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. 1994 Dec;14(12):8037–8050. doi: 10.1128/mcb.14.12.8037

Mutations in the Saccharomyces cerevisiae CDC1 gene affect double-strand-break-induced intrachromosomal recombination.

J Halbrook 1, M F Hoekstra 1
PMCID: PMC359342  PMID: 7969142

Abstract

To isolate Saccharomyces cerevisiae mutants defective in recombinational DNA repair, we constructed a strain that contains duplicated ura3 alleles that flank LEU2 and ADE5 genes at the ura3 locus on chromosome V. When a HO endonuclease cleavage site is located within one of the ura3 alleles, Ura+ recombination is increased over 100-fold in wild-type strains following HO induction from the GAL1, 10 promoter. This strain was used to screen for mutants that exhibited reduced levels of HO-induced intrachromosomal recombination without significantly affecting the spontaneous frequency of Ura+ recombination. One of the mutations isolated through this screen was found to affect the essential gene CDC1. This mutation, cdc1-100, completely eliminated HO-induced Ura+ recombination yet maintained both spontaneous preinduced recombination levels and cell viability, cdc1-100 mutants were moderately sensitive to killing by methyl methanesulfonate and gamma irradiation. The effect of the cdc1-100 mutation on recombinational double-strand break repair indicates that a recombinationally silent mechanism other than sister chromatid exchange was responsible for the efficient repair of DNA double-strand breaks.

