Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1986 Dec;168(3):1343–1351. doi: 10.1128/jb.168.3.1343-1351.1986

Mutations in an integration host factor-binding site: effect on lambda site-specific recombination and regulatory implications.

J F Thompson, D Waechter-Brulla, R I Gumport, J F Gardner, L Moitoso de Vargas, A Landy
PMCID: PMC213644  PMID: 2946666

Abstract

The manner in which integration host factor (IHF) regulates lambda site-specific recombination has been analyzed by examining the behavior of both wild-type and mutant DNAs in integrative and excisive recombination as well as in protein binding. While integrative recombination of an attP with two base changes in the H1 site required 8-fold more IHF than did wild type, binding to this site was lowered at least 500-fold, suggestive of cooperative interactions. A mutant attP with nine base changes did not integrate at all in vitro, with the defect being less severe in vivo. IHF inhibition of excisive recombination was relieved by both mutations in vitro and in vivo. These results imply that occupancy of the H1 site is critical for determining the direction of recombination. It is proposed that IHF inhibition of excision provides a monitor of the strength of the induction stimulus and the nutritional state of the cell; this would allow the prophage to excise selectively in conditions which favor successful completion of the lytic cycle.

Full text

PDF
1343

Images in this article

1345Image
on p.1345
1346Image
on p.1346
1347Image
on p.1347

Selected References

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

  1. Bailone A., Levine A., Devoret R. Inactivation of prophage lambda repressor in vivo. J Mol Biol. 1979 Jul 5;131(3):553–572. doi: 10.1016/0022-2836(79)90007-x. [DOI] [PubMed] [Google Scholar]
  2. Bauer C. E., Gardner J. F., Gumport R. I. Extent of sequence homology required for bacteriophage lambda site-specific recombination. J Mol Biol. 1985 Jan 20;181(2):187–197. doi: 10.1016/0022-2836(85)90084-1. [DOI] [PubMed] [Google Scholar]
  3. Bauer C. E., Hesse S. D., Waechter-Brulla D. A., Lynn S. P., Gumport R. I., Gardner J. F. A genetic enrichment for mutations constructed by oligodeoxynucleotide-directed mutagenesis. Gene. 1985;37(1-3):73–81. doi: 10.1016/0378-1119(85)90259-8. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Bushman W., Yin S., Thio L. L., Landy A. Determinants of directionality in lambda site-specific recombination. Cell. 1984 Dec;39(3 Pt 2):699–706. doi: 10.1016/0092-8674(84)90477-x. [DOI] [PubMed] [Google Scholar]
  6. Court D., Green L., Echols H. Positive and negative regulation by the cII and cIII gene products of bacteriophage lambda. Virology. 1975 Feb;63(2):484–491. doi: 10.1016/0042-6822(75)90321-9. [DOI] [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. Enquist L. W., Kikuchi A., Weisberg R. A. The role of lambda integrase in integration and excision. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):1115–1120. doi: 10.1101/sqb.1979.043.01.124. [DOI] [PubMed] [Google Scholar]
  10. Fried M., Crothers D. M. Equilibria and kinetics of lac repressor-operator interactions by polyacrylamide gel electrophoresis. Nucleic Acids Res. 1981 Dec 11;9(23):6505–6525. doi: 10.1093/nar/9.23.6505. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Friedman D. I., Olson E. J., Carver D., Gellert M. Synergistic effect of himA and gyrB mutations: evidence that him functions control expression of ilv and xyl genes. J Bacteriol. 1984 Feb;157(2):484–489. doi: 10.1128/jb.157.2.484-489.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Galas D. J., Schmitz A. DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res. 1978 Sep;5(9):3157–3170. doi: 10.1093/nar/5.9.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Garner M. M., Revzin A. A gel electrophoresis method for quantifying the binding of proteins to specific DNA regions: application to components of the Escherichia coli lactose operon regulatory system. Nucleic Acids Res. 1981 Jul 10;9(13):3047–3060. doi: 10.1093/nar/9.13.3047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Gillam S., Smith M. Site-specific mutagenesis using synthetic oligodeoxyribonucleotide primers: I. Optimum conditions and minimum ologodeoxyribonucleotide length. Gene. 1979 Dec;8(1):81–97. doi: 10.1016/0378-1119(79)90009-x. [DOI] [PubMed] [Google Scholar]
