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Genetics. Sep 2001; 159(1): 91–105.
PMCID: PMC1461803

Multiple interactions among the components of the recombinational DNA repair system in Schizosaccharomyces pombe.


Schizosaccharomyces pombe Rhp55 and Rhp57 are RecA-like proteins involved in double-strand break (DSB) repair. Here we demonstrate that Rhp55 and Rhp57 proteins strongly interact in vivo, similar to Saccharomyces cerevisiae Rad55p and Rad57p. Mutations in the conserved ATP-binding/hydrolysis folds of both the Rhp55 and Rhp57 proteins impaired their function in DNA repair but not in cell proliferation. However, when combined, ATPase fold mutations in Rhp55p and Rhp57p resulted in severe defects of both functions, characteristic of the deletion mutants. Yeast two-hybrid analysis also revealed other multiple in vivo interactions among S. pombe proteins involved in recombinational DNA repair. Similar to S. cerevisiae Rad51p-Rad54p, S. pombe Rhp51p and Rhp54p were found to interact. Both putative Rad52 homologs in S. pombe, Rad22p and Rti1p, were found to interact with the C-terminal region of Rhp51 protein. Moreover, Rad22p and Rti1p exhibited mutual, as well as self-, interactions. In contrast to the S. cerevisiae interacting pair Rad51p-Rad55p, S. pombe Rhp51 protein strongly interacted with Rhp57 but not with Rhp55 protein. In addition, the Rti1 and Rad22 proteins were found to form a complex with the large subunit of S. pombe RPA. Our data provide compelling evidence that most, but not all, of the protein-protein interactions found in S. cerevisiae DSB repair are evolutionarily conserved.

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Selected References

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  • Aihara H, Ito Y, Kurumizaka H, Yokoyama S, Shibata T. The N-terminal domain of the human Rad51 protein binds DNA: structure and a DNA binding surface as revealed by NMR. J Mol Biol. 1999 Jul 9;290(2):495–504. [PubMed]
  • Bai Y, Symington LS. A Rad52 homolog is required for RAD51-independent mitotic recombination in Saccharomyces cerevisiae. Genes Dev. 1996 Aug 15;10(16):2025–2037. [PubMed]
  • Bai Y, Davis AP, Symington LS. A novel allele of RAD52 that causes severe DNA repair and recombination deficiencies only in the absence of RAD51 or RAD59. Genetics. 1999 Nov;153(3):1117–1130. [PMC free article] [PubMed]
  • Clever B, Interthal H, Schmuckli-Maurer J, King J, Sigrist M, Heyer WD. Recombinational repair in yeast: functional interactions between Rad51 and Rad54 proteins. EMBO J. 1997 May 1;16(9):2535–2544. [PMC free article] [PubMed]
  • Donovan JW, Milne GT, Weaver DT. Homotypic and heterotypic protein associations control Rad51 function in double-strand break repair. Genes Dev. 1994 Nov 1;8(21):2552–2562. [PubMed]
  • Forsburg SL, Sherman DA. General purpose tagging vectors for fission yeast. Gene. 1997 Jun 3;191(2):191–195. [PubMed]
  • Game JC. DNA double-strand breaks and the RAD50-RAD57 genes in Saccharomyces. Semin Cancer Biol. 1993 Apr;4(2):73–83. [PubMed]
  • Golub EI, Kovalenko OV, Gupta RC, Ward DC, Radding CM. Interaction of human recombination proteins Rad51 and Rad54. Nucleic Acids Res. 1997 Oct 15;25(20):4106–4110. [PMC free article] [PubMed]
  • Gyuris J, Golemis E, Chertkov H, Brent R. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2. Cell. 1993 Nov 19;75(4):791–803. [PubMed]
