The dif/Xer recombination systems in proteobacteria - PubMed (original) (raw)

The dif/Xer recombination systems in proteobacteria

Christophe Carnoy et al. PLoS One. 2009.

Abstract

In E. coli, 10 to 15% of growing bacteria produce dimeric chromosomes during DNA replication. These dimers are resolved by XerC and XerD, two tyrosine recombinases that target the 28-nucleotide motif (dif) associated with the chromosome's replication terminus. In streptococci and lactococci, an alternative system is composed of a unique, Xer-like recombinase (XerS) genetically linked to a dif-like motif (dif(SL)) located at the replication terminus. Preliminary observations have suggested that the dif/Xer system is commonly found in bacteria with circular chromosomes but that assumption has not been confirmed in an exhaustive analysis. The aim of the present study was to extensively characterize the dif/Xer system in the proteobacteria, since this taxon accounts for the majority of genomes sequenced to date. To that end, we analyzed 234 chromosomes from 156 proteobacterial species and showed that most species (87.8%) harbor XerC and XerD-like recombinases and a dif-related sequence which (i) is located in non-coding sequences, (ii) is close to the replication terminus (as defined by the cumulative GC skew) (iii) has a palindromic structure, (iv) is encoded by a low G+C content and (v) contains a highly conserved XerD binding site. However, not all proteobacteria display this dif/XerCD system. Indeed, a sub-group of pathogenic epsilon-proteobacteria (including Helicobacter sp and Campylobacter sp) harbors a different recombination system, composed of a single recombinase (XerH) which is phylogenetically distinct from the other Xer recombinases and a motif (dif(H)) sharing homologies with dif(SL). Furthermore, no homologs to dif or Xer recombinases could be detected in small endosymbiont genomes or in certain bacteria with larger chromosomes like the Legionellales. This raises the question of the presence of other chromosomal deconcatenation systems in these species. Our study highlights the complexity of dif/Xer recombinase systems in proteobacteria and paves the way for systematic detection of these components in prokaryotes.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Nucleotide variability within _dif_-related sequences.

(A) Consensus sequence and dif nucleotide variability for 161 dif_-related sequences from 137 proteobacterial species. Nucleotide sequence characters in bold represent the dif sequence (28-mer). If the nucleotide frequency represents more than 50%, it is written in upper case letters; if not, the nucleotide is written in lower case letters. The nucleotide variability at each position in the 28-mer was defined as 1–_f, where f is the frequency of the most frequent nucleotide. Nucleotide frequencies at each position are given in Table S2. Black bars represent dif XerC and dif XerD nucleotides, whereas grey bars correspond to the the dif cent nucleotides. White bars represent nucleotides outside dif. (B) Degree of variability in the dif sequence in 21 multi-strain species and in 19 multi-chromosome species. The degree of variability was calculated for each nucleotide position, as described in the Methods section.

Figure 2. Phylogeny of proteobacterial XerC and XerD recombinases.

Representative proteobacterial species of each taxon were selected for the analysis (Table 1). β-proteobacterial species are represented in blue, with γ in red, δ in green, α in magenta and ε in black. Amino acid sequence alignments were performed using Clustal W (MEGA 4 [60]). The evolutionary history was inferred by using the Neighbor-Joining method conducted in MEGA4. Similar results were obtained using the Minimum Evolution method (data not shown). Only significant bootstrap values (≥90%) obtained with 1000 runs are indicated next to the branches (white with a grey background). The tree is drawn to scale, with branch lengths (below the branches) in the same units as those of the evolutionary distances used to infer the phylogenetic tree. Branch lengths below the value 0.05 are not shown. The evolutionary distances were computed using the Poisson correction method and are given as the number of amino acid substitutions per site.

Figure 3. Correlation between the position of the dif sequence and the terminus of replication as defined by cumulative GC skew.

The analysis was performed on the 161 proteobacterial chromosomes from the 137 representative dif + species (Table S1). Chromosome of Wolbachia endosymbiont of Drosophila melanogaster and chromosome 2 of Pseudoalteromonas haloplanktis were not included in the analysis since no terminus of replication could be located for these species by the method of the cumulative GC skew. The equation of the plot and the coefficient of determination (R2) are given.

Figure 4. Phylogenetic analysis of XerC, XerD, XerH and XerS recombinases.

XerH from the ε subgroup species (listed in Table 2) were compared with XerD and XerC recombinases from other ε species and representative bacteria from the α, β, δ and γ taxa (Table 1). XerS recombinases of S. pyogenes M1 GAS and L. lactis Il1403 were added for comparison. Amino acid sequence alignment (with Clustal W) and phylogenetic analyses were performed in MEGA4 . The phylogeny was built using the Neighbor-Joining method . The tree is drawn to scale, with branch lengths in the same units as those of the evolutionary distances used to infer the phylogenetic tree. The evolutionary distances were computed using the Poisson correction method and are in the units of the number of amino acid substitutions per site. The size range of the recombinases (in amino acids) is indicated under the recombinase name, in brackets.

Figure 5. Alignment of difH and difSL.

The difH sequence corresponds to the putative dif motif of H. pylori 26695 (Table 2), whereas difSL was described by Le Bourgeois et al. . Asterisks indicate the common nucleotides and arrows designate inverted repeats.

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The dif/Xer recombination systems in proteobacteria - PubMed (original) (raw)