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. 1991 Dec;129(4):1053–1060.

Extrachromosomal Recombination Is Deranged in the Rec2 Mutant of Ustilago Maydis

S Fotheringham 1, W K Holloman 1
PMCID: PMC1204770  PMID: 1783291

Abstract

Transformation of a leu1 auxotroph of Ustilago maydis to prototrophy with an autonomously replicating plasmid containing the selectable LEU1 gene was found to be efficient regardless of whether the transforming DNA was circular or linear. When pairs of autonomously replicating plasmids bearing noncomplementing leu1 alleles were used to cotransform strains deleted entirely for the genomic copy of the LEU1 gene, Leu(+) transformants were observed to arise by extrachromosomal recombination. The frequency of recombination increased severalfold when one plasmid of the pair was made linear by cleavage at one end of the leu1 gene, but increased 10-100-fold when both plasmids were first made linear. The increase in recombination noted in wild-type and rec1 strains was not apparent in the rec2 mutant unless the members of the pair of plasmids were cut at opposite ends of the leu1 gene to yield linear molecules offset in only one of the two possible configurations. Use of a pair of plasmid substrates designed to measure nonreciprocal and multiple exchange events revealed only a minor fraction of the total events arise through these modes, and further that no stimulation occurred when the plasmid DNA was linear. It is unlikely that the defect in rec2 lies in a mismatch correction step since a high yield of Leu(+) recombinants was obtained from the rec2 mutant when it was transformed with heteroduplex DNA constructed from plasmids with the two different leu1 alleles.

