Skip to main content
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
. 1988 Sep;85(17):6323–6327. doi: 10.1073/pnas.85.17.6323

Helical-repeat dependence of integrative recombination of bacteriophage lambda: role of the P1 and H1 protein binding sites.

J F Thompson 1, U K Snyder 1, A Landy 1
PMCID: PMC281962  PMID: 2842765

Abstract

The efficiency of site-specific recombination of bacteriophage lambda was found to depend on the spacing between distant protein binding sites. Insertions and deletions of up to 30 base pairs were made in the nonessential regions between the H1 and H2 protein binding sites. Recombination was found to occur in substrates with changes of integral multiples of a DNA helical repeat, whereas recombination was defective in substrates with nonintegral changes. The lambda recombinogenic complex is especially interesting because two different proteins are involved: integration host factor (IHF), which has been shown to bend DNA, and the phage-encoded integrase protein (Int), which has been shown to have two distinct DNA-binding domains. The importance of angular displacement of protein binding sites was confirmed by addition of ethidium bromide to defective substrates. Significant stimulation of recombination was observed when sufficient drug intercalated and unwound the DNA to allow improved orientation of sites. The orientation effects are dependent on supercoiling, as spacing is less important in conditions where supercoiling and the P1-H1 sites are not required for recombination.

Full text

PDF
6325

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Abremski K., Gottesman S. Purification of the bacteriophage lambda xis gene product required for lambda excisive recombination. J Biol Chem. 1982 Aug 25;257(16):9658–9662. [PubMed] [Google Scholar]
  2. Bauer C. E., Hesse S. D., Gumport R. I., Gardner J. F. Mutational analysis of integrase arm-type binding sites of bacteriophage lambda. Integration and excision involve distinct interactions of integrase with arm-type sites. J Mol Biol. 1986 Dec 5;192(3):513–527. doi: 10.1016/0022-2836(86)90273-1. [DOI] [PubMed] [Google Scholar]
  3. Better M., Lu C., Williams R. C., Echols H. Site-specific DNA condensation and pairing mediated by the int protein of bacteriophage lambda. Proc Natl Acad Sci U S A. 1982 Oct;79(19):5837–5841. doi: 10.1073/pnas.79.19.5837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Better M., Wickner S., Auerbach J., Echols H. Role of the Xis protein of bacteriophage lambda in a specific reactive complex at the attR prophage attachment site. Cell. 1983 Jan;32(1):161–168. doi: 10.1016/0092-8674(83)90506-8. [DOI] [PubMed] [Google Scholar]
  5. Borowiec J. A., Zhang L., Sasse-Dwight S., Gralla J. D. DNA supercoiling promotes formation of a bent repression loop in lac DNA. J Mol Biol. 1987 Jul 5;196(1):101–111. doi: 10.1016/0022-2836(87)90513-4. [DOI] [PubMed] [Google Scholar]
  6. Bushman W., Thompson J. F., Vargas L., Landy A. Control of directionality in lambda site specific recombination. Science. 1985 Nov 22;230(4728):906–911. doi: 10.1126/science.2932798. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Craig N. L., Nash H. A. E. coli integration host factor binds to specific sites in DNA. Cell. 1984 Dec;39(3 Pt 2):707–716. doi: 10.1016/0092-8674(84)90478-1. [DOI] [PubMed] [Google Scholar]
  8. Craig N. L., Nash H. A. The mechanism of phage lambda site-specific recombination: site-specific breakage of DNA by Int topoisomerase. Cell. 1983 Dec;35(3 Pt 2):795–803. doi: 10.1016/0092-8674(83)90112-5. [DOI] [PubMed] [Google Scholar]
  9. Dunn T. M., Hahn S., Ogden S., Schleif R. F. An operator at -280 base pairs that is required for repression of araBAD operon promoter: addition of DNA helical turns between the operator and promoter cyclically hinders repression. Proc Natl Acad Sci U S A. 1984 Aug;81(16):5017–5020. doi: 10.1073/pnas.81.16.5017. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Dynan W. S., Tjian R. Control of eukaryotic messenger RNA synthesis by sequence-specific DNA-binding proteins. 1985 Aug 29-Sep 4Nature. 316(6031):774–778. doi: 10.1038/316774a0. [DOI] [PubMed] [Google Scholar]
  11. Echols H. Multiple DNA-protein interactions governing high-precision DNA transactions. Science. 1986 Sep 5;233(4768):1050–1056. doi: 10.1126/science.2943018. [DOI] [PubMed] [Google Scholar]
  12. Griffith J. D., Nash H. A. Genetic rearrangement of DNA induces knots with a unique topology: implications for the mechanism of synapsis and crossing-over. Proc Natl Acad Sci U S A. 1985 May;82(10):3124–3128. doi: 10.1073/pnas.82.10.3124. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Griffith J., Hochschild A., Ptashne M. DNA loops induced by cooperative binding of lambda repressor. Nature. 1986 Aug 21;322(6081):750–752. doi: 10.1038/322750a0. [DOI] [PubMed] [Google Scholar]
  14. Hahn S., Hendrickson W., Schleif R. Transcription of Escherichia coli ara in vitro. The cyclic AMP receptor protein requirement for PBAD induction that depends on the presence and orientation of the araO2 site. J Mol Biol. 1986 Apr 5;188(3):355–367. doi: 10.1016/0022-2836(86)90160-9. [DOI] [PubMed] [Google Scholar]
  15. Herskowitz I., Hagen D. The lysis-lysogeny decision of phage lambda: explicit programming and responsiveness. Annu Rev Genet. 1980;14:399–445. doi: 10.1146/annurev.ge.14.120180.002151. [DOI] [PubMed] [Google Scholar]
  16. Hochschild A., Ptashne M. Cooperative binding of lambda repressors to sites separated by integral turns of the DNA helix. Cell. 1986 Mar 14;44(5):681–687. doi: 10.1016/0092-8674(86)90833-0. [DOI] [PubMed] [Google Scholar]
  17. Hsu P. L., Landy A. Resolution of synthetic att-site Holliday structures by the integrase protein of bacteriophage lambda. Nature. 1984 Oct 25;311(5988):721–726. doi: 10.1038/311721a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hsu P. L., Ross W., Landy A. The lambda phage att site: functional limits and interaction with Int protein. Nature. 1980 May 8;285(5760):85–91. doi: 10.1038/285085a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Johnson R. C., Glasgow A. C., Simon M. I. Spatial relationship of the Fis binding sites for Hin recombinational enhancer activity. Nature. 1987 Oct 1;329(6138):462–465. doi: 10.1038/329462a0. [DOI] [PubMed] [Google Scholar]
  20. Kikuchi Y., Nash H. A. Nicking-closing activity associated with bacteriophage lambda int gene product. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3760–3764. doi: 10.1073/pnas.76.8.3760. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Krämer H., Niemöller M., Amouyal M., Revet B., von Wilcken-Bergmann B., Müller-Hill B. lac repressor forms loops with linear DNA carrying two suitably spaced lac operators. EMBO J. 1987 May;6(5):1481–1491. doi: 10.1002/j.1460-2075.1987.tb02390.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Landy A., Ross W. Viral integration and excision: structure of the lambda att sites. Science. 1977 Sep 16;197(4309):1147–1160. doi: 10.1126/science.331474. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Mizuuchi K., Gellert M., Nash H. A. Involement of supertwisted DNA in integrative recombination of bacteriophage lambda. J Mol Biol. 1978 May 25;121(3):375–392. doi: 10.1016/0022-2836(78)90370-4. [DOI] [PubMed] [Google Scholar]
  24. Mossing M. C., Record M. T., Jr Upstream operators enhance repression of the lac promoter. Science. 1986 Aug 22;233(4766):889–892. doi: 10.1126/science.3090685. [DOI] [PubMed] [Google Scholar]
  25. Nash H. A. Integration and excision of bacteriophage lambda: the mechanism of conservation site specific recombination. Annu Rev Genet. 1981;15:143–167. doi: 10.1146/annurev.ge.15.120181.001043. [DOI] [PubMed] [Google Scholar]
  26. Nash H. A. Integrative recombination of bacteriophage lambda DNA in vitro. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1072–1076. doi: 10.1073/pnas.72.3.1072. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Nash H. A., Robertson C. A. Purification and properties of the Escherichia coli protein factor required for lambda integrative recombination. J Biol Chem. 1981 Sep 10;256(17):9246–9253. [PubMed] [Google Scholar]
  28. Pollock T. J., Abremski K. DNA without supertwists can be an in vitro substrate for site-specific recombination of bacteriophage lambda. J Mol Biol. 1979 Jul 5;131(3):651–654. doi: 10.1016/0022-2836(79)90013-5. [DOI] [PubMed] [Google Scholar]
  29. Prentki P., Chandler M., Galas D. J. Escherichia coli integration host factor bends the DNA at the ends of IS1 and in an insertion hotspot with multiple IHF binding sites. EMBO J. 1987 Aug;6(8):2479–2487. doi: 10.1002/j.1460-2075.1987.tb02529.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Ptashne M. Gene regulation by proteins acting nearby and at a distance. Nature. 1986 Aug 21;322(6081):697–701. doi: 10.1038/322697a0. [DOI] [PubMed] [Google Scholar]
  31. Richet E., Abcarian P., Nash H. A. The interaction of recombination proteins with supercoiled DNA: defining the role of supercoiling in lambda integrative recombination. Cell. 1986 Sep 26;46(7):1011–1021. doi: 10.1016/0092-8674(86)90700-2. [DOI] [PubMed] [Google Scholar]
  32. Robertson C. A., Nash H. A. Bending of the bacteriophage lambda attachment site by Escherichia coli integration host factor. J Biol Chem. 1988 Mar 15;263(8):3554–3557. [PubMed] [Google Scholar]
  33. Ross W., Landy A. Bacteriophage lambda int protein recognizes two classes of sequence in the phage att site: characterization of arm-type sites. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7724–7728. doi: 10.1073/pnas.79.24.7724. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Ross W., Landy A. Patterns of lambda Int recognition in the regions of strand exchange. Cell. 1983 May;33(1):261–272. doi: 10.1016/0092-8674(83)90355-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Shore D., Langowski J., Baldwin R. L. DNA flexibility studied by covalent closure of short fragments into circles. Proc Natl Acad Sci U S A. 1981 Aug;78(8):4833–4837. doi: 10.1073/pnas.78.8.4833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Singleton C. K., Wells R. D. The facile generation of covalently closed, circular DNAs with defined negative superhelical densities. Anal Biochem. 1982 May 15;122(2):253–257. doi: 10.1016/0003-2697(82)90277-9. [DOI] [PubMed] [Google Scholar]
  37. Spengler S. J., Stasiak A., Cozzarelli N. R. The stereostructure of knots and catenanes produced by phage lambda integrative recombination: implications for mechanism and DNA structure. Cell. 1985 Aug;42(1):325–334. doi: 10.1016/s0092-8674(85)80128-8. [DOI] [PubMed] [Google Scholar]
  38. Stenzel T. T., Patel P., Bastia D. The integration host factor of Escherichia coli binds to bent DNA at the origin of replication of the plasmid pSC101. Cell. 1987 Jun 5;49(5):709–717. doi: 10.1016/0092-8674(87)90547-2. [DOI] [PubMed] [Google Scholar]
  39. Takahashi K., Vigneron M., Matthes H., Wildeman A., Zenke M., Chambon P. Requirement of stereospecific alignments for initiation from the simian virus 40 early promoter. Nature. 1986 Jan 9;319(6049):121–126. doi: 10.1038/319121a0. [DOI] [PubMed] [Google Scholar]
  40. Thompson J. F., Moitoso de Vargas L., Koch C., Kahmann R., Landy A. Cellular factors couple recombination with growth phase: characterization of a new component in the lambda site-specific recombination pathway. Cell. 1987 Sep 11;50(6):901–908. doi: 10.1016/0092-8674(87)90516-2. [DOI] [PubMed] [Google Scholar]
  41. Thompson J. F., Waechter-Brulla D., Gumport R. I., Gardner J. F., Moitoso de Vargas L., Landy A. Mutations in an integration host factor-binding site: effect on lambda site-specific recombination and regulatory implications. J Bacteriol. 1986 Dec;168(3):1343–1351. doi: 10.1128/jb.168.3.1343-1351.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Thompson J. F., de Vargas L. M., Skinner S. E., Landy A. Protein-protein interactions in a higher-order structure direct lambda site-specific recombination. J Mol Biol. 1987 Jun 5;195(3):481–493. doi: 10.1016/0022-2836(87)90177-x. [DOI] [PubMed] [Google Scholar]
  43. Wang J. C., Giaever G. N. Action at a distance along a DNA. Science. 1988 Apr 15;240(4850):300–304. doi: 10.1126/science.3281259. [DOI] [PubMed] [Google Scholar]
  44. Wang J. C. The degree of unwinding of the DNA helix by ethidium. I. Titration of twisted PM2 DNA molecules in alkaline cesium chloride density gradients. J Mol Biol. 1974 Nov 15;89(4):783–801. doi: 10.1016/0022-2836(74)90053-9. [DOI] [PubMed] [Google Scholar]
  45. Yin S., Bushman W., Landy A. Interaction of the lambda site-specific recombination protein Xis with attachment site DNA. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1040–1044. doi: 10.1073/pnas.82.4.1040. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES