Control of Recombination Rate During Transformation of<i>Streptococcus pneumoniae</i>: An Overview

Mortier‐Barrière, Isabelle; Humbert, Odile; Martin, Bernard; Prudhomme, Marc; Claverys, Jean‐Pierre

doi:10.1089/mdr.1997.3.233

Cited by 29 publications

(30 citation statements)

References 46 publications

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“…In fact, the sequential reaction that we describe here, in which an initial RecA(Sp) protein-mediated invasion of a linear ssDNA into a homologous dsDNA is followed by a MmsA(Sp) proteinmediated branch migration of the three-stranded intermediate, is consistent with a hypothetical model for transformational recombination that was envisioned by Claverys and colleagues (12,20). However, although our results suggest that the RecA(Sp) and MmsA(Sp) protein can act together to carry out a complete strand exchange reaction between a polymeric linear ssDNA and homologous dsDNA, the efficiency of this coupled reaction is relatively low under our present reaction conditions.…”

Section: Discussionsupporting

confidence: 87%

“…One strand of the dsDNA fragment is then degraded further into smaller oligonucleotides, whereas the complementary linear ssDNA is transported through the cell wall and membrane into the cell cytosol (19). It is presumed that the RecA(Sp) protein will then mediate the assimilation of this ssDNA into a homologous region of the double-stranded S. pneumoniae chromosome (20).…”

Section: Discussionmentioning

confidence: 99%

“…The means by which the MmsA(Sp) protein would facilitate the RecA(Sp) protein-mediated transformational recombination reaction has not been clear inasmuch as it has been reported that the E. coli RecG protein inhibits the three-strand exchange activity of the RecA(Ec) protein (12)(13)20). Although our results indicate that the MmsA(Sp) protein can similarly inhibit the three-strand exchange activity of the RecA(Sp) protein, this inhibitory effect was observed only when the strand exchange reaction was carried out in the standard manner, with a circular ssDNA and a linear dsDNA as the starting substrates (Fig.…”

Section: Discussionmentioning

confidence: 99%

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Effect of the Streptococcus pneumoniae MmsA Protein on the RecA Protein-promoted Three-strand Exchange Reaction

Hedayati

Steffen

Bryant

2002

Journal of Biological Chemistry

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Streptococcus pneumoniae is a naturally transformable bacterium that is able to incorporate DNA from its environment into its own chromosome. This process, known as transformational recombination, is dependent in part on the mmsA gene, which encodes a protein having a sequence that is 40% identical to that of the Escherichia coli RecG protein, a junction-specific DNA helicase believed to be involved in the branch migration of recombinational intermediates. We have developed an expression system for the MmsA protein and have purified the MmsA protein to more than 99% homogeneity. The MmsA protein has DNA-dependent ATP hydrolysis and DNA junction-helicase activities that are similar to those of the E. coli RecG protein. The effect of the MmsA protein on the S. pneumoniae RecA protein-promoted three-strand exchange reaction was also investigated. In the standard direction (circular singlestranded (ss) DNA ؉ linear double-stranded (ds) DNA 3 linear ssDNA ؉ nicked circular dsDNA), the MmsA protein appears to promote the branch migration of partially exchanged intermediates in a direction opposite of the RecA protein, resulting in a nearly complete inhibition of the overall strand exchange reaction. In the reverse direction (linear ssDNA ؉ nicked circular dsDNA 3 circular ssDNA ؉ linear dsDNA), however, the MmsA protein appears to facilitate the conversion of partially exchanged intermediates into fully exchanged products, leading to a pronounced stimulation of the overall reaction. These results are discussed in terms of the molecular mechanism of transformational recombination.Streptococcus pneumoniae is a naturally transformable bacterium that is able to take up DNA from its environment (in the form of ssDNA) 1 and incorporate this DNA into its chromosome (1, 2). It has been proposed that this process, known as transformational recombination, has evolved as a general mechanism that allows S. pneumoniae to change its genetic composition in response to environmental changes and stresses (3). For example, transformational recombination is believed to have contributed to the recent emergence of penicillin-resistance in clinical isolates of S. pneumoniae (4,5).Genetic studies have shown that transformational recombination is dependent on the presence of the recA gene, which encodes a DNA recombinase analogous to the RecA protein from Escherichia coli (2, 6). We recently developed an expression system and purification protocol for the S. pneumoniae RecA protein (7). The purified S. pneumoniae RecA protein (RecA(Sp)) has an ATP-dependent three-strand exchange activity that is generally similar to that of the E. coli RecA protein (RecA(Ec)) (7). In the standard three-strand exchange reaction, a circular ssDNA and a homologous linear dsDNA are recombined to form a nicked circular dsDNA and a linear ssDNA. This reaction proceeds in three phases. In the first phase, RecA protein polymerizes onto the circular ssDNA (1 RecA monomer/3 nucleotides of ssDNA), forming a helical nucleoprotein filament known as the presynaptic comple...