  • Haber JE. The many interfaces of Mre11. Cell. 1998 Nov 25;95(5):583–586. [PubMed]
  • Harshman KD, Moye-Rowley WS, Parker CS. Transcriptional activation by the SV40 AP-1 recognition element in yeast is mediated by a factor similar to AP-1 that is distinct from GCN4. Cell. 1988 Apr 22;53(2):321–330. [PubMed]
  • Hays SL, Firmenich AA, Berg P. Complex formation in yeast double-strand break repair: participation of Rad51, Rad52, Rad55, and Rad57 proteins. Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):6925–6929. [PMC free article] [PubMed]
  • Hays SL, Firmenich AA, Massey P, Banerjee R, Berg P. Studies of the interaction between Rad52 protein and the yeast single-stranded DNA binding protein RPA. Mol Cell Biol. 1998 Jul;18(7):4400–4406. [PMC free article] [PubMed]
  • Heyer WD. The search for the right partner: homologous pairing and DNA strand exchange proteins in eukaryotes. Experientia. 1994 Mar 15;50(3):223–233. [PubMed]
  • Hopfner KP, Karcher A, Shin DS, Craig L, Arthur LM, Carney JP, Tainer JA. Structural biology of Rad50 ATPase: ATP-driven conformational control in DNA double-strand break repair and the ABC-ATPase superfamily. Cell. 2000 Jun 23;101(7):789–800. [PubMed]
  • Jiang H, Xie Y, Houston P, Stemke-Hale K, Mortensen UH, Rothstein R, Kodadek T. Direct association between the yeast Rad51 and Rad54 recombination proteins. J Biol Chem. 1996 Dec 27;271(52):33181–33186. [PubMed]
  • Johnson RD, Symington LS. Functional differences and interactions among the putative RecA homologs Rad51, Rad55, and Rad57. Mol Cell Biol. 1995 Sep;15(9):4843–4850. [PMC free article] [PubMed]
  • Khasanov FK, Savchenko GV, Bashkirova EV, Korolev VG, Heyer WD, Bashkirov VI. A new recombinational DNA repair gene from Schizosaccharomyces pombe with homology to Escherichia coli RecA. Genetics. 1999 Aug;152(4):1557–1572. [PMC free article] [PubMed]
  • Krejci L, Damborsky J, Thomsen B, Duno M, Bendixen C. Molecular dissection of interactions between Rad51 and members of the recombination-repair group. Mol Cell Biol. 2001 Feb;21(3):966–976. [PMC free article] [PubMed]
  • Schild D, Lio YC, Collins DW, Tsomondo T, Chen DJ. Evidence for simultaneous protein interactions between human Rad51 paralogs. J Biol Chem. 2000 Jun 2;275(22):16443–16449. [PubMed]
  • Kurumizaka H, Aihara H, Kagawa W, Shibata T, Yokoyama S. Human Rad51 amino acid residues required for Rad52 binding. J Mol Biol. 1999 Aug 20;291(3):537–548. [PubMed]
  • Shinohara A, Ogawa T. Homologous recombination and the roles of double-strand breaks. Trends Biochem Sci. 1995 Oct;20(10):387–391. [PubMed]
  • Mazin AV, Bornarth CJ, Solinger JA, Heyer WD, Kowalczykowski SC. Rad54 protein is targeted to pairing loci by the Rad51 nucleoprotein filament. Mol Cell. 2000 Sep;6(3):583–592. [PubMed]
  • Shinohara A, Ogawa T. Stimulation by Rad52 of yeast Rad51-mediated recombination. Nature. 1998 Jan 22;391(6665):404–407. [PubMed]
  • McCready SJ, Osman1 F, Yasui A. Repair of UV damage in the fission yeast Schizosaccharomyces pombe. Mutat Res. 2000 Jun 30;451(1-2):197–210. [PubMed]
  • 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. [PubMed]
  • Michel B, Ehrlich SD, Uzest M. DNA double-strand breaks caused by replication arrest. EMBO J. 1997 Jan 15;16(2):430–438. [PMC free article] [PubMed]
  • 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. [PubMed]