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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. Alani E., Cao L., Kleckner N. A method for gene disruption that allows repeated use of URA3 selection in the construction of multiply disrupted yeast strains. Genetics. 1987 Aug;116(4):541–545. doi: 10.1534/genetics.112.541.test. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Brunborg G., Resnick M. A., Williamson D. H. Cell-cycle-specific repair of DNA double strand breaks in Saccharomyces cerevisiae. Radiat Res. 1980 Jun;82(3):547–558. [PubMed] [Google Scholar]
  6. Brunborg G., Williamson D. H. The relevance of the nuclear division cycle to radiosensitivity in yeast. Mol Gen Genet. 1978 Jul 4;162(3):277–286. doi: 10.1007/BF00268853. [DOI] [PubMed] [Google Scholar]
  7. Byers B., Goetsch L. Duplication of spindle plaques and integration of the yeast cell cycle. Cold Spring Harb Symp Quant Biol. 1974;38:123–131. doi: 10.1101/sqb.1974.038.01.016. [DOI] [PubMed] [Google Scholar]
  8. Chen J., Derfler B., Samson L. Saccharomyces cerevisiae 3-methyladenine DNA glycosylase has homology to the AlkA glycosylase of E. coli and is induced in response to DNA alkylation damage. EMBO J. 1990 Dec;9(13):4569–4575. doi: 10.1002/j.1460-2075.1990.tb07910.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chlebowicz E., Jachymczyk W. J. Repair of MMS-induced DNA double-strand breaks in haploid cells of Saccharomyces cerevisiae, which requires the presence of a duplicate genome. Mol Gen Genet. 1979 Jan 2;167(3):279–286. doi: 10.1007/BF00267420. [DOI] [PubMed] [Google Scholar]
  10. Connolly B., White C. I., Haber J. E. Physical monitoring of mating type switching in Saccharomyces cerevisiae. Mol Cell Biol. 1988 Jun;8(6):2342–2349. doi: 10.1128/mcb.8.6.2342. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Dhillon N., Hoekstra M. F. Characterization of two protein kinases from Schizosaccharomyces pombe involved in the regulation of DNA repair. EMBO J. 1994 Jun 15;13(12):2777–2788. doi: 10.1002/j.1460-2075.1994.tb06571.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Elble R. A simple and efficient procedure for transformation of yeasts. Biotechniques. 1992 Jul;13(1):18–20. [PubMed] [Google Scholar]
  13. 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]
  14. 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]
  15. Gietz R. D., Sugino A. New yeast-Escherichia coli shuttle vectors constructed with in vitro mutagenized yeast genes lacking six-base pair restriction sites. Gene. 1988 Dec 30;74(2):527–534. doi: 10.1016/0378-1119(88)90185-0. [DOI] [PubMed] [Google Scholar]
  16. Hartwell L. H., Culotti J., Pringle J. R., Reid B. J. Genetic control of the cell division cycle in yeast. Science. 1974 Jan 11;183(4120):46–51. doi: 10.1126/science.183.4120.46. [DOI] [PubMed] [Google Scholar]
  17. Hartwell L. H., Culotti J., Reid B. Genetic control of the cell-division cycle in yeast. I. Detection of mutants. Proc Natl Acad Sci U S A. 1970 Jun;66(2):352–359. doi: 10.1073/pnas.66.2.352. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hartwell L. H. Genetic control of the cell division cycle in yeast. IV. Genes controlling bud emergence and cytokinesis. Exp Cell Res. 1971 Dec;69(2):265–276. doi: 10.1016/0014-4827(71)90223-0. [DOI] [PubMed] [Google Scholar]
  19. Hartwell L. H., Mortimer R. K., Culotti J., Culotti M. Genetic Control of the Cell Division Cycle in Yeast: V. Genetic Analysis of cdc Mutants. Genetics. 1973 Jun;74(2):267–286. doi: 10.1093/genetics/74.2.267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Henner W. D., Grunberg S. M., Haseltine W. A. Sites and structure of gamma radiation-induced DNA strand breaks. J Biol Chem. 1982 Oct 10;257(19):11750–11754. [PubMed] [Google Scholar]
  21. Ho K. S., Mortimer R. K. Induction of dominant lethality by x-rays in radiosensitive strain of yeast. Mutat Res. 1973 Oct;20(1):45–51. doi: 10.1016/0027-5107(73)90096-1. [DOI] [PubMed] [Google Scholar]
  22. Ho K. S., Mortimer R. K. Two mutations which confer temperature-sensitive radiation sensitivity in the yeast Saccharomyces cerevisiae. Mutat Res. 1975 Dec;33(2-3):157–164. doi: 10.1016/0027-5107(75)90190-6. [DOI] [PubMed] [Google Scholar]
  23. Hovland P., Flick J., Johnston M., Sclafani R. A. Galactose as a gratuitous inducer of GAL gene expression in yeasts growing on glucose. Gene. 1989 Nov 15;83(1):57–64. doi: 10.1016/0378-1119(89)90403-4. [DOI] [PubMed] [Google Scholar]
  24. Huisman O., Raymond W., Froehlich K. U., Errada P., Kleckner N., Botstein D., Hoyt M. A. A Tn10-lacZ-kanR-URA3 gene fusion transposon for insertion mutagenesis and fusion analysis of yeast and bacterial genes. Genetics. 1987 Jun;116(2):191–199. doi: 10.1093/genetics/116.2.191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Jensen R. E., Herskowitz I. Directionality and regulation of cassette substitution in yeast. Cold Spring Harb Symp Quant Biol. 1984;49:97–104. doi: 10.1101/sqb.1984.049.01.013. [DOI] [PubMed] [Google Scholar]