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Selected References

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

  1. Amundsen S. K., Neiman A. M., Thibodeaux S. M., Smith G. R. Genetic dissection of the biochemical activities of RecBCD enzyme. Genetics. 1990 Sep;126(1):25–40. doi: 10.1093/genetics/126.1.25. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bauchwitz R., Holloman W. K. Isolation of the REC2 gene controlling recombination in Ustilago maydis. Gene. 1990 Dec 15;96(2):285–288. doi: 10.1016/0378-1119(90)90265-s. [DOI] [PubMed] [Google Scholar]
  3. Bernstein H., Hopf F. A., Michod R. E. The molecular basis of the evolution of sex. Adv Genet. 1987;24:323–370. doi: 10.1016/s0065-2660(08)60012-7. [DOI] [PubMed] [Google Scholar]
  4. Brenner D. A., Smigocki A. C., Camerini-Otero R. D. Double-strand gap repair results in homologous recombination in mouse L cells. Proc Natl Acad Sci U S A. 1986 Mar;83(6):1762–1766. doi: 10.1073/pnas.83.6.1762. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Conley E. C., Saunders J. R. Recombination-dependent recircularization of linearized pBR322 plasmid DNA following transformation of Escherichia coli. Mol Gen Genet. 1984;194(1-2):211–218. doi: 10.1007/BF00383519. [DOI] [PubMed] [Google Scholar]
  6. Embretson J. E., Livingston D. M. A plasmid model to study genetic recombination in yeast. Gene. 1984 Sep;29(3):293–302. doi: 10.1016/0378-1119(84)90058-1. [DOI] [PubMed] [Google Scholar]
  7. Folger K. R., Thomas K., Capecchi M. R. Nonreciprocal exchanges of information between DNA duplexes coinjected into mammalian cell nuclei. Mol Cell Biol. 1985 Jan;5(1):59–69. doi: 10.1128/mcb.5.1.59. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fotheringham S., Holloman W. K. Cloning and disruption of Ustilago maydis genes. Mol Cell Biol. 1989 Sep;9(9):4052–4055. doi: 10.1128/mcb.9.9.4052. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fotheringham S., Holloman W. K. Pathways of transformation in Ustilago maydis determined by DNA conformation. Genetics. 1990 Apr;124(4):833–843. doi: 10.1093/genetics/124.4.833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Holliday R. Altered recombination frequencies in radiation sensitivie strains of Ustilago. Mutat Res. 1967 May-Jun;4(3):275–288. doi: 10.1016/0027-5107(67)90022-x. [DOI] [PubMed] [Google Scholar]
  11. Holliday R. Further evidence for an inducible recombination repair system in Ustilago maydis. Mutat Res. 1975 Jul;29(1):149–153. doi: 10.1016/0027-5107(75)90029-9. [DOI] [PubMed] [Google Scholar]
  12. Holliday R., Halliwell R. E., Evans M. W., Rowell V. Genetic characterization of rec-1, a mutant of Ustilago maydis defective in repair and recombination. Genet Res. 1976 Jun;27(3):413–453. doi: 10.1017/s0016672300016621. [DOI] [PubMed] [Google Scholar]
  13. Holloman W. K., Rowe T. C., Rusche J. R. Studies on nuclease alpha from Ustilago maydis. J Biol Chem. 1981 May 25;256(10):5087–5094. [PubMed] [Google Scholar]
  14. 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]
  15. Leaper S., Resnick M. A., Holliday R. Repair of double-strand breaks and lethal damage in DNA of Ustilago maydis. Genet Res. 1980 Jun;35(3):291–307. doi: 10.1017/s0016672300014154. [DOI] [PubMed] [Google Scholar]
  16. Lin F. L., Sperle K., Sternberg N. Intermolecular recombination between DNAs introduced into mouse L cells is mediated by a nonconservative pathway that leads to crossover products. Mol Cell Biol. 1990 Jan;10(1):103–112. doi: 10.1128/mcb.10.1.103. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Lin F. L., Sperle K., Sternberg N. Model for homologous recombination during transfer of DNA into mouse L cells: role for DNA ends in the recombination process. Mol Cell Biol. 1984 Jun;4(6):1020–1034. doi: 10.1128/mcb.4.6.1020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Maryon E., Carroll D. Characterization of recombination intermediates from DNA injected into Xenopus laevis oocytes: evidence for a nonconservative mechanism of homologous recombination. Mol Cell Biol. 1991 Jun;11(6):3278–3287. doi: 10.1128/mcb.11.6.3278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. 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]
  21. 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]
  22. 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]
  23. Rusche J. R., Rowe T. C., Holloman W. K. Purification and characterization of nuclease beta from Ustilago maydis. J Biol Chem. 1980 Oct 10;255(19):9117–9123. [PubMed] [Google Scholar]
  24. Seidman M. M. Intermolecular homologous recombination between transfected sequences in mammalian cells is primarily nonconservative. Mol Cell Biol. 1987 Oct;7(10):3561–3565. doi: 10.1128/mcb.7.10.3561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Sun H., Treco D., Szostak J. W. Extensive 3'-overhanging, single-stranded DNA associated with the meiosis-specific double-strand breaks at the ARG4 recombination initiation site. Cell. 1991 Mar 22;64(6):1155–1161. doi: 10.1016/0092-8674(91)90270-9. [DOI] [PubMed] [Google Scholar]
  26. Symington L. S., Morrison P. T., Kolodner R. Genetic recombination catalyzed by cell-free extracts of Saccharomyces cerevisiae. Cold Spring Harb Symp Quant Biol. 1984;49:805–814. doi: 10.1101/sqb.1984.049.01.091. [DOI] [PubMed] [Google Scholar]
  27. 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]
  28. Tsukuda T., Bauchwitz R., Holloman W. K. Isolation of the REC1 gene controlling recombination in Ustilago maydis. Gene. 1989 Dec 28;85(2):335–341. doi: 10.1016/0378-1119(89)90426-5. [DOI] [PubMed] [Google Scholar]
  29. Tsukuda T., Carleton S., Fotheringham S., Holloman W. K. Isolation and characterization of an autonomously replicating sequence from Ustilago maydis. Mol Cell Biol. 1988 Sep;8(9):3703–3709. doi: 10.1128/mcb.8.9.3703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wake C. T., Gudewicz T., Porter T., White A., Wilson J. H. How damaged is the biologically active subpopulation of transfected DNA? Mol Cell Biol. 1984 Mar;4(3):387–398. doi: 10.1128/mcb.4.3.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wake C. T., Vernaleone F., Wilson J. H. Topological requirements for homologous recombination among DNA molecules transfected into mammalian cells. Mol Cell Biol. 1985 Aug;5(8):2080–2089. doi: 10.1128/mcb.5.8.2080. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wang J., Holden D. W., Leong S. A. Gene transfer system for the phytopathogenic fungus Ustilago maydis. Proc Natl Acad Sci U S A. 1988 Feb;85(3):865–869. doi: 10.1073/pnas.85.3.865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. White C. I., Sedgwick S. G. The use of plasmid DNA to probe DNA repair functions in the yeast Saccharomyces cerevisiae. Mol Gen Genet. 1985;201(1):99–106. doi: 10.1007/BF00397993. [DOI] [PubMed] [Google Scholar]
  34. Yarnall M., Rowe T. C., Holloman W. K. Purification and properties of nuclease gamma from Ustilago maydis. J Biol Chem. 1984 Mar 10;259(5):3026–3032. [PubMed] [Google Scholar]

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