  15. Gottesman M. E., Yarmolinsky M. B. Integration-negative mutants of bacteriophage lambda. J Mol Biol. 1968 Feb 14;31(3):487–505. doi: 10.1016/0022-2836(68)90423-3. [DOI] [PubMed] [Google Scholar]
  16. Hardies S. C., Patient R. K., Klein R. D., Ho F., Reznikoff W. S., Wells R. D. Construction and mapping of recombinant plasmids used for the preparation of DNA fragments containing the Escherichia coli lactose operator and promoter. J Biol Chem. 1979 Jun 25;254(12):5527–5534. [PubMed] [Google Scholar]
  17. Ho Y. S., Rosenberg M. Characterization of a third, cII-dependent, coordinately activated promoter on phage lambda involved in lysogenic development. J Biol Chem. 1985 Sep 25;260(21):11838–11844. [PubMed] [Google Scholar]
  18. Hoopes B. C., McClure W. R. A cII-dependent promoter is located within the Q gene of bacteriophage lambda. Proc Natl Acad Sci U S A. 1985 May;82(10):3134–3138. doi: 10.1073/pnas.82.10.3134. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hoyt M. A., Knight D. M., Das A., Miller H. I., Echols H. Control of phage lambda development by stability and synthesis of cII protein: role of the viral cIII and host hflA, himA and himD genes. Cell. 1982 Dec;31(3 Pt 2):565–573. doi: 10.1016/0092-8674(82)90312-9. [DOI] [PubMed] [Google Scholar]
  20. Ito H., Ike Y., Ikuta S., Itakura K. Solid phase synthesis of polynucleotides. VI. Further studies on polystyrene copolymers for the solid support. Nucleic Acids Res. 1982 Mar 11;10(5):1755–1769. doi: 10.1093/nar/10.5.1755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Johnson A. D., Poteete A. R., Lauer G., Sauer R. T., Ackers G. K., Ptashne M. lambda Repressor and cro--components of an efficient molecular switch. Nature. 1981 Nov 19;294(5838):217–223. doi: 10.1038/294217a0. [DOI] [PubMed] [Google Scholar]
  22. Katzir N., Oppenheim A., Belfort M., Oppenheim A. B. Activation of the lambda int gene by the cii and ciii gene products. Virology. 1976 Oct 15;74(2):324–331. doi: 10.1016/0042-6822(76)90339-1. [DOI] [PubMed] [Google Scholar]
  23. Kikuchi A., Flamm E., Weisberg R. A. An Escherichia coli mutant unable to support site-specific recombination of bacteriophage lambda. J Mol Biol. 1985 May 25;183(2):129–140. doi: 10.1016/0022-2836(85)90207-4. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. Lange-Gustafson B. J., Nash H. A. Purification and properties of Int-h, a variant protein involved in site-specific recombination of bacteriophage lambda. J Biol Chem. 1984 Oct 25;259(20):12724–12732. [PubMed] [Google Scholar]
  26. Leong J. M., Nunes-Düby S., Lesser C. F., Youderian P., Susskind M. M., Landy A. The phi 80 and P22 attachment sites. Primary structure and interaction with Escherichia coli integration host factor. J Biol Chem. 1985 Apr 10;260(7):4468–4477. [PubMed] [Google Scholar]
  27. Mahajna J., Oppenheim A. B., Rattray A., Gottesman M. Translation initiation of bacteriophage lambda gene cII requires integration host factor. J Bacteriol. 1986 Jan;165(1):167–174. doi: 10.1128/jb.165.1.167-174.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Miller H. I., Friedman D. I. An E. coli gene product required for lambda site-specific recombination. Cell. 1980 Jul;20(3):711–719. doi: 10.1016/0092-8674(80)90317-7. [DOI] [PubMed] [Google Scholar]
  29. Miller H. I., Kirk M., Echols H. SOS induction and autoregulation of the himA gene for site-specific recombination in Escherichia coli. Proc Natl Acad Sci U S A. 1981 Nov;78(11):6754–6758. doi: 10.1073/pnas.78.11.6754. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Miller H. I., Mozola M. A., Friedman D. I. int-h: An int mutation of phage lambda that enhances site-specific recombination. Cell. 1980 Jul;20(3):721–729. doi: 10.1016/0092-8674(80)90318-9. [DOI] [PubMed] [Google Scholar]
  31. Miller H. I. Multilevel regulation of bacteriophage lambda lysogeny by the E. coli himA gene. Cell. 1981 Jul;25(1):269–276. doi: 10.1016/0092-8674(81)90252-x. [DOI] [PubMed] [Google Scholar]
  32. Mizuuchi K., Weisberg R., Enquist L., Mizuuchi M., Buraczynska M., Foeller C., Hsu P. L., Ross W., Landy A. Structure and function of the phage lambda att site: size, int-binding sites, and location of the crossover point. Cold Spring Harb Symp Quant Biol. 1981;45(Pt 1):429–437. doi: 10.1101/sqb.1981.045.01.057. [DOI] [PubMed] [Google Scholar]
  33. Mizuuchi M., Mizuuchi K. The extent of DNA sequence required for a functional bacterial attachment site of phage lambda. Nucleic Acids Res. 1985 Feb 25;13(4):1193–1208. doi: 10.1093/nar/13.4.1193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. 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]
  35. Peacock S., Weissbach H., Nash H. A. In vitro regulation of phage lambda cII gene expression by Escherichia coli integration host factor. Proc Natl Acad Sci U S A. 1984 Oct;81(19):6009–6013. doi: 10.1073/pnas.81.19.6009. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. 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]
  37. SUSSMAN R., JACOB F. [On a thermosensitive repression system in the Escherichia coli lambda bacteriophage]. C R Hebd Seances Acad Sci. 1962 Feb 19;254:1517–1519. [PubMed] [Google Scholar]
  38. Schmeissner U., Court D., Shimatake H., Rosenberg M. Promoter for the establishment of repressor synthesis in bacteriophage lambda. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3191–3195. doi: 10.1073/pnas.77.6.3191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Shea M. A., Ackers G. K. The OR control system of bacteriophage lambda. A physical-chemical model for gene regulation. J Mol Biol. 1985 Jan 20;181(2):211–230. doi: 10.1016/0022-2836(85)90086-5. [DOI] [PubMed] [Google Scholar]
  40. Shimada K., Weisberg R. A., Gottesman M. E. Prophage lambda at unusual chromosomal locations. I. Location of the secondary attachment sites and the properties of the lysogens. J Mol Biol. 1972 Feb 14;63(3):483–503. doi: 10.1016/0022-2836(72)90443-3. [DOI] [PubMed] [Google Scholar]
  41. Shulman M. J., Mizuuchi K., Gottesman M. M. New att mutants of phage lambda. Virology. 1976 Jul 1;72(1):13–22. doi: 10.1016/0042-6822(76)90307-x. [DOI] [PubMed] [Google Scholar]
  42. Shulman M., Gottesman M. Attachment site mutants of bacteriophage lambda. J Mol Biol. 1973 Dec 25;81(4):461–482. doi: 10.1016/0022-2836(73)90517-2. [DOI] [PubMed] [Google Scholar]
  43. Simatake H., Rosenberg M. Purified lambda regulatory protein cII positively activates promoters for lysogenic development. Nature. 1981 Jul 9;292(5819):128–132. doi: 10.1038/292128a0. [DOI] [PubMed] [Google Scholar]
  44. Sly W. S., Rabideau K. Mechanism of lambda-c17cI virulence. J Mol Biol. 1969 Jun 28;42(3):385–400. doi: 10.1016/0022-2836(69)90231-9. [DOI] [PubMed] [Google Scholar]
  45. Stephenson F. H. A CII-responsive promoter within the Q gene of bacteriophage lambda. Gene. 1985;35(3):313–320. doi: 10.1016/0378-1119(85)90010-1. [DOI] [PubMed] [Google Scholar]
  46. Weisberg R. A., Enquist L. W., Foeller C., Landy A. Role for DNA homology in site-specific recombination. The isolation and characterization of a site affinity mutant of coliphage lambda. J Mol Biol. 1983 Oct 25;170(2):319–342. doi: 10.1016/s0022-2836(83)80151-x. [DOI] [PubMed] [Google Scholar]
  47. Wu H. M., Crothers D. M. The locus of sequence-directed and protein-induced DNA bending. Nature. 1984 Apr 5;308(5959):509–513. doi: 10.1038/308509a0. [DOI] [PubMed] [Google Scholar]
  48. Wulff D. L., Mahoney M., Shatzman A., Rosenberg M. Mutational analysis of a regulatory region in bacteriophage lambda that has overlapping signals for the initiation of transcription and translation. Proc Natl Acad Sci U S A. 1984 Jan;81(2):555–559. doi: 10.1073/pnas.81.2.555. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. 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 Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES