1887

Abstract

During natural transformation of , the genomic integration of foreign (non-homologous) DNA is possible when the DNA contains a single segment homologous to the recipient genome (anchor) through homologous recombination in the anchor facilitating illegitimate recombination in the neighbouring foreign DNA (homology-facilitated illegitimate recombination; HFIR). DNA integration by HFIR occurs about 10 000 times less frequently than fully homologous recombination, but at least 100 000-fold more frequently than integration in the absence of any homology. We investigated the influence of the RecBCD enzyme (DNase/helicase) and SbcCD DNase (DNA-structure-specific single-strand endonuclease and exonuclease) on HFIR. In a null mutant the acquisition of foreign DNA was elevated 11-fold relative to wild-type cells by a 6.9-fold increased HFIR frequency and by the integration of longer stretches of foreign DNA in each event. In an null mutant, the foreign DNA acquisition was 4.5-fold higher than in the wild-type, while homologous transformation with large DNA molecules was unaffected and increased 3.2-fold with small DNA fragments. The mutation partially suppressed the high UV sensitivity and low viability of the mutant and also decreased its foreign DNA acquisition by HFIR to the lower level of the mutant. We propose that suppression of HFIR results from the elimination of double-stranded intermediates of the HFIR process during transformation by RecBCD, and by SbcCD interfering with branched molecules. Our results provide evidence that the homologous recombination enzymes RecBCD and SbcCD control the level of foreign DNA acquisition by HFIR.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.2008/018382-0
2008-08-01
2019-10-17
Loading full text...

Full text loading...

/deliver/fulltext/micro/154/8/2437.html?itemId=/content/journal/micro/10.1099/mic.0.2008/018382-0&mimeType=html&fmt=ahah

