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Nucleotide sequence, functional characterization and evolution of pFKN, a virulence plasmid in Pseudomonas syringae
Laurence Rohmer1, Susanne Kjemtrup, Patrizia Marchesini
1Department of Biology, Coker Hall 108, CB#3280, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599-3280, USA.
Abstract:
Pseudomonas syringae pv. maculicola strain M6 (Psm M6) carries the avrRpm1 gene, encoding a type III effector, on a 40 kb plasmid, pFKN. We hypothesized that this plasmid might carry additional genes required for pathogenesis on plants. We report the sequence and features of pFKN. In addition to avrRpm1, pFKN carries an allele of another type III effector, termed avrPphE, and a gene of unknown function (ORF8), expression of which is induced in planta, suggesting a role in the plant-pathogen interaction. The region of pFKN carrying avrRpm1, avrPphE and ORF8 exhibits several features of pathogenicity islands (PAIs). Curing of pFKN (creating Psm M6C) caused a significant reduction in virulence on Arabidopsis leaves. However, complementation studies using Psm M6C demonstrated an obvious virulence function only for avrRpm1. pFKN can integrate and excise from the chromosome of Psm M6 at low frequency via homologous recombination between identical sequence segments located on the chromosome and on pFKN. These segments are part of two nearly identical transposons carrying avrPphE. The avrPphE transposon was also detected in other strains of P. s. pv. maculicola and in P. s. tomato strain DC3000. The avrPphE transposon was found inserted at different loci in different strains. The analysis of sequences surrounding the avrPphE transposon insertion site in the chromosome of Psm M6 indicates that pFKN integrates into a PAI that encodes type III effectors. The integration of pFKN into this chromosomal region may therefore be seen as an evolutionary process determining the formation of a new PAI in the chromosome of Psm M6.
Insights
The Pseudomonas syringae plasmid pFKN carries avrRpm1 and avrPphE effector genes, contributing to virulence. This plasmid integrates into the plant pathogen
Area of Science:
- Microbiology
- Plant Pathology
- Genetics
Background:
- Pseudomonas syringae pv. maculicola strain M6 (Psm M6) possesses the avrRpm1 gene on the pFKN plasmid.
- Type III effectors are crucial for bacterial virulence in plants.
Purpose of the Study:
- To sequence and characterize the pFKN plasmid from Psm M6.
- To investigate the role of pFKN and its genes in Psm M6 pathogenesis.
- To understand the integration and evolutionary significance of pFKN.
Main Methods:
- Plasmid sequencing and feature analysis.
- Bacterial virulence assays on Arabidopsis.
- Complementation studies.
- Analysis of transposon and integration sites.
Main Results:
- The pFKN plasmid contains avrRpm1, avrPphE, and ORF8, with ORF8 expression induced in planta.
- Curing pFKN significantly reduced Psm M6 virulence, though only avrRpm1 showed a clear virulence function upon complementation.
- pFKN integrates and excises from the Psm M6 chromosome via homologous recombination involving avrPphE-carrying transposons.
- The avrPphE transposon is present in other P. syringae strains, inserted at various chromosomal locations.
Conclusions:
- The pFKN plasmid harbors multiple type III effectors and contributes to Psm M6 virulence.
- The avrPphE transposon facilitates plasmid integration, potentially driving the evolution of new pathogenicity islands.
- The study reveals insights into the genetic basis of Pseudomonas syringae pathogenicity and evolution.