Replication regions from plant-pathogenic Pseudomonas syringae plasmids are similar to ColE2-related replicons

Abstract

Many strains of the phytopathogen Pseudomonas syringae contain mutually compatible plasmids that share extensive regions of sequence homology and essential replication determinants. The replication regions of two compatible large plasmids involved in virulence or pathogenicity, pPT23A from P. syringae pv. tomato strain PT23 and pAV505 from P. syringae pv. phaseolicola strain HR11302A, were isolated. DNA sequencing of the origins of replication revealed homologous ORFs, designated ORF-Pto and ORF-Pph, respectively. Both ORFs are 1311 bp long and encode peptides of 437 amino acids with predicted molecular masses of 48259 (Pto) and 48334 (Pph) Da. Expression of the two ORFs in Escherichia coli produced peptides of 50 kDa (Pto) and 56 kDa (Pph). The predicted peptides showed an overall identity of 897 %, being highly conserved from residues 1 to 373, but showing considerable variation in their C-terminal regions (50% identity over the last 64 aa). The two ORFs had significant similarity with the putative replication protein from plasmid pTiKl2 of Thiobacillus intermedius and other ColE2-related plasmids. However, both peptides were 100 residues longer than any of the known ColE2-related rep sequences. Subcloning of fragments from the replication region of pPT23A revealed the presence of a t least three incompatibility determinants, designated IncA, lncB and IncC. Partial sequencing of the region downstream of ORF-Pto revealed homology to the rulAB genes, involved in UV resistance, from plasmid pPSR1. It is proposed that the replication origin of pPT23A serves as the prototype of a family of related plasmids.

Genetic and pathogenic diversity of Pseudomonas syringae strains isolated from cucurbits

“…There are publications that suggest pathogenicity islands are plasmidborne (Jackson et al 1999;Gibbon et al 1999;Ma et al 2007). However, in this study some highly virulent strains had no plasmid, and some non-virulent strains had a plasmid (Fig.…”

Phylogeny of the replication regions of pPT23A-like plasmids from Pseudomonas syringae The EMBL accession numbers for the sequences reported in this paper are AJ276998–AJ277021.

“…The repA gene from pAV505 was not cloned because its complete sequence was already available (accession no. AJ222648 ; Gibbon et al, 1999). The amplicon cloned from total plasmid DNA from strain OK-1 was confirmed to be derived from plasmid pOK1B, by comparison of the restriction profile of repA amplicons from purified DNA of each of the two native plasmids of OK-1 with that of the cloned amplicon.…”

“…The majority of the native plasmids identified in P. syringae belong to the recently described pPT23A-like family ; these plasmids share replication sequences and, in most cases, additional areas of homology Glickmann et al, 1998 ;Sesma et al, 1998 ;Gibbon et al, 1999). Furthermore, many P. syringae strains contain two to six coexisting pPT23A-like plasmids Sesma et al, 1998), suggesting their potential role in the bacterial life cycle and underlying their capacity to overcome incompatibility.…”

“…A 1399 bp fragment containing 1279 bp of the 1311 bp repA coding region plus 120 bp upstream of the putative start codon ( Fig. 1 ; Gibbon et al, 1999) was amplified from the pPT23A-like plasmids and separately cloned in vector pCR2.1 which, in our hands, did not replicate in P. syringae. All the cloned amplicons, except those originating from p5D425A and pB76A, were able to replicate in the plasmidless strain P. syringae pv.…”

Temperature-responsive genetic loci in the plant pathogen Pseudomonas syringae pv. glycinea The GenBank accession numbers for the nucleotide sequences of mutants 560, 561, 562, 563, 564, 568, 570, 574, 590, 591, 593, 596, 599, 601, 605, 608, 613, 617, 618, 626 and 632 determined in this work are AF274322–AF274342, respectively.

“…Two additional loci, designated p601 and p632 (Table 2), showed relatedness, respectively, to the plasmid replication protein RepA of P. syringae pv. phaseolicola (Gibbon et al, 1999) and to the replication region of the P. aeruginosa plasmid R91-5 (Moore & Krishnapillai, 1982), suggesting plasmid-borne locations. Agarose gel electrophoresis of undigested plasmid preparations from mutants 632, 601 and PG4180(pRGMU3) (Fig.…”