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. 2000 Apr;154(4):1587–1596. doi: 10.1093/genetics/154.4.1587

RAD51 is required for propagation of the germinal nucleus in Tetrahymena thermophila.

T C Marsh 1, E S Cole 1, K R Stuart 1, C Campbell 1, D P Romero 1
PMCID: PMC1461009  PMID: 10747055

Abstract

RAD51, the eukaryote homolog of the Escherichia coli recA recombinase, participates in homologous recombination during mitosis, meiosis, and in the repair of double-stranded DNA breaks. The Tetrahymena thermophila RAD51 gene was recently cloned, and the in vitro activities and induction of Rad51p following DNA damage were shown to be similar to that of RAD51 from other species. This study describes the pattern of Tetrahymena RAD51 expression during both the cell cycle and conjugation. Tetrahymena RAD51 mRNA abundance is elevated during macronuclear S phase during vegetative cell growth and with both meiotic prophase and new macronuclear development during conjugation. Gene disruption of the macronuclear RAD51 locus leads to severe abnormalities during both vegetative growth and conjugation. rad51 nulls divide slowly and incur rapid deterioration of their micronuclear chromosomes. Conjugation of two rad51 nulls leads to an arrest early during prezygotic development (meiosis I). We discuss the potential usefulness of the ciliates' characteristic nuclear duality for further analyses of the potentially unique roles of Tetrahymena RAD51.

