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PMID: 10490626 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

The essential functions of human Rad51 are independent of ATP hydrolysis.

Molecular and cellular biology ·Vol. 19 ·No. 10 ·1999-10-00 ·Pages 6891-7

Morrison C, Shinohara A, Sonoda E, Yamaguchi-Iwai Y, Takata M, Weichselbaum RR, Takeda S

Abstract

Genetic recombination and the repair of double-strand DNA breaks in Saccharomyces cerevisiae require Rad51, a homologue of the Escherichia coli RecA protein. In vitro, Rad51 binds DNA to form an extended nucleoprotein filament and catalyzes the ATP-dependent exchange of DNA between molecules with homologous sequences. Vertebrate Rad51 is essential for cell proliferation. Using site-directed mutagenesis of highly conserved residues of human Rad51 (hRad51) and gene targeting of the RAD51 locus in chicken DT40 cells, we examined the importance of Rad51's highly conserved ATP-binding domain. Mutant hRad51 incapable of ATP hydrolysis (hRad51K-133R) binds DNA less efficiently than the wild type but catalyzes strand exchange between homologous DNAs. hRad51 does not need to hydrolyze ATP to allow vertebrate cell proliferation, form nuclear foci, or repair radiation-induced DNA damage. However, cells expressing hRad51K-133R show greatly reduced targeted integration frequencies. These findings show that ATP hydrolysis is involved in DNA binding by hRad51 and suggest that the extent of DNA complexed with hRad51 in nucleoprotein influences the efficiency of recombination.