  26. Kadyk L. C., Hartwell L. H. Sister chromatids are preferred over homologs as substrates for recombinational repair in Saccharomyces cerevisiae. Genetics. 1992 Oct;132(2):387–402. doi: 10.1093/genetics/132.2.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Kostriken R., Heffron F. The product of the HO gene is a nuclease: purification and characterization of the enzyme. Cold Spring Harb Symp Quant Biol. 1984;49:89–96. doi: 10.1101/sqb.1984.049.01.012. [DOI] [PubMed] [Google Scholar]
  28. Kramer K. M., Brock J. A., Bloom K., Moore J. K., Haber J. E. Two different types of double-strand breaks in Saccharomyces cerevisiae are repaired by similar RAD52-independent, nonhomologous recombination events. Mol Cell Biol. 1994 Feb;14(2):1293–1301. doi: 10.1128/mcb.14.2.1293. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. 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]
  30. Lin Y. H., Keil R. L. Mutations affecting RNA polymerase I-stimulated exchange and rDNA recombination in yeast. Genetics. 1991 Jan;127(1):31–38. doi: 10.1093/genetics/127.1.31. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Luchnik A. N., Glaser V. M., Shestakov S. V. Repair of DNA double-strand breaks requires two homologous DNA duplexes. Mol Biol Rep. 1977 Dec;3(6):437–442. doi: 10.1007/BF00808385. [DOI] [PubMed] [Google Scholar]
  32. 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]
  33. Matsumoto K., Yoshimatsu T., Oshima Y. Recessive mutations conferring resistance to carbon catabolite repression of galactokinase synthesis in Saccharomyces cerevisiae. J Bacteriol. 1983 Mar;153(3):1405–1414. doi: 10.1128/jb.153.3.1405-1414.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. 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]
  35. Ozenberger B. A., Roeder G. S. A unique pathway of double-strand break repair operates in tandemly repeated genes. Mol Cell Biol. 1991 Mar;11(3):1222–1231. doi: 10.1128/mcb.11.3.1222. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Ray A., Siddiqi I., Kolodkin A. L., Stahl F. W. Intra-chromosomal gene conversion induced by a DNA double-strand break in Saccharomyces cerevisiae. J Mol Biol. 1988 May 20;201(2):247–260. doi: 10.1016/0022-2836(88)90136-2. [DOI] [PubMed] [Google Scholar]
  37. Repine J. E., Pfenninger O. W., Talmage D. W., Berger E. M., Pettijohn D. E. Dimethyl sulfoxide prevents DNA nicking mediated by ionizing radiation or iron/hydrogen peroxide-generated hydroxyl radical. Proc Natl Acad Sci U S A. 1981 Feb;78(2):1001–1003. doi: 10.1073/pnas.78.2.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. 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]
  39. 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]
  40. 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]
  41. Rose M. D., Broach J. R. Cloning genes by complementation in yeast. Methods Enzymol. 1991;194:195–230. doi: 10.1016/0076-6879(91)94017-7. [DOI] [PubMed] [Google Scholar]
  42. 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]
  43. 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]
  44. Sikorski R. S., Hieter P. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics. 1989 May;122(1):19–27. doi: 10.1093/genetics/122.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Simchen G. Are mitotic functions required in meiosis? Genetics. 1974 Apr;76(4):745–753. doi: 10.1093/genetics/76.4.745. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Sugawara N., Haber J. E. Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation. Mol Cell Biol. 1992 Feb;12(2):563–575. doi: 10.1128/mcb.12.2.563. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. 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]
  48. 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]
  49. Weinert T. A., Hartwell L. H. The RAD9 gene controls the cell cycle response to DNA damage in Saccharomyces cerevisiae. Science. 1988 Jul 15;241(4863):317–322. doi: 10.1126/science.3291120. [DOI] [PubMed] [Google Scholar]
  50. White C. I., Haber J. E. Intermediates of recombination during mating type switching in Saccharomyces cerevisiae. EMBO J. 1990 Mar;9(3):663–673. doi: 10.1002/j.1460-2075.1990.tb08158.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Xiao W., Derfler B., Chen J., Samson L. Primary sequence and biological functions of a Saccharomyces cerevisiae O6-methylguanine/O4-methylthymine DNA repair methyltransferase gene. EMBO J. 1991 Aug;10(8):2179–2186. doi: 10.1002/j.1460-2075.1991.tb07753.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Zehfus B. R., McWilliams A. D., Lin Y. H., Hoekstra M. F., Keil R. L. Genetic control of RNA polymerase I-stimulated recombination in yeast. Genetics. 1990 Sep;126(1):41–52. doi: 10.1093/genetics/126.1.41. [DOI] [PMC free article] [PubMed] [Google Scholar]

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