  • Morrison C, Takeda S. Genetic analysis of homologous DNA recombination in vertebrate somatic cells. Int J Biochem Cell Biol. 2000 Aug;32(8):817–831. [PubMed]
  • Shinohara A, Shinohara M, Ohta T, Matsuda S, Ogawa T. Rad52 forms ring structures and co-operates with RPA in single-strand DNA annealing. Genes Cells. 1998 Mar;3(3):145–156. [PubMed]
  • Mortensen UH, Bendixen C, Sunjevaric I, Rothstein R. DNA strand annealing is promoted by the yeast Rad52 protein. Proc Natl Acad Sci U S A. 1996 Oct 1;93(20):10729–10734. [PMC free article] [PubMed]
  • Sonoda E, Sasaki MS, Morrison C, Yamaguchi-Iwai Y, Takata M, Takeda S. Sister chromatid exchanges are mediated by homologous recombination in vertebrate cells. Mol Cell Biol. 1999 Jul;19(7):5166–5169. [PMC free article] [PubMed]
  • Muris DF, Vreeken K, Carr AM, Broughton BC, Lehmann AR, Lohman PH, Pastink A. Cloning the RAD51 homologue of Schizosaccharomyces pombe. Nucleic Acids Res. 1993 Sep 25;21(19):4586–4591. [PMC free article] [PubMed]
  • Muris DF, Vreeken K, Carr AM, Murray JM, Smit C, Lohman PH, Pastink A. Isolation of the Schizosaccharomyces pombe RAD54 homologue, rhp54+, a gene involved in the repair of radiation damage and replication fidelity. J Cell Sci. 1996 Jan;109(Pt 1):73–81. [PubMed]
  • Sugawara N, Haber JE. Characterization of double-strand break-induced recombination: homology requirements and single-stranded DNA formation. Mol Cell Biol. 1992 Feb;12(2):563–575. [PMC free article] [PubMed]
  • Needleman SB, Wunsch CD. A general method applicable to the search for similarities in the amino acid sequence of two proteins. J Mol Biol. 1970 Mar;48(3):443–453. [PubMed]
  • Sugawara N, Ira G, Haber JE. DNA length dependence of the single-strand annealing pathway and the role of Saccharomyces cerevisiae RAD59 in double-strand break repair. Mol Cell Biol. 2000 Jul;20(14):5300–5309. [PMC free article] [PubMed]
  • New JH, Sugiyama T, Zaitseva E, Kowalczykowski SC. Rad52 protein stimulates DNA strand exchange by Rad51 and replication protein A. Nature. 1998 Jan 22;391(6665):407–410. [PubMed]
  • Ogawa T, Yu X, Shinohara A, Egelman EH. Similarity of the yeast RAD51 filament to the bacterial RecA filament. Science. 1993 Mar 26;259(5103):1896–1899. [PubMed]
  • Ostermann K, Lorentz A, Schmidt H. The fission yeast rad22 gene, having a function in mating-type switching and repair of DNA damages, encodes a protein homolog to Rad52 of Saccharomyces cerevisiae. Nucleic Acids Res. 1993 Dec 25;21(25):5940–5944. [PMC free article] [PubMed]
  • Sung P. Catalysis of ATP-dependent homologous DNA pairing and strand exchange by yeast RAD51 protein. Science. 1994 Aug 26;265(5176):1241–1243. [PubMed]
  • Pâques F, Haber JE. Multiple pathways of recombination induced by double-strand breaks in Saccharomyces cerevisiae. Microbiol Mol Biol Rev. 1999 Jun;63(2):349–404. [PMC free article] [PubMed]
  • Park MS, Ludwig DL, Stigger E, Lee SH. Physical interaction between human RAD52 and RPA is required for homologous recombination in mammalian cells. J Biol Chem. 1996 Aug 2;271(31):18996–19000. [PubMed]
  • Sung P, Robberson DL. DNA strand exchange mediated by a RAD51-ssDNA nucleoprotein filament with polarity opposite to that of RecA. Cell. 1995 Aug 11;82(3):453–461. [PubMed]
  • Parker AE, Clyne RK, Carr AM, Kelly TJ. The Schizosaccharomyces pombe rad11+ gene encodes the large subunit of replication protein A. Mol Cell Biol. 1997 May;17(5):2381–2390. [PMC free article] [PubMed]