References

  1. Amundsen, S. K., Taylor, A. F., Chaudhury, A. M. & Smith, G. R. ( 1986; ). recD: the gene for an essential third subunit of exonuclease V. Proc Natl Acad Sci U S A 83, 5558–5562.[CrossRef]
    [Google Scholar]
  2. Anderson, D. G. & Kowalczykowski, S. C. ( 1997; ). The translocating RecBCD enzyme stimulates recombination by directing RecA protein onto ssDNA in a chi-regulated manner. Cell 90, 77–86.[CrossRef]
    [Google Scholar]
  3. Arber, W. ( 2000; ). Genetic variation: molecular mechanisms and impact on microbial evolution. FEMS Microbiol Rev 24, 1–7.[CrossRef]
    [Google Scholar]
  4. Barbe, V., Vallenet, D., Fonknechten, N., Kreimeyer, A., Oztas, S., Labarre, L., Cruveiller, S., Robert, C., Duprat, S. & other authors ( 2004; ). Unique features revealed by the genome sequence of Acinetobacter sp. ADP1, a versatile and naturally transformation competent bacterium. Nucleic Acids Res 32, 5766–5779.[CrossRef]
    [Google Scholar]
  5. Bentchikou, E., Servant, P., Coste, G. & Sommer, S. ( 2007; ). Additive effects of SbcCD and PolX deficiencies in the in vivo repair of DNA double-strand breaks in Deinococcus radiodurans. J Bacteriol 189, 4784–4790.[CrossRef]
    [Google Scholar]
  6. Bierne, H., Ehrlich, S. D. & Michel, B. ( 1997; ). Deletions at stalled replication forks occur by two different pathways. EMBO J 16, 3332–3340.[CrossRef]
    [Google Scholar]
  7. Burdett, V., Baitinger, C., Viswanathan, M., Lovett, S. T. & Modrich, P. ( 2001; ). In vivo requirement for RecJ, ExoVII, ExoI, and ExoX in methyl-directed mismatch repair. Proc Natl Acad Sci U S A 98, 6765–6770.[CrossRef]
    [Google Scholar]
  8. Chen, I. & Dubnau, D. ( 2004; ). DNA uptake during bacterial transformation. Nat Rev Microbiol 2, 241–249.[CrossRef]
    [Google Scholar]
  9. Churchill, J. J., Anderson, D. G. & Kowalczykowski, S. C. ( 1999; ). The RecBC enzyme loads RecA protein onto ssDNA asymmetrically and independently of chi, resulting in constitutive recombination activation. Genes Dev 13, 901–911.[CrossRef]
    [Google Scholar]
  10. Claverys, J. P., Lefevre, J. C. & Sicard, A. M. ( 1980; ). Transformation of Streptococcus pneumoniae with S. pneumoniae-lambda phage hybrid DNA: induction of deletions. Proc Natl Acad Sci U S A 77, 3534–3538.[CrossRef]
    [Google Scholar]
  11. Connelly, J. C. & Leach, D. R. ( 1996; ). The sbcC and sbcD genes of Escherichia coli encode a nuclease involved in palindrome inviability and genetic recombination. Genes Cells 1, 285–291.[CrossRef]
    [Google Scholar]
  12. Connelly, J. C., de Leau, E. S., Okely, E. A. & Leach, D. R. ( 1997; ). Overexpression, purification, and characterization of the SbcCD protein from Escherichia coli. J Biol Chem 272, 19819–19826.[CrossRef]
    [Google Scholar]
  13. Connelly, J. C., Kirkham, L. A. & Leach, D. R. ( 1998; ). The SbcCD nuclease of Escherichia coli is a structural maintenance of chromosomes (SMC) family protein that cleaves hairpin DNA. Proc Natl Acad Sci U S A 95, 7969–7974.[CrossRef]
    [Google Scholar]
  14. Connelly, J. C., de Leau, E. S. & Leach, D. R. ( 1999; ). DNA cleavage and degradation by the SbcCD protein complex from Escherichia coli. Nucleic Acids Res 27, 1039–1046.[CrossRef]
    [Google Scholar]
  15. de Vries, J. & Wackernagel, W. ( 1998; ). Detection of nptII (kanamycin resistance) genes in genomes of transgenic plants by marker rescue transformation. Mol Gen Genet 257, 606–613.[CrossRef]
    [Google Scholar]
  16. de Vries, J. & Wackernagel, W. ( 2002; ). Integration of foreign DNA during natural transformation of Acinetobacter sp. by homology-facilitated illegitimate recombination. Proc Natl Acad Sci U S A 99, 2094–2099.[CrossRef]
    [Google Scholar]
  17. de Vries, J., Heine, M., Harms, K. & Wackernagel, W. ( 2003; ). Spread of recombinant DNA by roots and pollen of transgenic potato plants, identified by highly specific biomonitoring using natural transformation of Acinetobacter sp. Appl Environ Microbiol 69, 4455–4462.[CrossRef]
    [Google Scholar]
  18. de Vries, J., Herzfeld, T. & Wackernagel, W. ( 2004; ). Transfer of plastid DNA from tobacco to the soil bacterium Acinetobacter sp. by natural transformation. Mol Microbiol 53, 323–334.[CrossRef]
    [Google Scholar]
  19. Dubnau, D. & Provvedi, R. ( 2000; ). Internalizing DNA. Res Microbiol 151, 475–480.[CrossRef]
    [Google Scholar]
  20. Harms, K., Schön, V., Kickstein, E. & Wackernagel, W. ( 2007; ). The RecJ DNase strongly suppresses genomic integration of short but not long DNA fragments by homology-facilitated illegitimate recombination during transformation of Acinetobacter baylyi. Mol Microbiol 64, 691–702.[CrossRef]
    [Google Scholar]
