Skip to main content
Genetics logoLink to Genetics
. 1995 Mar;139(3):1189–1199. doi: 10.1093/genetics/139.3.1189

Modulation of Saccharomyces Cerevisiae DNA Double-Strand Break Repair by Srs2 and Rad51

G T Milne 1, T Ho 1, D T Weaver 1
PMCID: PMC1206449  PMID: 7768432

Abstract

RAD52 function is required for virtually all DNA double-strand break repair and recombination events in Saccharomyces cerevisiae. To gain greater insight into the mechanism of RAD52-mediated repair, we screened for genes that suppress partially active alleles of RAD52 when mutant or overexpressed. Described here is the isolation of a phenotypic null allele of SRS2 that suppressed multiple alleles of RAD52 (rad52B, rad52D, rad52-1 and KlRAD52) and RAD51 (KlRAD51) but failed to suppress either a rad52δ or a rad51δ. These results indicate that SRS2 antagonizes RAD51 and RAD52 function in recombinational repair. The mechanism of suppression of RAD52 alleles by srs2 is distinct from that which has been previously described for RAD51 overexpression, as both conditions were shown to act additively with respect to the rad52B allele. Furthermore, overexpression of either RAD52 or RAD51 enhanced the recombination-dependent sensitivity of an srs2δ RAD52 strain, suggesting that RAD52 and RAD51 positively influence recombinational repair mechanisms. Thus, RAD52-dependent recombinational repair is controlled both negatively and positively.

Full Text

The Full Text of this article is available as a PDF (1.1 MB).

Selected References

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

  1. Aguilera A., Klein H. L. Genetic control of intrachromosomal recombination in Saccharomyces cerevisiae. I. Isolation and genetic characterization of hyper-recombination mutations. Genetics. 1988 Aug;119(4):779–790. doi: 10.1093/genetics/119.4.779. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Barnes G., Rine J. Regulated expression of endonuclease EcoRI in Saccharomyces cerevisiae: nuclear entry and biological consequences. Proc Natl Acad Sci U S A. 1985 Mar;82(5):1354–1358. doi: 10.1073/pnas.82.5.1354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Becker D. M., Fikes J. D., Guarente L. A cDNA encoding a human CCAAT-binding protein cloned by functional complementation in yeast. Proc Natl Acad Sci U S A. 1991 Mar 1;88(5):1968–1972. doi: 10.1073/pnas.88.5.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Berg C. M., Vartak N. B., Wang G., Xu X., Liu L., MacNeil D. J., Gewain K. M., Wiater L. A., Berg D. E. The m gamma delta-1 element, a small gamma delta (Tn1000) derivative useful for plasmid mutagenesis, allele replacement and DNA sequencing. Gene. 1992 Apr 1;113(1):9–16. doi: 10.1016/0378-1119(92)90664-b. [DOI] [PubMed] [Google Scholar]
  5. Carlson M., Botstein D. Two differentially regulated mRNAs with different 5' ends encode secreted with intracellular forms of yeast invertase. Cell. 1982 Jan;28(1):145–154. doi: 10.1016/0092-8674(82)90384-1. [DOI] [PubMed] [Google Scholar]
  6. Chien C. T., Bartel P. L., Sternglanz R., Fields S. The two-hybrid system: a method to identify and clone genes for proteins that interact with a protein of interest. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9578–9582. doi: 10.1073/pnas.88.21.9578. [DOI] [PMC free article] [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. Game J. C., Cox B. S. Synergistic interactions between rad mutations in yeast. Mutat Res. 1973 Oct;20(1):35–44. doi: 10.1016/0027-5107(73)90095-x. [DOI] [PubMed] [Google Scholar]
  9. Gietz D., St Jean A., Woods R. A., Schiestl R. H. Improved method for high efficiency transformation of intact yeast cells. Nucleic Acids Res. 1992 Mar 25;20(6):1425–1425. doi: 10.1093/nar/20.6.1425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Glassner B. J., Mortimer R. K. Synergistic interactions between RAD5, RAD16 and RAD54, three partially homologous yeast DNA repair genes each in a different repair pathway. Radiat Res. 1994 Jul;139(1):24–33. [PubMed] [Google Scholar]
  11. Herskowitz I., Jensen R. E. Putting the HO gene to work: practical uses for mating-type switching. Methods Enzymol. 1991;194:132–146. doi: 10.1016/0076-6879(91)94011-z. [DOI] [PubMed] [Google Scholar]
  12. Hill J., Donald K. A., Griffiths D. E., Donald G. DMSO-enhanced whole cell yeast transformation. Nucleic Acids Res. 1991 Oct 25;19(20):5791–5791. doi: 10.1093/nar/19.20.5791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hoffman C. S., Winston F. A ten-minute DNA preparation from yeast efficiently releases autonomous plasmids for transformation of Escherichia coli. Gene. 1987;57(2-3):267–272. doi: 10.1016/0378-1119(87)90131-4. [DOI] [PubMed] [Google Scholar]
  14. Lawrence C. W., Christensen R. B. Metabolic suppressors of trimethoprim and ultraviolet light sensitivities of Saccharomyces cerevisiae rad6 mutants. J Bacteriol. 1979 Sep;139(3):866–876. doi: 10.1128/jb.139.3.866-876.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Milne G. T., Weaver D. T. Dominant negative alleles of RAD52 reveal a DNA repair/recombination complex including Rad51 and Rad52. Genes Dev. 1993 Sep;7(9):1755–1765. doi: 10.1101/gad.7.9.1755. [DOI] [PubMed] [Google Scholar]
  16. Nakai S., Matsumoto S. Two types of radiation-sensitive mutant in yeast. Mutat Res. 1967 Mar-Apr;4(2):129–136. doi: 10.1016/0027-5107(67)90064-4. [DOI] [PubMed] [Google Scholar]
  17. 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]
  18. Palladino F., Klein H. L. Analysis of mitotic and meiotic defects in Saccharomyces cerevisiae SRS2 DNA helicase mutants. Genetics. 1992 Sep;132(1):23–37. doi: 10.1093/genetics/132.1.23. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rong L., Klein H. L. Purification and characterization of the SRS2 DNA helicase of the yeast Saccharomyces cerevisiae. J Biol Chem. 1993 Jan 15;268(2):1252–1259. [PubMed] [Google Scholar]
  20. Rong L., Palladino F., Aguilera A., Klein H. L. The hyper-gene conversion hpr5-1 mutation of Saccharomyces cerevisiae is an allele of the SRS2/RADH gene. Genetics. 1991 Jan;127(1):75–85. doi: 10.1093/genetics/127.1.75. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Rose M. D., Novick P., Thomas J. H., Botstein D., Fink G. R. A Saccharomyces cerevisiae genomic plasmid bank based on a centromere-containing shuttle vector. Gene. 1987;60(2-3):237–243. doi: 10.1016/0378-1119(87)90232-0. [DOI] [PubMed] [Google Scholar]
  22. Rothstein R. Targeting, disruption, replacement, and allele rescue: integrative DNA transformation in yeast. Methods Enzymol. 1991;194:281–301. doi: 10.1016/0076-6879(91)94022-5. [DOI] [PubMed] [Google Scholar]
  23. Schiestl R. H., Prakash S. Interactions of the RAD7 and RAD23 excision repair genes of Saccharomyces cerevisiae with DNA repair genes in different epistasis groups. Curr Genet. 1989 Oct;16(4):219–223. doi: 10.1007/BF00422107. [DOI] [PubMed] [Google Scholar]
  24. 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]
  25. 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]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES