Abstract
Mobile genetic elements are widespread in Pseudomonas syringae, and often associate with virulence genes. Genome reannotation of the model bean pathogen P. syringae pv. phaseolicola 1448A identified seventeen types of insertion sequences and two miniature inverted-repeat transposable elements (MITEs) with a biased distribution, representing 2.8% of the chromosome, 25.8% of the 132-kb virulence plasmid and 2.7% of the 52-kb plasmid. Employing an entrapment vector containing sacB, we estimated that transposition frequency oscillated between 2.6×10−5 and 1.1×10−6, depending on the clone, although it was stable for each clone after consecutive transfers in culture media. Transposition frequency was similar for bacteria grown in rich or minimal media, and from cells recovered from compatible and incompatible plant hosts, indicating that growth conditions do not influence transposition in strain 1448A. Most of the entrapped insertions contained a full-length IS801 element, with the remaining insertions corresponding to sequences smaller than any transposable element identified in strain 1448A, and collectively identified as miniature sequences. From these, fragments of 229, 360 and 679-nt of the right end of IS801 ended in a consensus tetranucleotide and likely resulted from one-ended transposition of IS801. An average 0.7% of the insertions analyzed consisted of IS801 carrying a fragment of variable size from gene PSPPH_0008/PSPPH_0017, showing that IS801 can mobilize DNA in vivo. Retrospective analysis of complete plasmids and genomes of P. syringae suggests, however, that most fragments of IS801 are likely the result of reorganizations rather than one-ended transpositions, and that this element might preferentially contribute to genome flexibility by generating homologous regions of recombination. A further miniature sequence previously found to affect host range specificity and virulence, designated MITEPsy1 (100-nt), represented an average 2.4% of the total number of insertions entrapped in sacB, demonstrating for the first time the mobilization of a MITE in bacteria.
Highlights
Insertion sequences (IS) are probably the simplest autonomous mobile DNA elements and generally consist of a transposase coding gene, responsible for their transposition, which is bound by terminal inverted repeats [1,2]
P. syringae pv. phaseolicola 1448A contains at least seventeen insertion sequences and two miniature inverted-repeat transposable elements (MITEs)
A comparison of the genome of P. syringae pv. phaseolicola 1448A with the databases indicates that it harbors at least seventeen different types of insertion sequences and two MITEs (Table 1 and Annotation files S1, S2 and S3)
Summary
Insertion sequences (IS) are probably the simplest autonomous mobile DNA elements and generally consist of a transposase coding gene, responsible for their transposition, which is bound by terminal inverted repeats [1,2]. ISs can generate significant variability in bacteria and contribute to their evolution [3], in part because they are usually present in more than one copy per genome and represent mobile regions of recombination Their mobility, together with their capacity to mobilize unrelated DNA in their proximity, can lead to a panoply of mutations and rearrangements in the host bacteria, which include insertions, deletions, duplications, translocations, cointegrations, inversions and gene activation [4]. From these activities, it follows that they have an enormous potential to alter the genome and influence bacterial evolution. The impact of MITE activity in the prokaryotic genome is potentially very high and their small size allows them to contribute to phenotypic variation in many different and creative ways, such as generating new gene alleles and functions, or new regulatory signals for preexisting genes [9]
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