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Selected References

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  1. Adl S. M., Berger J. D. Commitment to division in ciliate cell cycles. J Eukaryot Microbiol. 1996 Mar-Apr;43(2):77–86. doi: 10.1111/j.1550-7408.1996.tb04484.x. [DOI] [PubMed] [Google Scholar]
  2. Akaboshi E., Inoue Y., Ryo H. Cloning of the cDNA and genomic DNA that correspond to the recA-like gene of Drosophila melanogaster. Jpn J Genet. 1994 Dec;69(6):663–670. doi: 10.1266/jjg.69.663. [DOI] [PubMed] [Google Scholar]
  3. Allen S. L. Cytogenetics of genomic exclusion in Tetrahymena. Genetics. 1967 Apr;55(4):797–822. doi: 10.1093/genetics/55.4.797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Allis C. D., Colavito-Shepanski M., Gorovsky M. A. Scheduled and unscheduled DNA synthesis during development in conjugating Tetrahymena. Dev Biol. 1987 Dec;124(2):469–480. doi: 10.1016/0012-1606(87)90500-8. [DOI] [PubMed] [Google Scholar]
  5. Basile G., Aker M., Mortimer R. K. Nucleotide sequence and transcriptional regulation of the yeast recombinational repair gene RAD51. Mol Cell Biol. 1992 Jul;12(7):3235–3246. doi: 10.1128/mcb.12.7.3235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Baumann P., Benson F. E., West S. C. Human Rad51 protein promotes ATP-dependent homologous pairing and strand transfer reactions in vitro. Cell. 1996 Nov 15;87(4):757–766. doi: 10.1016/s0092-8674(00)81394-x. [DOI] [PubMed] [Google Scholar]
  7. Bodenbender J., Prohaska A., Jauker F., Hipke H., Cleffmann G. DNA elimination and its relation to quantities in the macronucleus of Tetrahymena. Dev Genet. 1992;13(2):103–110. doi: 10.1002/dvg.1020130203. [DOI] [PubMed] [Google Scholar]
  8. Cassidy-Hanley D., Bowen J., Lee J. H., Cole E., VerPlank L. A., Gaertig J., Gorovsky M. A., Bruns P. J. Germline and somatic transformation of mating Tetrahymena thermophila by particle bombardment. Genetics. 1997 May;146(1):135–147. doi: 10.1093/genetics/146.1.135. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cole E. S., Cassidy-Hanley D., Hemish J., Tuan J., Bruns P. J. A mutational analysis of conjugation in Tetrahymena thermophila. 1. Phenotypes affecting early development: meiosis to nuclear selection. Dev Biol. 1997 Sep 15;189(2):215–232. doi: 10.1006/dbio.1997.8648. [DOI] [PubMed] [Google Scholar]
  10. Cole E. S., Soelter T. A. A mutational analysis of conjugation in Tetrahymena thermophila. 2. Phenotypes affecting middle and late development: third prezygotic nuclear division, pronuclear exchange, pronuclear fusion, and postzygotic development. Dev Biol. 1997 Sep 15;189(2):233–245. doi: 10.1006/dbio.1997.8649. [DOI] [PubMed] [Google Scholar]
  11. Coyne R. S., Chalker D. L., Yao M. C. Genome downsizing during ciliate development: nuclear division of labor through chromosome restructuring. Annu Rev Genet. 1996;30:557–578. doi: 10.1146/annurev.genet.30.1.557. [DOI] [PubMed] [Google Scholar]
  12. Deak J. C., Doerder F. P. High frequency intragenic recombination during macronuclear development in Tetrahymena thermophila restores the wild-type SerH1 gene. Genetics. 1998 Mar;148(3):1109–1115. doi: 10.1093/genetics/148.3.1109. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Doerder F. P., Debault L. E. Cytofluorimetric analysis of nuclear DNA during meiosis, fertilization and macronuclear development in the ciliate Tetrahymena pyriformis, syngen 1. J Cell Sci. 1975 May;17(3):471–493. doi: 10.1242/jcs.17.3.471. [DOI] [PubMed] [Google Scholar]
  14. Dresser M. E., Ewing D. J., Conrad M. N., Dominguez A. M., Barstead R., Jiang H., Kodadek T. DMC1 functions in a Saccharomyces cerevisiae meiotic pathway that is largely independent of the RAD51 pathway. Genetics. 1997 Oct;147(2):533–544. doi: 10.1093/genetics/147.2.533. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gaertig J., Thatcher T. H., Gu L., Gorovsky M. A. Electroporation-mediated replacement of a positively and negatively selectable beta-tubulin gene in Tetrahymena thermophila. Proc Natl Acad Sci U S A. 1994 May 10;91(10):4549–4553. doi: 10.1073/pnas.91.10.4549. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Hai B., Gorovsky M. A. Germ-line knockout heterokaryons of an essential alpha-tubulin gene enable high-frequency gene replacement and a test of gene transfer from somatic to germ-line nuclei in Tetrahymena thermophila. Proc Natl Acad Sci U S A. 1997 Feb 18;94(4):1310–1315. doi: 10.1073/pnas.94.4.1310. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kirk K. E., Harmon B. P., Reichardt I. K., Sedat J. W., Blackburn E. H. Block in anaphase chromosome separation caused by a telomerase template mutation. Science. 1997 Mar 7;275(5305):1478–1481. doi: 10.1126/science.275.5305.1478. [DOI] [PubMed] [Google Scholar]
