmids, pRAS3.1 and pRAS3.2

“…Despite the Tn1012 insertion in orfX, the 5.6-kb mobilization region of Rms149 remained functional (18), but without an untruncated orfX the effect of orfX on mobilization frequency could not be tested. We therefore tested whether the insertion of EZ-Tn5 into orf3 of pRAS3.1, which was used to indentify orf3 as the cause of incompatibility, affected the mobilization frequency.…”

“…orf3 is 753 bp long and encodes a predicted protein of 250 aa that is preceded by a putative ribosomebinding site (GGAGG) 5 bp upstream of the ATG start. A BLAST analysis against the nonredundant NCBI protein database indicated two strong hits of 98 and 97% amino acid identity along the entire length of a hypothetical protein (240 aa) from an uncultured bacterium and to ORFX (163 aa) of plasmid Rms149 from Pseudomonas aeruginosa (18). Rms149 is a 57-kb IncP-6 plasmid that has a different replicon from IncQ plasmids but a 5.6-kb mobilization region that is very similar to that of the pRAS3 plasmids and pTF-FC2.…”

Pyocin S2 (Sa) Kills Pseudomonas aeruginosa Strains via the FpvA Type I Ferripyoverdine Receptor

“…The fragment also contained the genes PA1152 (encoding a hypothetical protein) and PA1153 (carrying the COG3617 prophage antirepressor motif), as annotated in the P. aeruginosa PAO1 genome (http://www.pseudomonas.com). Via a BLASTX analysis, we found an incomplete reading frame (ЈorfA) corresponding to part of the ofn44 gene (Q4W1R3_ PSEAE) from the IncP-6 plasmid Rsm149 (15) in the intergenic region between the imm2 gene and the PA1152 gene, which codes for a conserved hypothetical protein corresponding to the Tn3 family of transposons (Fig. 1).…”

Complete Nucleotide Sequence of TOL Plasmid pDK1 Provides Evidence for Evolutionary History of IncP-7 Catabolic Plasmids

“…Transposon Tn4662 was very closely related to Tn5501-Tn5502 (27) and Tn5503 (18) in terms of gene content and sequence similarity. Tn4662 differed from Tn5503 on Rms149, an IncP-6 Pseudomonas plasmid, only in the repeat number of the 7-bp sequence (5Ј-CCCAGAG-3Ј) in ofn19; the repeat number was 16 in Tn4662 and 9 in the corresponding gene of Tn5503.…”