Skip to main content
The EMBO Journal logoLink to The EMBO Journal
. 1997 May 1;16(9):2535–2544. doi: 10.1093/emboj/16.9.2535

Recombinational repair in yeast: functional interactions between Rad51 and Rad54 proteins.

B Clever 1, H Interthal 1, J Schmuckli-Maurer 1, J King 1, M Sigrist 1, W D Heyer 1
PMCID: PMC1169853  PMID: 9171366

Abstract

Rad51p is a eukaryotic homolog of RecA, the central homologous pairing and strand exchange protein in Escherichia coli. Rad54p belongs to the Swi2p/Snf2p family of DNA-stimulated ATPases. Both proteins are also important members of the RAD52 group which controls recombinational DNA damage repair of double-strand breaks and other DNA lesions in Saccharomyces cerevisiae. Here we demonstrate by genetic, molecular and biochemical criteria that Rad51 and Rad54 proteins interact. Strikingly, overexpression of Rad54p can functionally suppress the UV and methyl methanesulfonate sensitivity caused by a deletion of the RAD51 gene. However, no suppression was observed for the defects of rad51 cells in the repair of gamma-ray-induced DNA damage, mating type switching or spontaneous hetero-allelic recombination. This suppression is genetically dependent on the presence of two other members of the recombinational repair group, RAD55 and RAD57. Our data provide compelling evidence that Rad51 and Rad54 proteins interact in vivo and that this interaction is functionally important for recombinational DNA damage repair. As both proteins are conserved throughout evolution from yeasts to humans, a similar protein-protein interaction may be expected in other organisms.

Full Text

The Full Text of this article is available as a PDF (393.2 KB).

Selected References

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

  1. Aboussekhra A., Chanet R., Adjiri A., Fabre F. Semidominant suppressors of Srs2 helicase mutations of Saccharomyces cerevisiae map in the RAD51 gene, whose sequence predicts a protein with similarities to procaryotic RecA proteins. Mol Cell Biol. 1992 Jul;12(7):3224–3234. doi: 10.1128/mcb.12.7.3224. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Auble D. T., Hansen K. E., Mueller C. G., Lane W. S., Thorner J., Hahn S. Mot1, a global repressor of RNA polymerase II transcription, inhibits TBP binding to DNA by an ATP-dependent mechanism. Genes Dev. 1994 Aug 15;8(16):1920–1934. doi: 10.1101/gad.8.16.1920. [DOI] [PubMed] [Google Scholar]
  3. Basile G., Aker M., Mortimer R. K. Nucleotide sequence and transcriptional regulation of the yeast recombinational repair gene RAD51. Mol Cell Biol. 1992 Jul;12(7):3235–3246. doi: 10.1128/mcb.12.7.3235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bishop D. K., Park D., Xu L., Kleckner N. DMC1: a meiosis-specific yeast homolog of E. coli recA required for recombination, synaptonemal complex formation, and cell cycle progression. Cell. 1992 May 1;69(3):439–456. doi: 10.1016/0092-8674(92)90446-j. [DOI] [PubMed] [Google Scholar]
  5. Broach J. R., Strathern J. N., Hicks J. B. Transformation in yeast: development of a hybrid cloning vector and isolation of the CAN1 gene. Gene. 1979 Dec;8(1):121–133. doi: 10.1016/0378-1119(79)90012-x. [DOI] [PubMed] [Google Scholar]
  6. Bähler J., Hagens G., Holzinger G., Scherthan H., Heyer W. D. Saccharomyces cerevisiae cells lacking the homologous pairing protein p175SEP1 arrest at pachytene during meiotic prophase. Chromosoma. 1994 Apr;103(2):129–141. doi: 10.1007/BF00352322. [DOI] [PubMed] [Google Scholar]
  7. Donovan J. W., Milne G. T., Weaver D. T. Homotypic and heterotypic protein associations control Rad51 function in double-strand break repair. Genes Dev. 1994 Nov 1;8(21):2552–2562. doi: 10.1101/gad.8.21.2552. [DOI] [PubMed] [Google Scholar]
  8. Eisen J. A., Sweder K. S., Hanawalt P. C. Evolution of the SNF2 family of proteins: subfamilies with distinct sequences and functions. Nucleic Acids Res. 1995 Jul 25;23(14):2715–2723. doi: 10.1093/nar/23.14.2715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Emery H. S., Schild D., Kellogg D. E., Mortimer R. K. Sequence of RAD54, a Saccharomyces cerevisiae gene involved in recombination and repair. Gene. 1991 Jul 31;104(1):103–106. doi: 10.1016/0378-1119(91)90473-o. [DOI] [PubMed] [Google Scholar]
  10. Fields S., Song O. A novel genetic system to detect protein-protein interactions. Nature. 1989 Jul 20;340(6230):245–246. doi: 10.1038/340245a0. [DOI] [PubMed] [Google Scholar]
  11. Firmenich A. A., Elias-Arnanz M., Berg P. A novel allele of Saccharomyces cerevisiae RFA1 that is deficient in recombination and repair and suppressible by RAD52. Mol Cell Biol. 1995 Mar;15(3):1620–1631. doi: 10.1128/mcb.15.3.1620. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Fishman-Lobell J., Rudin N., Haber J. E. Two alternative pathways of double-strand break repair that are kinetically separable and independently modulated. Mol Cell Biol. 1992 Mar;12(3):1292–1303. doi: 10.1128/mcb.12.3.1292. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Game J. C. DNA double-strand breaks and the RAD50-RAD57 genes in Saccharomyces. Semin Cancer Biol. 1993 Apr;4(2):73–83. [PubMed] [Google Scholar]
  14. Gyuris J., Golemis E., Chertkov H., Brent R. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 1993 Nov 19;75(4):791–803. doi: 10.1016/0092-8674(93)90498-f. [DOI] [PubMed] [Google Scholar]
  15. Harshman K. D., Moye-Rowley W. S., Parker C. S. Transcriptional activation by the SV40 AP-1 recognition element in yeast is mediated by a factor similar to AP-1 that is distinct from GCN4. Cell. 1988 Apr 22;53(2):321–330. doi: 10.1016/0092-8674(88)90393-5. [DOI] [PubMed] [Google Scholar]
  16. Hoeijmakers J. H. Nucleotide excision repair. II: From yeast to mammals. Trends Genet. 1993 Jun;9(6):211–217. doi: 10.1016/0168-9525(93)90121-w. [DOI] [PubMed] [Google Scholar]
  17. Holler A., Bashkirov V. I., Solinger J. A., Reinhart U., Heyer W. D. Use of monoclonal antibodies in the functional characterization of the Saccharomyces cerevisiae Sep1 protein. Eur J Biochem. 1995 Jul 15;231(2):329–336. doi: 10.1111/j.1432-1033.1995.tb20704.x. [DOI] [PubMed] [Google Scholar]
  18. Jiang H., Xie Y., Houston P., Stemke-Hale K., Mortensen U. H., Rothstein R., Kodadek T. Direct association between the yeast Rad51 and Rad54 recombination proteins. J Biol Chem. 1996 Dec 27;271(52):33181–33186. doi: 10.1074/jbc.271.52.33181. [DOI] [PubMed] [Google Scholar]
  19. Johnson R. D., Symington L. S. Functional differences and interactions among the putative RecA homologs Rad51, Rad55, and Rad57. Mol Cell Biol. 1995 Sep;15(9):4843–4850. doi: 10.1128/mcb.15.9.4843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kans J. A., Mortimer R. K. Nucleotide sequence of the RAD57 gene of Saccharomyces cerevisiae. Gene. 1991 Aug 30;105(1):139–140. doi: 10.1016/0378-1119(91)90527-i. [DOI] [PubMed] [Google Scholar]
  21. Kowalczykowski S. C., Eggleston A. K. Homologous pairing and DNA strand-exchange proteins. Annu Rev Biochem. 1994;63:991–1043. doi: 10.1146/annurev.bi.63.070194.005015. [DOI] [PubMed] [Google Scholar]
  22. Muris D. F., Bezzubova O., Buerstedde J. M., Vreeken K., Balajee A. S., Osgood C. J., Troelstra C., Hoeijmakers J. H., Ostermann K., Schmidt H. Cloning of human and mouse genes homologous to RAD52, a yeast gene involved in DNA repair and recombination. Mutat Res. 1994 Nov;315(3):295–305. doi: 10.1016/0921-8777(94)90040-x. [DOI] [PubMed] [Google Scholar]
  23. Muris D. F., Vreeken K., Carr A. M., Murray J. M., Smit C., Lohman P. H., Pastink A. Isolation of the Schizosaccharomyces pombe RAD54 homologue, rhp54+, a gene involved in the repair of radiation damage and replication fidelity. J Cell Sci. 1996 Jan;109(Pt 1):73–81. doi: 10.1242/jcs.109.1.73. [DOI] [PubMed] [Google Scholar]
  24. Ogawa T., Yu X., Shinohara A., Egelman E. H. Similarity of the yeast RAD51 filament to the bacterial RecA filament. Science. 1993 Mar 26;259(5103):1896–1899. doi: 10.1126/science.8456314. [DOI] [PubMed] [Google Scholar]
  25. Peterson C. L., Tamkun J. W. The SWI-SNF complex: a chromatin remodeling machine? Trends Biochem Sci. 1995 Apr;20(4):143–146. doi: 10.1016/s0968-0004(00)88990-2. [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. Salinas F., Kodadek T. Phage T4 homologous strand exchange: a DNA helicase, not the strand transferase, drives polar branch migration. Cell. 1995 Jul 14;82(1):111–119. doi: 10.1016/0092-8674(95)90057-8. [DOI] [PubMed] [Google Scholar]
  28. Sancar A., Hearst J. E. Molecular matchmakers. Science. 1993 Mar 5;259(5100):1415–1420. doi: 10.1126/science.8451638. [DOI] [PubMed] [Google Scholar]
  29. Sandler S. J., Satin L. H., Samra H. S., Clark A. J. recA-like genes from three archaean species with putative protein products similar to Rad51 and Dmc1 proteins of the yeast Saccharomyces cerevisiae. Nucleic Acids Res. 1996 Jun 1;24(11):2125–2132. doi: 10.1093/nar/24.11.2125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Santos-Rosa H., Clever B., Heyer W. D., Aguilera A. The yeast HRS1 gene encodes a polyglutamine-rich nuclear protein required for spontaneous and hpr1-induced deletions between direct repeats. Genetics. 1996 Mar;142(3):705–716. doi: 10.1093/genetics/142.3.705. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Schiestl R. H., Gietz R. D. High efficiency transformation of intact yeast cells using single stranded nucleic acids as a carrier. Curr Genet. 1989 Dec;16(5-6):339–346. doi: 10.1007/BF00340712. [DOI] [PubMed] [Google Scholar]
  32. Schild D. Suppression of a new allele of the yeast RAD52 gene by overexpression of RAD51, mutations in srs2 and ccr4, or mating-type heterozygosity. Genetics. 1995 May;140(1):115–127. doi: 10.1093/genetics/140.1.115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Shinohara A., Ogawa H., Matsuda Y., Ushio N., Ikeo K., Ogawa T. Cloning of human, mouse and fission yeast recombination genes homologous to RAD51 and recA. Nat Genet. 1993 Jul;4(3):239–243. doi: 10.1038/ng0793-239. [DOI] [PubMed] [Google Scholar]
  34. Shinohara A., Ogawa H., Ogawa T. Rad51 protein involved in repair and recombination in S. cerevisiae is a RecA-like protein. Cell. 1992 May 1;69(3):457–470. doi: 10.1016/0092-8674(92)90447-k. [DOI] [PubMed] [Google Scholar]
  35. Shinohara A., Ogawa T. Homologous recombination and the roles of double-strand breaks. Trends Biochem Sci. 1995 Oct;20(10):387–391. doi: 10.1016/s0968-0004(00)89085-4. [DOI] [PubMed] [Google Scholar]
  36. Story R. M., Bishop D. K., Kleckner N., Steitz T. A. Structural relationship of bacterial RecA proteins to recombination proteins from bacteriophage T4 and yeast. Science. 1993 Mar 26;259(5103):1892–1896. doi: 10.1126/science.8456313. [DOI] [PubMed] [Google Scholar]
  37. Story R. M., Steitz T. A. Structure of the recA protein-ADP complex. Nature. 1992 Jan 23;355(6358):374–376. doi: 10.1038/355374a0. [DOI] [PubMed] [Google Scholar]
  38. Story R. M., Weber I. T., Steitz T. A. The structure of the E. coli recA protein monomer and polymer. Nature. 1992 Jan 23;355(6358):318–325. doi: 10.1038/355318a0. [DOI] [PubMed] [Google Scholar]
  39. Struhl K. Yeast transcriptional regulatory mechanisms. Annu Rev Genet. 1995;29:651–674. doi: 10.1146/annurev.ge.29.120195.003251. [DOI] [PubMed] [Google Scholar]
  40. Subramanya H. S., Bird L. E., Brannigan J. A., Wigley D. B. Crystal structure of a DExx box DNA helicase. Nature. 1996 Nov 28;384(6607):379–383. doi: 10.1038/384379a0. [DOI] [PubMed] [Google Scholar]
  41. Sugawara N., Ivanov E. L., Fishman-Lobell J., Ray B. L., Wu X., Haber J. E. DNA structure-dependent requirements for yeast RAD genes in gene conversion. Nature. 1995 Jan 5;373(6509):84–86. doi: 10.1038/373084a0. [DOI] [PubMed] [Google Scholar]
  42. Sung P. Catalysis of ATP-dependent homologous DNA pairing and strand exchange by yeast RAD51 protein. Science. 1994 Aug 26;265(5176):1241–1243. doi: 10.1126/science.8066464. [DOI] [PubMed] [Google Scholar]
  43. Sung P., Robberson D. L. DNA strand exchange mediated by a RAD51-ssDNA nucleoprotein filament with polarity opposite to that of RecA. Cell. 1995 Aug 11;82(3):453–461. doi: 10.1016/0092-8674(95)90434-4. [DOI] [PubMed] [Google Scholar]
  44. Svejstrup J. Q., Wang Z., Feaver W. J., Wu X., Bushnell D. A., Donahue T. F., Friedberg E. C., Kornberg R. D. Different forms of TFIIH for transcription and DNA repair: holo-TFIIH and a nucleotide excision repairosome. Cell. 1995 Jan 13;80(1):21–28. doi: 10.1016/0092-8674(95)90447-6. [DOI] [PubMed] [Google Scholar]
  45. Wach A., Brachat A., Pöhlmann R., Philippsen P. New heterologous modules for classical or PCR-based gene disruptions in Saccharomyces cerevisiae. Yeast. 1994 Dec;10(13):1793–1808. doi: 10.1002/yea.320101310. [DOI] [PubMed] [Google Scholar]
  46. Weaver D. T. What to do at an end: DNA double-strand-break repair. Trends Genet. 1995 Oct;11(10):388–392. doi: 10.1016/s0168-9525(00)89121-0. [DOI] [PubMed] [Google Scholar]
  47. West S. C. Enzymes and molecular mechanisms of genetic recombination. Annu Rev Biochem. 1992;61:603–640. doi: 10.1146/annurev.bi.61.070192.003131. [DOI] [PubMed] [Google Scholar]
  48. Wood R. D. DNA repair in eukaryotes. Annu Rev Biochem. 1996;65:135–167. doi: 10.1146/annurev.bi.65.070196.001031. [DOI] [PubMed] [Google Scholar]

Articles from The EMBO Journal are provided here courtesy of Nature Publishing Group

RESOURCES