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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1990 Apr;87(7):2790–2794. doi: 10.1073/pnas.87.7.2790

Evidence for the double-strand break repair model of bacteriophage lambda recombination.

N Takahashi 1, I Kobayashi 1
PMCID: PMC53776  PMID: 2138786

Abstract

We have obtained evidence for the repair of double-strand gaps promoted by the Red function of bacteriophage lambda. A double-strand gap was made in one of the two regions of homology in an inverted orientation on a plasmid DNA molecule. The gapped plasmid was introduced into Escherichia coli cells expressing the red alpha (exo) and red beta (bet) genes of lambda. The gap was repaired by DNA synthesis copying an intact duplex. This gap repair was sometimes accompanied by reciprocal recombination (crossing over). The gap stimulated recombination about 100-fold. Our results are compatible with previous proposals that lambda homologous recombination involves the following early steps: (i) generation of double-stranded ends by the packaging machinery or by the replication machinery; (ii) production of a single-stranded tail with a 3'-hydroxyl end by 5'----3' degradation by lambda exonuclease (red alpha gene product); (iii) pairing of the single-stranded tail with a complementary strand from a homologous duplex with the help of beta protein (red beta gene product); (iv) priming of DNA synthesis at this 3'-hydroxyl end to copy the second DNA molecule.