  18. Lee S., Wisniewski J. C., Dentler W. L., Asai D. J. Gene knockouts reveal separate functions for two cytoplasmic dyneins in Tetrahymena thermophila. Mol Biol Cell. 1999 Mar;10(3):771–784. doi: 10.1091/mbc.10.3.771. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. McCormick-Graham M., Romero D. P. A single telomerase RNA is sufficient for the synthesis of variable telomeric DNA repeats in ciliates of the genus Paramecium. Mol Cell Biol. 1996 Apr;16(4):1871–1879. doi: 10.1128/mcb.16.4.1871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Muris D. F., Vreeken K., Carr A. M., Broughton B. C., Lehmann A. R., Lohman P. H., Pastink A. Cloning the RAD51 homologue of Schizosaccharomyces pombe. Nucleic Acids Res. 1993 Sep 25;21(19):4586–4591. doi: 10.1093/nar/21.19.4586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. 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]
  22. Orias E., Flacks M. Macronuclear genetics of Tetrahymena. I. Random distribution of macronuclear genecopies in T. pyriformis, syngen 1. Genetics. 1975 Feb;79(2):187–206. doi: 10.1093/genetics/79.2.187. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Prescott D. M. The DNA of ciliated protozoa. Microbiol Rev. 1994 Jun;58(2):233–267. doi: 10.1128/mr.58.2.233-267.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rinaldo C., Ederle S., Rocco V., La Volpe A. The Caenorhabditis elegans RAD51 homolog is transcribed into two alternative mRNAs potentially encoding proteins of different sizes. Mol Gen Genet. 1998 Nov;260(2-3):289–294. doi: 10.1007/s004380050897. [DOI] [PubMed] [Google Scholar]
  25. Scully R., Chen J., Plug A., Xiao Y., Weaver D., Feunteun J., Ashley T., Livingston D. M. Association of BRCA1 with Rad51 in mitotic and meiotic cells. Cell. 1997 Jan 24;88(2):265–275. doi: 10.1016/s0092-8674(00)81847-4. [DOI] [PubMed] [Google Scholar]
  26. Shinohara A., Ogawa H., Matsuda Y., Ushio N., Ikeo K., Ogawa T. Cloning of human, mouse and fission yeast recombination genes homologous to RAD51 and recA. Nat Genet. 1993 Jul;4(3):239–243. doi: 10.1038/ng0793-239. [DOI] [PubMed] [Google Scholar]
  27. 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]
  28. Sonoda E., Sasaki M. S., Buerstedde J. M., Bezzubova O., Shinohara A., Ogawa H., Takata M., Yamaguchi-Iwai Y., Takeda S. Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death. EMBO J. 1998 Jan 15;17(2):598–608. doi: 10.1093/emboj/17.2.598. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stürzbecher H. W., Donzelmann B., Henning W., Knippschild U., Buchhop S. p53 is linked directly to homologous recombination processes via RAD51/RecA protein interaction. EMBO J. 1996 Apr 15;15(8):1992–2002. [PMC free article] [PubMed] [Google Scholar]
  30. 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]
  31. Taki T., Ohnishi T., Yamamoto A., Hiraga S., Arita N., Izumoto S., Hayakawa T., Morita T. Antisense inhibition of the RAD51 enhances radiosensitivity. Biochem Biophys Res Commun. 1996 Jun 14;223(2):434–438. doi: 10.1006/bbrc.1996.0911. [DOI] [PubMed] [Google Scholar]
  32. Tsuzuki T., Fujii Y., Sakumi K., Tominaga Y., Nakao K., Sekiguchi M., Matsushiro A., Yoshimura Y., MoritaT Targeted disruption of the Rad51 gene leads to lethality in embryonic mice. Proc Natl Acad Sci U S A. 1996 Jun 25;93(13):6236–6240. doi: 10.1073/pnas.93.13.6236. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vispé S., Cazaux C., Lesca C., Defais M. Overexpression of Rad51 protein stimulates homologous recombination and increases resistance of mammalian cells to ionizing radiation. Nucleic Acids Res. 1998 Jun 15;26(12):2859–2864. doi: 10.1093/nar/26.12.2859. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wei Y., Yu L., Bowen J., Gorovsky M. A., Allis C. D. Phosphorylation of histone H3 is required for proper chromosome condensation and segregation. Cell. 1999 Apr 2;97(1):99–109. doi: 10.1016/s0092-8674(00)80718-7. [DOI] [PubMed] [Google Scholar]
  35. Wu M., Allis C. D., Gorovsky M. A. Cell-cycle regulation as a mechanism for targeting proteins to specific DNA sequences in Tetrahymena thermophila. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2205–2209. doi: 10.1073/pnas.85.7.2205. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Xia S. J., Shammas M. A., Shmookler Reis R. J. Elevated recombination in immortal human cells is mediated by HsRAD51 recombinase. Mol Cell Biol. 1997 Dec;17(12):7151–7158. doi: 10.1128/mcb.17.12.7151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Xu L., Weiner B. M., Kleckner N. Meiotic cells monitor the status of the interhomolog recombination complex. Genes Dev. 1997 Jan 1;11(1):106–118. doi: 10.1101/gad.11.1.106. [DOI] [PubMed] [Google Scholar]
  38. Yuan Z. M., Huang Y., Ishiko T., Nakada S., Utsugisawa T., Kharbanda S., Wang R., Sung P., Shinohara A., Weichselbaum R. Regulation of Rad51 function by c-Abl in response to DNA damage. J Biol Chem. 1998 Feb 13;273(7):3799–3802. doi: 10.1074/jbc.273.7.3799. [DOI] [PubMed] [Google Scholar]

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