Skip to main content
PMC home page
Genetics logoLink to Genetics
. 1989 Dec;123(4):725–738. doi: 10.1093/genetics/123.4.725

The Genetic Control of Direct-Repeat Recombination in Saccharomyces: The Effect of Rad52 and Rad1 on Mitotic Recombination at Gal10, a Transcriptionally Regulated Gene

B J Thomas 1, R Rothstein 1
PMCID: PMC1203884  PMID: 2693208

Abstract

We have previously shown direct-repeat recombination events leading to loss of a plasmid integrated at the GAL10 locus in Saccharomyces cerevisiae are stimulated by transcription of the region. We have examined the role of two recombination- and repair-defective mutations, rad1 and rad52, on direct repeat recombination in transcriptionally active and inactive sequences. We show that the RAD52 gene is required for transcription-stimulated recombination events leading to loss of the integrated plasmid. Similarly, Gal(+) events between the duplicated repeats that retain the integrated plasmid DNA (Gal(+) Ura(+) replacement events) are reduced 20-fold in the rad52 mutant in sequences that are constitutively expressed. In contrast, in sequences that are not expressed, the rad52 mutation reduces plasmid loss events by only twofold and Gal(+) Ura(+) replacements by fourfold. We also observe an increase in disome-associated plasmid loss events in the rad52 mutant, indicative of chromosome gain. This event is not affected by expression of the region. Plasmid loss events in rad1 mutant strains are reduced only twofold in transcriptionally active sequences and are not affected in sequences that are repressed. However, the rad1 and rad52 double mutant shows a decrease in plasmid loss events greater than the sum of the decreases in the rates of this event displayed by either single mutant in both constitutive and repressed DNA, indicating a synergistic interaction between these two genes. The synergism is limited to recombination since the rad1 rad52 double mutant is no more sensitive when compared with either single mutant in its ability to survive radiation damage. Finally, the recombination pathway that remains in the double mutant is positively affected by transcription of the region.

Full Text

The Full Text of this article is available as a PDF (2.6 MB).