  21. Hülter, N. & Wackernagel, W. ( 2008; ). Double illegitimate recombination events integrate DNA segments through two different mechanisms during natural transformation of Acinetobacter baylyi. Mol Microbiol 67, 984–995.[CrossRef]
    [Google Scholar]
  22. Kickstein, E., Harms, K. & Wackernagel, W. ( 2007; ). Deletions of recBCD and recD influence genetic transformation differently and are lethal together with a recJ deletion in Acinetobacter baylyi. Microbiology 153, 2259–2270.[CrossRef]
    [Google Scholar]
  23. Kowalczykowski, S. C., Dixon, D. A., Eggleston, A. K., Lauder, S. D. & Rehrauer, W. M. ( 1994; ). Biochemistry of homologous recombination in Escherichia coli. Microbiol Rev 58, 401–465.
    [Google Scholar]
  24. Kushner, S. R., Nagaishi, H., Templin, A. & Clark, A. J. ( 1971; ). Genetic recombination in Escherichia coli: the role of exonuclease I. Proc Natl Acad Sci U S A 68, 824–827.[CrossRef]
    [Google Scholar]
  25. Kushner, S. R., Nagaishi, H. & Clark, A. J. ( 1972; ). Indirect suppression of recB and recC mutations by exonuclease I deficiency. Proc Natl Acad Sci U S A 69, 1366–1370.[CrossRef]
    [Google Scholar]
  26. Kuzminov, A. ( 1999; ). Recombinational repair of DNA damage in Escherichia coli and bacteriophage lambda. Microbiol Mol Biol Rev 63, 751–813.
    [Google Scholar]
  27. Lehman, I. R. & Nussbaum, A. L. ( 1964; ). The deoxyribonucleases of Escherichia coli. V. On the specificity of exonuclease I (phosphodiesterase). J Biol Chem 239, 2628–2636.
    [Google Scholar]
  28. Lloyd, R. G. & Buckman, C. ( 1985; ). Identification and genetic analysis of sbcC mutations in commonly used recBC sbcB strains of Escherichia coli K-12. J Bacteriol 164, 836–844.
    [Google Scholar]
  29. Lorenz, M. G. & Wackernagel, W. ( 1994; ). Bacterial gene transfer by natural genetic transformation in the environment. Microbiol Rev 58, 563–602.
    [Google Scholar]
  30. Mascarenhas, J., Sanchez, H., Tadesse, S., Kidane, D., Krisnamurthy, M., Alonso, J. C. & Graumann, P. L. ( 2006; ). Bacillus subtilis SbcC protein plays an important role in DNA inter-strand cross-link repair. BMC Mol Biol 7, 20 [CrossRef]
    [Google Scholar]
  31. Meier, P. & Wackernagel, W. ( 2003; ). Mechanisms of homology-facilitated illegitimate recombination for foreign DNA acquisition in transformable Pseudomonas stutzeri. Mol Microbiol 48, 1107–1118.[CrossRef]
    [Google Scholar]
  32. Meier, P. & Wackernagel, W. ( 2005; ). Impact of mutS inactivation on foreign DNA acquisition by natural transformation in Pseudomonas stutzeri. J Bacteriol 187, 143–154.[CrossRef]
    [Google Scholar]
  33. Prudhomme, M., Libante, V. & Claverys, J. P. ( 2002; ). Homologous recombination at the border: insertion-deletions and the trapping of foreign DNA in Streptococcus pneumoniae. Proc Natl Acad Sci U S A 99, 2100–2105.[CrossRef]
    [Google Scholar]
  34. Romanowski, G., Lorenz, M. G. & Wackernagel, W. ( 1993; ). Use of polymerase chain reaction and electroporation of Escherichia coli to monitor the persistence of extracellular plasmid DNA introduced into natural soils. Appl Environ Microbiol 59, 3438–3446.
    [Google Scholar]
  35. Sambrook, J., Fritsch, E. F. & Maniatis, T. ( 1989; ). Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory.
  36. Sharples, G. J. & Leach, D. R. ( 1995; ). Structural and functional similarities between the SbcCD proteins of Escherichia coli and the RAD50 and MRE11 (RAD32) recombination and repair proteins of yeast. Mol Microbiol 17, 1215–1217.[CrossRef]
    [Google Scholar]
  37. Shiraishi, K., Imai, Y., Yoshizaki, S. & Ikeda, H. ( 2005; ). Rep helicase suppresses short-homology-dependent illegitimate recombination in Escherichia coli. Genes Cells 10, 1015–1023.[CrossRef]
    [Google Scholar]
  38. Thoms, B. & Wackernagel, W. ( 1982; ). UV-induced alleviation of lambda restriction in Escherichia coli K-12: kinetics of induction and specificity of this SOS function. Mol Gen Genet 186, 111–117.[CrossRef]
    [Google Scholar]
  39. Thoms, B., Borchers, I. & Wackernagel, W. ( 2008; ). Effects of single-strand DNases ExoI, RecJ, ExoVII, and SbcCD on homologous recombination of recBCD + strains of Escherichia coli and roles of SbcB15 and XonA2 mutant enzymes of ExoI. J Bacteriol 190, 179–192.[CrossRef]
    [Google Scholar]
  40. Tønjum, T., Higgins, J. D. & Gibson, T. J. ( 1995; ). Fastidious Gram-negative bacteria: meeting the diagnostic challenge with nucleic acid analysis. APMIS 103, 609–627.[CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.2008/018382-0
Loading
/content/journal/micro/10.1099/mic.0.2008/018382-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error