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Section: Discussionsupporting

confidence: 87%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

Effect of the Streptococcus pneumoniae MmsA Protein on the RecA Protein-promoted Three-strand Exchange Reaction

Hedayati

Steffen

Bryant

2002

Journal of Biological Chemistry

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“…S. pneumoniae is frequently challenged by adverse ecological changes, including frequent exposure to antibiotics and the host immune system (3,11), and it is known that bacterial stress promotes the emergence and selection of hypermutable variants (7,12). Hypermutation in S. pneumoniae has been described to be the result of inactivation of the hexA and hexB genes in the Hex repair system, which is responsible for mutation avoidance through the correction of errors resulting from nucleotide misincorporation during replication (4,8). Even though the increase in mutation frequency, on average, rarely exceeds 10-fold that of the wild-type proficient control strain (4,8,10), the hex-deficient variants have a clear advantage to acquire adaptive mutations leading to antibiotic resistance (10).…”

mentioning

confidence: 99%

Frequency of Mutation to Rifampin Resistance in Streptococcus pneumoniae Clinical Strains: hexA and hexB Polymorphisms Do Not Account for Hypermutation

Morosini

Baquero

Sánchez-Romero

et al. 2003

Antimicrob Agents Chemother

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The frequency of mutation to rifampin resistance of 200 clinical Streptococcus pneumoniae isolates was examined. Two peaks were observed in the distribution, with mode frequencies of 2.5 ؋ 10 ؊7 (20% of isolates) and 2.5 ؋ 10 ؊8 . The hexA and hexB gene entire sequences were analyzed in 13 isolates. Sequences from both hypermutable and "normomutable" strains were conserved relative to that of the R6 S. pneumoniae control strain. The phenotypic Hex system proficiency, in terms of transforming efficiency, was also maintained irrespective of the variations in mutation frequency values.

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“…Could such a trapping occur in nature? Inasmuch as formation of InsDels requires only a short homologous recombination anchor, we suggested previously that any piece of homologous DNA, including a mobile genetic element (e.g., an insertion sequence, IS) present both in the incoming DNA and the recipient chromosome, can serve as a recombination anchor (5). Taking into account the high incidence of IS in the chromosome of S. pneumoniae (27,28), and the wide interspecies distribution of these elements, IS can promote the trapping of DNA from distantly related species by generating InsDels through homology-directed illegitimate recombination.…”

Section: A D-loop Resolvase In Transformation?mentioning

confidence: 99%

Homologous recombination at the border: Insertion-deletions and the trapping of foreign DNA in Streptococcus pneumoniae

Prudhomme

Libante

Claverys

2002

Proc. Natl. Acad. Sci. U.S.A.

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Integration of foreign DNA was observed in the Gram-positive human pathogen Streptococcus pneumoniae (pneumococcus) after transformation with DNA from a recombinant Escherichia coli bacteriophage carrying a pneumococcal insert. Segments of DNA replaced chromosomal sequences adjacent to the region homologous with the pneumococcal insert, whence the name insertion-deletion. Here we report that a pneumococcal insert was absolutely required for insertion-deletion formation, but could be as short as 153 bp; that the sizes of foreign DNA insertions (289 -2,474 bp) and concomitant chromosomal deletions (45-1,485 bp) were not obviously correlated; that novel joints clustered preferentially within segments of high GC content; and that the crossovers in 29 independent novel joints were located 1 bp from the border or within short (3-10 nt long) stretches of identity (microhomology) between resident and foreign DNA. The data are consistent with a model in which the insert serving as a homologous recombination anchor favors interaction and subsequent illegitimate recombination events at microhomologies between foreign and resident sequences. The potential of homologydirected illegitimate recombination for genome evolution was illustrated by the trapping of functional heterologous genes. G enetic transformation, which was discovered in the Grampositive Streptococcus pneumoniae (pneumococcus) (1), is believed to play a central role in the biology of this human pathogen through its contribution to genetic plasticity (see ref.2 for a review). Transformation with naked DNA allows intraspecies and interspecies gene transfer. As such exchanges involve homologous recombination, they are generally ''conservative,'' i.e., they do not result in the creation of novel sequences but simply in a redistribution of previously existing genes. However, a pre-existing gene also can be modified when the two interacting sequences are partly divergent. Homologous recombination then leads to the production of mosaic genes, as exemplified in the case of the pbp genes of S. pneumoniae that encode altered penicillin-binding proteins with decreased affinity for ␤-lactam antibiotics (see ref. 3 for a review).Besides these conservative facets of transformation, is there any potential for the creation of novel sequence combinations or genes? The observation that transformation with chimeric donor DNA is mutagenic (4) suggested that shuffling and reassembly of previously unrelated sequences could readily occur in S. pneumoniae. The chimeric DNA extracted from a recombinant Escherichia coli bacteriophage carrying a pneumococcal insert produced illegitimate recombinants at a frequency of about 0.5% that of homologous recombinants (4). Illegitimate recombinants resulted from simultaneous insertion of heterologous vector (i.e., ) sequences and deletion of chromosomal sequences adjacent to the region homologous to the insert. These illegitimate events were therefore termed insertion-deletions (InsDels). As the pneumococcal insert in the chimeric donor seemed r...

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Control of Recombination Rate During Transformation ofStreptococcus pneumoniae: An Overview

Cited by 29 publications

References 46 publications

Effect of the Streptococcus pneumoniae MmsA Protein on the RecA Protein-promoted Three-strand Exchange Reaction

Effect of the Streptococcus pneumoniae MmsA Protein on the RecA Protein-promoted Three-strand Exchange Reaction

Frequency of Mutation to Rifampin Resistance in Streptococcus pneumoniae Clinical Strains: hexA and hexB Polymorphisms Do Not Account for Hypermutation

Homologous recombination at the border: Insertion-deletions and the trapping of foreign DNA in Streptococcus pneumoniae

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