Selected References

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

  1. Boeke J. D., LaCroute F., Fink G. R. A positive selection for mutants lacking orotidine-5'-phosphate decarboxylase activity in yeast: 5-fluoro-orotic acid resistance. Mol Gen Genet. 1984;197(2):345–346. doi: 10.1007/BF00330984. [DOI] [PubMed] [Google Scholar]
  2. Bohr V. A., Smith C. A., Okumoto D. S., Hanawalt P. C. DNA repair in an active gene: removal of pyrimidine dimers from the DHFR gene of CHO cells is much more efficient than in the genome overall. Cell. 1985 Feb;40(2):359–369. doi: 10.1016/0092-8674(85)90150-3. [DOI] [PubMed] [Google Scholar]
  3. Douglas H. C., Hawthorne D. C. Regulation of genes controlling synthesis of the galactose pathway enzymes in yeast. Genetics. 1966 Sep;54(3):911–916. doi: 10.1093/genetics/54.3.911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Esposito M. S., Hosoda J., Golin J., Moise H., Bjornstad K., Maleas D. Recombination in Saccharomyces cerevisiae: REC-gene mutants and DNA-binding proteins. Cold Spring Harb Symp Quant Biol. 1984;49:41–48. doi: 10.1101/sqb.1984.049.01.007. [DOI] [PubMed] [Google Scholar]
  5. Game J. C., Mortimer R. K. A genetic study of x-ray sensitive mutants in yeast. Mutat Res. 1974 Sep;24(3):281–292. doi: 10.1016/0027-5107(74)90176-6. [DOI] [PubMed] [Google Scholar]
  6. Haber J. E., Hearn M. Rad52-independent mitotic gene conversion in Saccharomyces cerevisiae frequently results in chromosomal loss. Genetics. 1985 Sep;111(1):7–22. doi: 10.1093/genetics/111.1.7. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hartwell L. H., Smith D. Altered fidelity of mitotic chromosome transmission in cell cycle mutants of S. cerevisiae. Genetics. 1985 Jul;110(3):381–395. doi: 10.1093/genetics/110.3.381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Ho K. S. Induction of DNA double-strand breaks by X-rays in a radiosensitive strain of the yeast Saccharomyces cerevisiae. Mutat Res. 1975 Dec;30(3):327–334. [PubMed] [Google Scholar]
  9. Hoekstra M. F., Naughton T., Malone R. E. Properties of spontaneous mitotic recombination occurring in the presence of the rad52-1 mutation of Saccharomyces cerevisiae. Genet Res. 1986 Aug;48(1):9–17. doi: 10.1017/s0016672300024599. [DOI] [PubMed] [Google Scholar]
  10. Hollingsworth N. M., Byers B. HOP1: a yeast meiotic pairing gene. Genetics. 1989 Mar;121(3):445–462. doi: 10.1093/genetics/121.3.445. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jackson J. A., Fink G. R. Gene conversion between duplicated genetic elements in yeast. Nature. 1981 Jul 23;292(5821):306–311. doi: 10.1038/292306a0. [DOI] [PubMed] [Google Scholar]
  13. Jinks-Robertson S., Petes T. D. Chromosomal translocations generated by high-frequency meiotic recombination between repeated yeast genes. Genetics. 1986 Nov;114(3):731–752. doi: 10.1093/genetics/114.3.731. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Keil R. L., Roeder G. S. Cis-acting, recombination-stimulating activity in a fragment of the ribosomal DNA of S. cerevisiae. Cell. 1984 Dec;39(2 Pt 1):377–386. doi: 10.1016/0092-8674(84)90016-3. [DOI] [PubMed] [Google Scholar]
  15. Klein H. L. Different types of recombination events are controlled by the RAD1 and RAD52 genes of Saccharomyces cerevisiae. Genetics. 1988 Oct;120(2):367–377. doi: 10.1093/genetics/120.2.367. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Kunz B. A., Haynes R. H. Phenomenology and genetic control of mitotic recombination in yeast. Annu Rev Genet. 1981;15:57–89. doi: 10.1146/annurev.ge.15.120181.000421. [DOI] [PubMed] [Google Scholar]
  17. Lichten M., Borts R. H., Haber J. E. Meiotic gene conversion and crossing over between dispersed homologous sequences occurs frequently in Saccharomyces cerevisiae. Genetics. 1987 Feb;115(2):233–246. doi: 10.1093/genetics/115.2.233. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Luria S. E., Delbrück M. Mutations of Bacteria from Virus Sensitivity to Virus Resistance. Genetics. 1943 Nov;28(6):491–511. doi: 10.1093/genetics/28.6.491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Madhani H. D., Bohr V. A., Hanawalt P. C. Differential DNA repair in transcriptionally active and inactive proto-oncogenes: c-abl and c-mos. Cell. 1986 May 9;45(3):417–423. doi: 10.1016/0092-8674(86)90327-2. [DOI] [PubMed] [Google Scholar]
  20. Malone R. E., Esposito R. E. The RAD52 gene is required for homothallic interconversion of mating types and spontaneous mitotic recombination in yeast. Proc Natl Acad Sci U S A. 1980 Jan;77(1):503–507. doi: 10.1073/pnas.77.1.503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Mellon I., Bohr V. A., Smith C. A., Hanawalt P. C. Preferential DNA repair of an active gene in human cells. Proc Natl Acad Sci U S A. 1986 Dec;83(23):8878–8882. doi: 10.1073/pnas.83.23.8878. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Mellon I., Spivak G., Hanawalt P. C. Selective removal of transcription-blocking DNA damage from the transcribed strand of the mammalian DHFR gene. Cell. 1987 Oct 23;51(2):241–249. doi: 10.1016/0092-8674(87)90151-6. [DOI] [PubMed] [Google Scholar]