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Selected References

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  1. Carter D. M., Radding C. M. The role of exonuclease and beta protein of phage lambda in genetic recombination. II. Substrate specificity and the mode of action of lambda exonuclease. J Biol Chem. 1971 Apr 25;246(8):2502–2512. [PubMed] [Google Scholar]
  2. Dower W. J., Miller J. F., Ragsdale C. W. High efficiency transformation of E. coli by high voltage electroporation. Nucleic Acids Res. 1988 Jul 11;16(13):6127–6145. doi: 10.1093/nar/16.13.6127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Duckett D. R., Murchie A. I., Diekmann S., von Kitzing E., Kemper B., Lilley D. M. The structure of the Holliday junction, and its resolution. Cell. 1988 Oct 7;55(1):79–89. doi: 10.1016/0092-8674(88)90011-6. [DOI] [PubMed] [Google Scholar]
  4. Feiss M., Kobayashi I., Widner W. Separate sites for binding and nicking of bacteriophage lambda DNA by terminase. Proc Natl Acad Sci U S A. 1983 Feb;80(4):955–959. doi: 10.1073/pnas.80.4.955. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Greer H. The kil gene of bacteriophage lambda. Virology. 1975 Aug;66(2):589–604. doi: 10.1016/0042-6822(75)90231-7. [DOI] [PubMed] [Google Scholar]
  6. Jayaram M. Association of reciprocal exchange with gene conversion between the repeated segments of 2-micron circle. J Mol Biol. 1986 Oct 5;191(3):341–354. doi: 10.1016/0022-2836(86)90131-2. [DOI] [PubMed] [Google Scholar]
  7. Jayaram M. Mating type-like conversion promoted by the 2 micrograms circle site-specific recombinase: implications for the double-strand-gap repair model. Mol Cell Biol. 1986 Nov;6(11):3831–3837. doi: 10.1128/mcb.6.11.3831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Karu A. E., MacKay V., Goldmark P. J., Linn S. The recBC deoxyribonuclease of Escherichia coli K-12. Substrate specificity and reaction intermediates. J Biol Chem. 1973 Jul 25;248(14):4874–4884. [PubMed] [Google Scholar]
  9. Kmiec E., Holloman W. K. Beta protein of bacteriophage lambda promotes renaturation of DNA. J Biol Chem. 1981 Dec 25;256(24):12636–12639. [PubMed] [Google Scholar]
  10. Kobayashi I., Ikeda H. Double Holliday structure: a possible in vivo intermediate form of general recombination in Escherichia coli. Mol Gen Genet. 1983;191(2):213–220. doi: 10.1007/BF00334816. [DOI] [PubMed] [Google Scholar]
  11. Kobayashi I., Murialdo H., Crasemann J. M., Stahl M. M., Stahl F. W. Orientation of cohesive end site cos determines the active orientation of chi sequence in stimulating recA . recBC-mediated recombination in phage lambda lytic infections. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5981–5985. doi: 10.1073/pnas.79.19.5981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kobayashi I., Stahl M. M., Fairfield F. R., Stahl F. W. Coupling with packaging explains apparent nonreciprocality of Chi-stimulated recombination of bacteriophage lambda by RecA and RecBC functions. Genetics. 1984 Dec;108(4):773–794. doi: 10.1093/genetics/108.4.773. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kobayashi I., Stahl M. M., Leach D., Stahl F. W. The interaction of cos with Chi is separable from DNA packaging in recA-recBC-mediated recombination of bacteriophage lambda. Genetics. 1983 Aug;104(4):549–570. doi: 10.1093/genetics/104.4.549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Kobayashi I., Stahl M. M., Stahl F. W. The mechanism of the chi-cos interaction in RecA-RecBC-mediated recombination in phage lambda. Cold Spring Harb Symp Quant Biol. 1984;49:497–506. doi: 10.1101/sqb.1984.049.01.056. [DOI] [PubMed] [Google Scholar]
  15. Kobayashi I., Takahashi N. Double-stranded gap repair of DNA by gene conversion in Escherichia coli. Genetics. 1988 Aug;119(4):751–757. doi: 10.1093/genetics/119.4.751. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kolodkin A. L., Klar A. J., Stahl F. W. Double-strand breaks can initiate meiotic recombination in S. cerevisiae. Cell. 1986 Aug 29;46(5):733–740. doi: 10.1016/0092-8674(86)90349-1. [DOI] [PubMed] [Google Scholar]
  17. Little J. W. An exonuclease induced by bacteriophage lambda. II. Nature of the enzymatic reaction. J Biol Chem. 1967 Feb 25;242(4):679–686. [PubMed] [Google Scholar]
  18. MESELSON M. ON THE MECHANISM OF GENETIC RECOMBINATION BETWEEN DNA MOLECULES. J Mol Biol. 1964 Sep;9:734–745. doi: 10.1016/s0022-2836(64)80178-9. [DOI] [PubMed] [Google Scholar]
  19. Muniyappa K., Radding C. M. The homologous recombination system of phage lambda. Pairing activities of beta protein. J Biol Chem. 1986 Jun 5;261(16):7472–7478. [PubMed] [Google Scholar]
  20. Nickoloff J. A., Chen E. Y., Heffron F. A 24-base-pair DNA sequence from the MAT locus stimulates intergenic recombination in yeast. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7831–7835. doi: 10.1073/pnas.83.20.7831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Yeast transformation: a model system for the study of recombination. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6354–6358. doi: 10.1073/pnas.78.10.6354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Orr-Weaver T. L., Szostak J. W. Yeast recombination: the association between double-strand gap repair and crossing-over. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4417–4421. doi: 10.1073/pnas.80.14.4417. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Poteete A. R., Fenton A. C. Lambda red-dependent growth and recombination of phage P22. Virology. 1984 Apr 15;134(1):161–167. doi: 10.1016/0042-6822(84)90281-2. [DOI] [PubMed] [Google Scholar]
  24. Poteete A. R., Fenton A. C., Murphy K. C. Modulation of Escherichia coli RecBCD activity by the bacteriophage lambda Gam and P22 Abc functions. J Bacteriol. 1988 May;170(5):2012–2021. doi: 10.1128/jb.170.5.2012-2021.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Radding C. M., Shreffler D. C. Regulation of lambda exonuclease. II. Joint regulation of exonuclease and a new lambda antigen. J Mol Biol. 1966 Jul;18(2):251–261. doi: 10.1016/s0022-2836(66)80244-9. [DOI] [PubMed] [Google Scholar]