  • Sung P, Stratton SA. Yeast Rad51 recombinase mediates polar DNA strand exchange in the absence of ATP hydrolysis. J Biol Chem. 1996 Nov 8;271(45):27983–27986. [PubMed]
  • Sung P, Higgins D, Prakash L, Prakash S. Mutation of lysine-48 to arginine in the yeast RAD3 protein abolishes its ATPase and DNA helicase activities but not the ability to bind ATP. EMBO J. 1988 Oct;7(10):3263–3269. [PMC free article] [PubMed]
  • Petukhova G, Stratton SA, Sung P. Single strand DNA binding and annealing activities in the yeast recombination factor Rad59. J Biol Chem. 1999 Nov 26;274(48):33839–33842. [PubMed]
  • Petukhova G, Van Komen S, Vergano S, Klein H, Sung P. Yeast Rad54 promotes Rad51-dependent homologous DNA pairing via ATP hydrolysis-driven change in DNA double helix conformation. J Biol Chem. 1999 Oct 8;274(41):29453–29462. [PubMed]
  • Takata M, Sasaki MS, Tachiiri S, Fukushima T, Sonoda E, Schild D, Thompson LH, Takeda S. Chromosome instability and defective recombinational repair in knockout mutants of the five Rad51 paralogs. Mol Cell Biol. 2001 Apr;21(8):2858–2866. [PMC free article] [PubMed]
  • Pringle JR, Adams AE, Drubin DG, Haarer BK. Immunofluorescence methods for yeast. Methods Enzymol. 1991;194:565–602. [PubMed]
  • Tan TL, Essers J, Citterio E, Swagemakers SM, de Wit J, Benson FE, Hoeijmakers JH, Kanaar R. Mouse Rad54 affects DNA conformation and DNA-damage-induced Rad51 foci formation. Curr Biol. 1999 Mar 25;9(6):325–328. [PubMed]
  • Tanaka K, Hiramoto T, Fukuda T, Miyagawa K. A novel human rad54 homologue, Rad54B, associates with Rad51. J Biol Chem. 2000 Aug 25;275(34):26316–26321. [PubMed]
  • Rijkers T, Van Den Ouweland J, Morolli B, Rolink AG, Baarends WM, Van Sloun PP, Lohman PH, Pastink A. Targeted inactivation of mouse RAD52 reduces homologous recombination but not resistance to ionizing radiation. Mol Cell Biol. 1998 Nov;18(11):6423–6429. [PMC free article] [PubMed]
  • Tavassoli M, Shayeghi M, Nasim A, Watts FZ. Cloning and characterisation of the Schizosaccharomyces pombe rad32 gene: a gene required for repair of double strand breaks and recombination. Nucleic Acids Res. 1995 Feb 11;23(3):383–388. [PMC free article] [PubMed]
  • Santos-Rosa H, Clever B, Heyer WD, Aguilera A. The yeast HRS1 gene encodes a polyglutamine-rich nuclear protein required for spontaneous and hpr1-induced deletions between direct repeats. Genetics. 1996 Mar;142(3):705–716. [PMC free article] [PubMed]
  • Thompson LH, Schild D. The contribution of homologous recombination in preserving genome integrity in mammalian cells. Biochimie. 1999 Jan-Feb;81(1-2):87–105. [PubMed]
  • Van Komen S, Petukhova G, Sigurdsson S, Stratton S, Sung P. Superhelicity-driven homologous DNA pairing by yeast recombination factors Rad51 and Rad54. Mol Cell. 2000 Sep;6(3):563–572. [PubMed]
  • Yamaguchi-Iwai Y, Sonoda E, Buerstedde JM, Bezzubova O, Morrison C, Takata M, Shinohara A, Takeda S. Homologous recombination, but not DNA repair, is reduced in vertebrate cells deficient in RAD52. Mol Cell Biol. 1998 Nov;18(11):6430–6435. [PMC free article] [PubMed]
  • van den Bosch M, Vreeken K, Zonneveld JB, Brandsma JA, Lombaerts M, Murray JM, Lohman PH, Pastink A. Characterization of RAD52 homologs in the fission yeast Schizosaccharomyces pombe. Mutat Res. 2001 Jan 5;461(4):311–323. [PubMed]

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