  23. Meselson M. S., Radding C. M. A general model for genetic recombination. Proc Natl Acad Sci U S A. 1975 Jan;72(1):358–361. doi: 10.1073/pnas.72.1.358. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Mikus M. D., Petes T. D. Recombination between genes located on nonhomologous chromosomes in Saccharomyces cerevisiae. Genetics. 1982 Jul-Aug;101(3-4):369–404. doi: 10.1093/genetics/101.3-4.369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Modrich P. DNA mismatch correction. Annu Rev Biochem. 1987;56:435–466. doi: 10.1146/annurev.bi.56.070187.002251. [DOI] [PubMed] [Google Scholar]
  26. Mortimer R. K., Contopoulou R., Schild D. Mitotic chromosome loss in a radiation-sensitive strain of the yeast Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1981 Sep;78(9):5778–5782. doi: 10.1073/pnas.78.9.5778. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Nickoloff J. A., Singer J. D., Hoekstra M. F., Heffron F. Double-strand breaks stimulate alternative mechanisms of recombination repair. J Mol Biol. 1989 Jun 5;207(3):527–541. doi: 10.1016/0022-2836(89)90462-2. [DOI] [PubMed] [Google Scholar]
  28. 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]
  29. Resnick M. A. Genetic control of radiation sensitivity in Saccharomyces cerevisiae. Genetics. 1969 Jul;62(3):519–531. doi: 10.1093/genetics/62.3.519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Resnick M. A., Martin P. The repair of double-strand breaks in the nuclear DNA of Saccharomyces cerevisiae and its genetic control. Mol Gen Genet. 1976 Jan 16;143(2):119–129. doi: 10.1007/BF00266917. [DOI] [PubMed] [Google Scholar]
  31. 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]
  32. Reynolds R. J., Friedberg E. C. Molecular mechanisms of pyrimidine dimer excision in Saccharomyces cerevisiae: incision of ultraviolet-irradiated deoxyribonucleic acid in vivo. J Bacteriol. 1981 May;146(2):692–704. doi: 10.1128/jb.146.2.692-704.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Ronne H., Rothstein R. Mitotic sectored colonies: evidence of heteroduplex DNA formation during direct repeat recombination. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2696–2700. doi: 10.1073/pnas.85.8.2696. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Rothstein R. J. One-step gene disruption in yeast. Methods Enzymol. 1983;101:202–211. doi: 10.1016/0076-6879(83)01015-0. [DOI] [PubMed] [Google Scholar]
  35. Rothstein R., Helms C., Rosenberg N. Concerted deletions and inversions are caused by mitotic recombination between delta sequences in Saccharomyces cerevisiae. Mol Cell Biol. 1987 Mar;7(3):1198–1207. doi: 10.1128/mcb.7.3.1198. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Rudin N., Haber J. E. Efficient repair of HO-induced chromosomal breaks in Saccharomyces cerevisiae by recombination between flanking homologous sequences. Mol Cell Biol. 1988 Sep;8(9):3918–3928. doi: 10.1128/mcb.8.9.3918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Schiestl R. H., Igarashi S., Hastings P. J. Analysis of the mechanism for reversion of a disrupted gene. Genetics. 1988 Jun;119(2):237–247. doi: 10.1093/genetics/119.2.237. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Schiestl R. H., Prakash S. RAD1, an excision repair gene of Saccharomyces cerevisiae, is also involved in recombination. Mol Cell Biol. 1988 Sep;8(9):3619–3626. doi: 10.1128/mcb.8.9.3619. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Smith G. R. Mechanism and control of homologous recombination in Escherichia coli. Annu Rev Genet. 1987;21:179–201. doi: 10.1146/annurev.ge.21.120187.001143. [DOI] [PubMed] [Google Scholar]
  40. Stark G. R., Wahl G. M. Gene amplification. Annu Rev Biochem. 1984;53:447–491. doi: 10.1146/annurev.bi.53.070184.002311. [DOI] [PubMed] [Google Scholar]
  41. Thomas B. J., Rothstein R. Elevated recombination rates in transcriptionally active DNA. Cell. 1989 Feb 24;56(4):619–630. doi: 10.1016/0092-8674(89)90584-9. [DOI] [PubMed] [Google Scholar]
  42. Torchia T. E., Hamilton R. W., Cano C. L., Hopper J. E. Disruption of regulatory gene GAL80 in Saccharomyces cerevisiae: effects on carbon-controlled regulation of the galactose/melibiose pathway genes. Mol Cell Biol. 1984 Aug;4(8):1521–1527. doi: 10.1128/mcb.4.8.1521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Weiffenbach B., Haber J. E. Homothallic mating type switching generates lethal chromosome breaks in rad52 strains of Saccharomyces cerevisiae. Mol Cell Biol. 1981 Jun;1(6):522–534. doi: 10.1128/mcb.1.6.522. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Wilcox D. R., Prakash L. Incision and postincision steps of pyrimidine dimer removal in excision-defective mutants of Saccharomyces cerevisiae. J Bacteriol. 1981 Nov;148(2):618–623. doi: 10.1128/jb.148.2.618-623.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Willis K. K., Klein H. L. Intrachromosomal recombination in Saccharomyces cerevisiae: reciprocal exchange in an inverted repeat and associated gene conversion. Genetics. 1987 Dec;117(4):633–643. doi: 10.1093/genetics/117.4.633. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Yang E., Friedberg E. C. Molecular cloning and nucleotide sequence analysis of the Saccharomyces cerevisiae RAD1 gene. Mol Cell Biol. 1984 Oct;4(10):2161–2169. doi: 10.1128/mcb.4.10.2161. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Genetics are provided here courtesy of Oxford University Press

RESOURCES