  26. Resnick M. A. The repair of double-strand breaks in DNA; a model involving recombination. J Theor Biol. 1976 Jun;59(1):97–106. doi: 10.1016/s0022-5193(76)80025-2. [DOI] [PubMed] [Google Scholar]
  27. Russo V. E. On the physical structure of lambda recombinant DNA. Mol Gen Genet. 1973 May 28;122(4):353–366. doi: 10.1007/BF00269436. [DOI] [PubMed] [Google Scholar]
  28. Sarthy P. V., Meselson M. Single burst study of rec- and red-mediated recombination in bacteriophage lambda. Proc Natl Acad Sci U S A. 1976 Dec;73(12):4613–4617. doi: 10.1073/pnas.73.12.4613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Simmon V. F., Lederberg S. Degradation of bacteriophage lambda deoxyribonucleic acid after restriction by Escherichia coli K-12. J Bacteriol. 1972 Oct;112(1):161–169. doi: 10.1128/jb.112.1.161-169.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Smith G. R. Homologous recombination in procaryotes. Microbiol Rev. 1988 Mar;52(1):1–28. doi: 10.1128/mr.52.1.1-28.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Sriprakash K. S., Lundh N., Huh MM-O, Radding C. M. The specificity of lambda exonuclease. Interactions with single-stranded DNA. J Biol Chem. 1975 Jul 25;250(14):5438–5445. [PubMed] [Google Scholar]
  32. Stahl F. W., Kobayashi I., Stahl M. M. Distance from cohesive end site cos determines the replication requirement for recombination in phage lambda. Proc Natl Acad Sci U S A. 1982 Oct;79(20):6318–6321. doi: 10.1073/pnas.79.20.6318. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Stahl F. W., Kobayashi I., Stahl M. M. In phage lambda, cos is a recombinator in the red pathway. J Mol Biol. 1985 Jan 20;181(2):199–209. doi: 10.1016/0022-2836(85)90085-3. [DOI] [PubMed] [Google Scholar]
  34. Stahl F. W., McMilin K. D., Stahl M. M., Crasemann J. M., Lam S. The distribution of crossovers along unreplicated lambda bacteriophage chromosomes. Genetics. 1974 Jul;77(3):395–408. doi: 10.1093/genetics/77.3.395. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Stahl F. W., McMilin K. D., Stahl M. M., Nozu Y. An enhancing role for DNA synthesis in formation of bacteriophage lambda recombinants. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3598–3601. doi: 10.1073/pnas.69.12.3598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Stahl F. W. Roles of double-strand breaks in generalized genetic recombination. Prog Nucleic Acid Res Mol Biol. 1986;33:169–194. doi: 10.1016/s0079-6603(08)60023-9. [DOI] [PubMed] [Google Scholar]
  37. Stahl F. W., Stahl M. M. DNA synthesis at the site of a Red-mediated exchange in phage lambda. Genetics. 1986 May;113(1):1–12. doi: 10.1093/genetics/113.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Stahl M. M., Kobayashi I., Stahl F. W., Huntington S. K. Activation of Chi, a recombinator, by the action of an endonuclease at a distant site. Proc Natl Acad Sci U S A. 1983 Apr;80(8):2310–2313. doi: 10.1073/pnas.80.8.2310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Strathern J. N., Klar A. J., Hicks J. B., Abraham J. A., Ivy J. M., Nasmyth K. A., McGill C. Homothallic switching of yeast mating type cassettes is initiated by a double-stranded cut in the MAT locus. Cell. 1982 Nov;31(1):183–192. doi: 10.1016/0092-8674(82)90418-4. [DOI] [PubMed] [Google Scholar]
  40. Szostak J. W., Orr-Weaver T. L., Rothstein R. J., Stahl F. W. The double-strand-break repair model for recombination. Cell. 1983 May;33(1):25–35. doi: 10.1016/0092-8674(83)90331-8. [DOI] [PubMed] [Google Scholar]
  41. Takahashi S. Structure of the oligomeric circular molecules of a bacteriophage lambda DNA. Virology. 1975 Apr;64(2):319–329. doi: 10.1016/0042-6822(75)90108-7. [DOI] [PubMed] [Google Scholar]
  42. Taylor A. F., Schultz D. W., Ponticelli A. S., Smith G. R. RecBC enzyme nicking at Chi sites during DNA unwinding: location and orientation-dependence of the cutting. Cell. 1985 May;41(1):153–163. doi: 10.1016/0092-8674(85)90070-4. [DOI] [PubMed] [Google Scholar]
  43. Thaler D. S., Sampson E., Siddiqi I., Rosenberg S. M., Thomason L. C., Stahl F. W., Stahl M. M. Recombination of bacteriophage lambda in recD mutants of Escherichia coli. Genome. 1989;31(1):53–67. doi: 10.1139/g89-013. [DOI] [PubMed] [Google Scholar]
  44. Thaler D. S., Stahl F. W. DNA double-chain breaks in recombination of phage lambda and of yeast. Annu Rev Genet. 1988;22:169–197. doi: 10.1146/annurev.ge.22.120188.001125. [DOI] [PubMed] [Google Scholar]
  45. Thaler D. S., Stahl M. M., Stahl F. W. Double-chain-cut sites are recombination hotspots in the Red pathway of phage lambda. J Mol Biol. 1987 May 5;195(1):75–87. doi: 10.1016/0022-2836(87)90328-7. [DOI] [PubMed] [Google Scholar]
  46. Thaler D. S., Stahl M. M., Stahl F. W. Tests of the double-strand-break repair model for red-mediated recombination of phage lambda and plasmid lambda dv. Genetics. 1987 Aug;116(4):501–511. doi: 10.1093/genetics/116.4.501. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Weil J. Reciprocal and non-reciprocal recombination in bacteriopahge lambda. J Mol Biol. 1969 Jul 28;43(2):351–355. doi: 10.1016/0022-2836(69)90276-9. [DOI] [PubMed] [Google Scholar]
  48. White R. L., Fox M. S. On the molecular basis of high negative interference. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1544–1548. doi: 10.1073/pnas.71.4.1544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Yamamoto K., Takahashi N., Yoshikura H., Kobayashi I. Homologous recombination involving a large heterology in Escherichia coli. Genetics. 1988 Aug;119(4):759–769. doi: 10.1093/genetics/119.4.759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Yamamoto K., Yoshikura H., Takahashi N., Kobayashi I. Apparent gene conversion in an Escherichia coli rec+ strain is explained by multiple rounds of reciprocal crossing-over. Mol Gen Genet. 1988 Jun;212(3):393–404. doi: 10.1007/BF00330842. [DOI] [PubMed] [Google Scholar]
  51. Zinn A. R., Butow R. A. Nonreciprocal exchange between alleles of the yeast mitochondrial 21S rRNA gene: kinetics and the involvement of a double-strand break. Cell. 1985 Apr;40(4):887–895. doi: 10.1016/0092-8674(85)90348-4. [DOI] [PubMed] [Google Scholar]

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