Abstract
Double-strand breaks in DNA are known to promote recombination in Saccharomyces cerevisiae. Yeast mating type switching, which is a highly efficient gene conversion event, is apparently initiated by a site-specific double-strand break. The 2 micrograms circle site-specific recombinase, FLP, has been shown to make double-strand breaks in its substrate DNA. By using a hybrid 2 micrograms circle::Tn5 plasmid, a portion of which resembles, in its DNA organization, the active (MAT) and the silent (HML) yeast mating type loci, it is shown that FLP mediates a conversion event analogous to mating type switching. Whereas the FLP site-specific recombination is not dependent on the RAD52 gene product, the FLP-induced conversion is abolished in a rad52 background. The FLP-promoted conversion in vivo can be faithfully reproduced by making a double-stranded gap in vitro in the vicinity of the FLP site and allowing the gap to be repaired in vivo.
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- Andrews B. J., Proteau G. A., Beatty L. G., Sadowski P. D. The FLP recombinase of the 2 micron circle DNA of yeast: interaction with its target sequences. Cell. 1985 Apr;40(4):795–803. doi: 10.1016/0092-8674(85)90339-3. [DOI] [PubMed] [Google Scholar]
- Beggs J. D. Transformation of yeast by a replicating hybrid plasmid. Nature. 1978 Sep 14;275(5676):104–109. doi: 10.1038/275104a0. [DOI] [PubMed] [Google Scholar]
- Broach J. R., Guarascio V. R., Jayaram M. Recombination within the yeast plasmid 2mu circle is site-specific. Cell. 1982 May;29(1):227–234. doi: 10.1016/0092-8674(82)90107-6. [DOI] [PubMed] [Google Scholar]
- Cryer D. R., Eccleshall R., Marmur J. Isolation of yeast DNA. Methods Cell Biol. 1975;12:39–44. doi: 10.1016/s0091-679x(08)60950-4. [DOI] [PubMed] [Google Scholar]
- Gronostajski R. M., Sadowski P. D. Determination of DNA sequences essential for FLP-mediated recombination by a novel method. J Biol Chem. 1985 Oct 5;260(22):12320–12327. [PubMed] [Google Scholar]
- HAWTHORNE D. C. A DELETION IN YEAST AND ITS BEARING ON THE STRUCTURE OF THE MATING TYPE LOCUS. Genetics. 1963 Dec;48:1727–1729. doi: 10.1093/genetics/48.12.1727. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hartley J. L., Donelson J. E. Nucleotide sequence of the yeast plasmid. Nature. 1980 Aug 28;286(5776):860–865. doi: 10.1038/286860a0. [DOI] [PubMed] [Google Scholar]
- 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]
- Jayaram M., Broach J. R. Yeast plasmid 2-micron circle promotes recombination within bacterial transposon Tn5. Proc Natl Acad Sci U S A. 1983 Dec;80(23):7264–7268. doi: 10.1073/pnas.80.23.7264. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jayaram M., Li Y. Y., Broach J. R. The yeast plasmid 2mu circle encodes components required for its high copy propagation. Cell. 1983 Aug;34(1):95–104. doi: 10.1016/0092-8674(83)90139-3. [DOI] [PubMed] [Google Scholar]
- Jayaram M. Two-micrometer circle site-specific recombination: the minimal substrate and the possible role of flanking sequences. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5875–5879. doi: 10.1073/pnas.82.17.5875. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kikuchi Y. Yeast plasmid requires a cis-acting locus and two plasmid proteins for its stable maintenance. Cell. 1983 Dec;35(2 Pt 1):487–493. doi: 10.1016/0092-8674(83)90182-4. [DOI] [PubMed] [Google Scholar]
- Klar A. J., Strathern J. N. Resolution of recombination intermediates generated during yeast mating type switching. 1984 Aug 30-Sep 5Nature. 310(5980):744–748. doi: 10.1038/310744a0. [DOI] [PubMed] [Google Scholar]
- Klein H. L. Lack of association between intrachromosomal gene conversion and reciprocal exchange. 1984 Aug 30-Sep 5Nature. 310(5980):748–753. doi: 10.1038/310748a0. [DOI] [PubMed] [Google Scholar]
- Klein H. L., Petes T. D. Intrachromosomal gene conversion in yeast. Nature. 1981 Jan 15;289(5794):144–148. doi: 10.1038/289144a0. [DOI] [PubMed] [Google Scholar]
- Kostriken R., Strathern J. N., Klar A. J., Hicks J. B., Heffron F. A site-specific endonuclease essential for mating-type switching in Saccharomyces cerevisiae. Cell. 1983 Nov;35(1):167–174. doi: 10.1016/0092-8674(83)90219-2. [DOI] [PubMed] [Google Scholar]
- 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]
- Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
- Nasmyth K. Molecular analysis of a cell lineage. Nature. 1983 Apr 21;302(5910):670–676. doi: 10.1038/302670a0. [DOI] [PubMed] [Google Scholar]
- 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]
- Orr-Weaver T. L., Szostak J. W. Yeast recombination: the association between double-strand gap repair and crossing-over. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4417–4421. doi: 10.1073/pnas.80.14.4417. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Prakash S., Prakash L., Burke W., Montelone B. A. Effects of the RAD52 Gene on Recombination in SACCHAROMYCES CEREVISIAE. Genetics. 1980 Jan;94(1):31–50. doi: 10.1093/genetics/94.1.31. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Ruvkun G. B., Ausubel F. M. A general method for site-directed mutagenesis in prokaryotes. Nature. 1981 Jan 1;289(5793):85–88. doi: 10.1038/289085a0. [DOI] [PubMed] [Google Scholar]
- Senecoff J. F., Bruckner R. C., Cox M. M. The FLP recombinase of the yeast 2-micron plasmid: characterization of its recombination site. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7270–7274. doi: 10.1073/pnas.82.21.7270. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sigurdson D. C., Gaarder M. E., Livingston D. M. Characterization of the transmission during cytoductant formation of the 2 micrometers DNA plasmid from Saccharomyces. Mol Gen Genet. 1981;183(1):59–65. doi: 10.1007/BF00270139. [DOI] [PubMed] [Google Scholar]
- Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
- Strathern J. N., Klar A. J., Hicks J. B., Abraham J. A., Ivy J. M., Nasmyth K. A., McGill C. Homothallic switching of yeast mating type cassettes is initiated by a double-stranded cut in the MAT locus. Cell. 1982 Nov;31(1):183–192. doi: 10.1016/0092-8674(82)90418-4. [DOI] [PubMed] [Google Scholar]
- Strathern J. N., Newlon C. S., Herskowitz I., Hicks J. B. Isolation of a circular derivative of yeast chromosome III: implications for the mechanism of mating type interconversion. Cell. 1979 Oct;18(2):309–319. doi: 10.1016/0092-8674(79)90050-3. [DOI] [PubMed] [Google Scholar]
- Szostak J. W., Orr-Weaver T. L., Rothstein R. J., Stahl F. W. The double-strand-break repair model for recombination. Cell. 1983 May;33(1):25–35. doi: 10.1016/0092-8674(83)90331-8. [DOI] [PubMed] [Google Scholar]
- 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]