The unconventional Xer recombination machinery of Streptococci/Lactococci - PubMed (original) (raw)
Comparative Study
The unconventional Xer recombination machinery of Streptococci/Lactococci
Pascal Le Bourgeois et al. PLoS Genet. 2007 Jul.
Abstract
Homologous recombination between circular sister chromosomes during DNA replication in bacteria can generate chromosome dimers that must be resolved into monomers prior to cell division. In Escherichia coli, dimer resolution is achieved by site-specific recombination, Xer recombination, involving two paralogous tyrosine recombinases, XerC and XerD, and a 28-bp recombination site (dif) located at the junction of the two replication arms. Xer recombination is tightly controlled by the septal protein FtsK. XerCD recombinases and FtsK are found on most sequenced eubacterial genomes, suggesting that the Xer recombination system as described in E. coli is highly conserved among prokaryotes. We show here that Streptococci and Lactococci carry an alternative Xer recombination machinery, organized in a single recombination module. This corresponds to an atypical 31-bp recombination site (dif(SL)) associated with a dedicated tyrosine recombinase (XerS). In contrast to the E. coli Xer system, only a single recombinase is required to recombine dif(SL), suggesting a different mechanism in the recombination process. Despite this important difference, XerS can only perform efficient recombination when dif(SL) sites are located on chromosome dimers. Moreover, the XerS/dif(SL) recombination requires the streptococcal protein FtsK(SL), probably without the need for direct protein-protein interaction, which we demonstrated to be located at the division septum of Lactococcus lactis. Acquisition of the XerS recombination module can be considered as a landmark of the separation of Streptococci/Lactococci from other firmicutes and support the view that Xer recombination is a conserved cellular function in bacteria, but that can be achieved by functional analogs.
Conflict of interest statement
Competing interests. The authors have declared that no competing interests exist.
Figures
Figure 1. Identification of the Streptococcal dif SL Site by Comparative Genomics
(A) Cumulative GC skew diagrams of three streptococcal genomes. Lla, L. lactis IL1403; Spyo, S. pyogenes M1 GAS; Spn, S. pneumoniae R6. (B) Multiple DNA comparison is presented for the 10-kb regions encompassing the region of GC skew shift. The result is scaled to the gene organization of the L. lactis IL1403 terC region, and the conserved ORF is indicated in black. The lactococcal ORFs A, B, and C, not found in other streptococcal genomes, showed significant homology to the YmfD protein when fused together, suggesting ancient duplication of the ymfD region in L. lactis. The conserved ∼50-bp sequence is indicated by an asterisk. (C) Sequence similarity of putative dif SL sites from 49 publicly available streptococcal genomes (December 2006) and comparison with the E. coli and B. subtilis dif sites are presented. Nucleotides conserved in all except one species are indicated in bold, and those conserved in all species are indicated in bold and underlined. S. pyogenes M1 strains were M1 GAS, MGAS10750, MGAS5005, and MGAS10394. S. pyogenes M3 strains were MGAS315, MGAS2096, MGAS10270, MGAS9429, MGAS6180, M49–591, and SSI-1. S. agalactiae sequences of strains A909, H36B, COH1, CJB111, and 18RS21 were identical to strain 2603V/R. Asterisks indicate unfinished genomes.
Figure 2. Gene Context Analysis of the 10-kb terC Region of Different Streptococcal Species
Lla, L. lactis IL1403; Spn, S. pneumoniae R6; Spyo, S. pyogenes M1 GAS; Sagal, S. agalactiae NEM316; Sth, S. salivarius subsp_. thermophilus_ CNRZ1066; Smut, S. mutans UA159. The ORF coding for the putative tyrosine recombinase is shown in black. Only putative ρ-independent transcription terminators with free energy (ΔG) <−12 kcal.mol−1 are indicated.
Figure 3. XerS/dif SL Recombination in E. coli
(A) KmR cassette excision mediated by XerS in E. coli in different genetic backgrounds is shown. The +/− signs indicated presence/absence of plasmid expressing the L. lactis xerS (pCL297) or the E. coli ftsK (pCL263) genes. When available the number of independent experiments (n) is indicated below each excision frequency mean value. Error bars correspond to the standard deviation (σn−1). (B) Effect of the lactococcal XerS/dif SL system on chromosome dimer resolution in E. coli as measured by growth competition assays is presented. E368 (XerS + dif SL +) was mixed with E367 (XerS + dif SL −, squares), E378 (XerS+ dif SL + ftsK C −, triangles), or E375 (WT strain with XerS, circles) at a ratio of 1:1 and grown in serial culture for 60 generations. Values were calculated from two independent assays.
Figure 4. Subcellular Localization of FtsKSL-GFP Proteins in L. lactis
Phase-contrast (A) and fluorescence (B) microscopy of L. lactis NZ900 overexpressing full-length lactococcal FtsK-GFP (upper) or N-ter Ftsk-GFP (lower) are presented. Cells were analyzed by microscopy on midexponential growth phase.
References
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