MeSH Terms
Adenosine Triphosphate/metabolism Amino Acid Sequence Avian Proteins Binding Sites/genetics Cell Division/radiation effects Cell Survival Conserved Sequence DNA Nucleotidyltransferases/genetics,metabolism DNA Repair DNA-Binding Proteins/genetics,metabolism Gamma Rays Genes, Essential Genetic Complementation Test Humans Hydrolysis Mutagenesis, Site-Directed Protein Binding Rad51 Recombinase
Chemicals
Avian Proteins DNA-Binding Proteins Adenosine Triphosphate DNA Nucleotidyltransferases RAD51 protein, Gallus gallus RAD51 protein, human Rad51 Recombinase
Authors & Affiliations
7 authors, click to expand affiliations / ORCID
Morrison C
Bayer-Chair Department of Molecular Immunology and Allergology, Faculty of Medicine, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan.
Shinohara A
Sonoda E
Yamaguchi-Iwai Y
Takata M
Weichselbaum R R
Takeda S
References (53)
53 references, click to expand
  1. Catalysis of ATP-dependent homologous DNA pairing and strand exchange by yeast RAD51 protein.
    Science. 1994 Aug 26;265(5176):1241-3 PMID: 8066464
  2. Interaction of recA protein with single-stranded DNA. Quantitative aspects of binding affinity modulation by nucleotide cofactors.
    J Mol Biol. 1985 Jan 20;181(2):281-95 PMID: 3981638
  3. Homologous pairing and DNA strand-exchange proteins.
    Annu Rev Biochem. 1994;63:991-1043 PMID: 7979259
  4. Purification and characterization of the human Rad51 protein, an analogue of E. coli RecA.
    EMBO J. 1994 Dec 1;13(23):5764-71 PMID: 7988572
  5. Nuclear foci of mammalian Rad51 recombination protein in somatic cells after DNA damage and its localization in synaptonemal complexes.
    Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):2298-302 PMID: 7892263
  6. DNA-strand exchange promoted by RecA protein in the absence of ATP: implications for the mechanism of energy transduction in protein-promoted nucleic acid transactions.
    Proc Natl Acad Sci U S A. 1995 Apr 11;92(8):3478-82 PMID: 7724585
  7. DNA strand exchange mediated by a RAD51-ssDNA nucleoprotein filament with polarity opposite to that of RecA.
    Cell. 1995 Aug 11;82(3):453-61 PMID: 7634335
  8. DNA strand exchange promoted by RecA K72R. Two reaction phases with different Mg2+ requirements.
    J Biol Chem. 1996 Mar 8;271(10):5712-24 PMID: 8621437
  9. Evidence for the coupling of ATP hydrolysis to the final (extension) phase of RecA protein-mediated DNA strand exchange.
    J Biol Chem. 1996 Mar 8;271(10):5725-32 PMID: 8621438
  10. S phase specific formation of the human Rad51 protein nuclear foci in lymphocytes.
    Oncogene. 1996 May 16;12(10):2165-70 PMID: 8668342
  11. Stable DNA heteroduplex formation catalyzed by the Escherichia coli RecA protein in the absence of ATP hydrolysis.
    Proc Natl Acad Sci U S A. 1990 Jan;87(1):21-5 PMID: 2404275
  12. The RecA protein: structure and function.
    Crit Rev Biochem Mol Biol. 1990;25(6):415-56 PMID: 2292186
  13. The structure of the E. coli recA protein monomer and polymer.
    Nature. 1992 Jan 23;355(6358):318-25 PMID: 1731246
  14. Structure of the recA protein-ADP complex.
    Nature. 1992 Jan 23;355(6358):374-6 PMID: 1731253
  15. Rad51 protein involved in repair and recombination in S. cerevisiae is a RecA-like protein.
    Cell. 1992 May 1;69(3):457-70 PMID: 1581961
  16. Semidominant suppressors of Srs2 helicase mutations of Saccharomyces cerevisiae map in the RAD51 gene, whose sequence predicts a protein with similarities to procaryotic RecA proteins.
    Mol Cell Biol. 1992 Jul;12(7):3224-34 PMID: 1620127
  17. Nucleotide sequence and transcriptional regulation of the yeast recombinational repair gene RAD51.
    Mol Cell Biol. 1992 Jul;12(7):3235-46 PMID: 1620128
  18. Alteration of the nucleoside triphosphate (NTP) catalytic domain within Escherichia coli recA protein attenuates NTP hydrolysis but not joint molecule formation.
    J Biol Chem. 1993 Jan 15;268(2):1292-7 PMID: 8419331
  19. Structural relationship of bacterial RecA proteins to recombination proteins from bacteriophage T4 and yeast.
    Science. 1993 Mar 26;259(5103):1892-6 PMID: 8456313
  20. Similarity of the yeast RAD51 filament to the bacterial RecA filament.
    Science. 1993 Mar 26;259(5103):1896-9 PMID: 8456314
  21. Cloning of human, mouse and fission yeast recombination genes homologous to RAD51 and recA.
    Nat Genet. 1993 Jul;4(3):239-43 PMID: 8358431
  22. Dominant negative alleles of RAD52 reveal a DNA repair/recombination complex including Rad51 and Rad52.
    Genes Dev. 1993 Sep;7(9):1755-65 PMID: 8370524
  23. Mutagenesis of the P-loop motif in the ATP binding site of the RecA protein from Escherichia coli.
    J Mol Biol. 1993 Aug 20;232(4):1048-59 PMID: 8371266
  24. Functional characterization of residues in the P-loop motif of the RecA protein ATP binding site.
    J Mol Biol. 1994 Mar 18;237(1):20-34 PMID: 8133517
  25. Tyrosine kinases Lyn and Syk regulate B cell receptor-coupled Ca2+ mobilization through distinct pathways.
    EMBO J. 1994 Mar 15;13(6):1341-9 PMID: 8137818
  26. Syk activation by the Src-family tyrosine kinase in the B cell receptor signaling.
    J Exp Med. 1994 May 1;179(5):1725-9 PMID: 7513017
  27. Why does RecA protein hydrolyse ATP?
    Trends Biochem Sci. 1994 May;19(5):217-22 PMID: 8048163
  28. Activities of human recombination protein Rad51.
    Proc Natl Acad Sci U S A. 1997 Jan 21;94(2):463-8 PMID: 9012806
  29. DNA double-strand breaks caused by replication arrest.
    EMBO J. 1997 Jan 15;16(2):430-8 PMID: 9029161
  30. Reduced X-ray resistance and homologous recombination frequencies in a RAD54-/- mutant of the chicken DT40 cell line.
    Cell. 1997 Apr 18;89(2):185-93 PMID: 9108474
  31. Evolutionary comparisons of RecA-like proteins across all major kingdoms of living organisms.
    J Mol Evol. 1997 May;44(5):528-41 PMID: 9115177
  32. RecA as a motor protein. Testing models for the role of ATP hydrolysis in DNA strand exchange.
    J Biol Chem. 1997 Jul 11;272(28):17675-85 PMID: 9211918
  33. Holliday junctions accumulate in replication mutants via a RecA homolog-independent mechanism.
    Cell. 1997 Jul 11;90(1):87-96 PMID: 9230305
  34. Purification of human Rad51 protein by selective spermidine precipitation.
    Mutat Res. 1997 Aug;384(2):65-72 PMID: 9298115
  35. Rad51-deficient vertebrate cells accumulate chromosomal breaks prior to cell death.
    EMBO J. 1998 Jan 15;17(2):598-608 PMID: 9430650
  36. Stimulation by Rad52 of yeast Rad51-mediated recombination.
    Nature. 1998 Jan 22;391(6665):404-7 PMID: 9450759
  37. Binding of Rad51p to DNA. Interaction of Rad51p with single- and double-stranded DNA.
    J Biol Chem. 1998 Mar 13;273(11):6177-82 PMID: 9497339
  38. XRCC2 and XRCC3, new human Rad51-family members, promote chromosome stability and protect against DNA cross-links and other damages.
    Mol Cell. 1998 May;1(6):783-93 PMID: 9660962
  39. Role of the human RAD51 protein in homologous recombination and double-stranded-break repair.
    Trends Biochem Sci. 1998 Jul;23(7):247-51 PMID: 9697414
  40. Xrcc3 is required for assembly of Rad51 complexes in vivo.
    J Biol Chem. 1998 Aug 21;273(34):21482-8 PMID: 9705276
  41. Homologous recombination and non-homologous end-joining pathways of DNA double-strand break repair have overlapping roles in the maintenance of chromosomal integrity in vertebrate cells.
    EMBO J. 1998 Sep 15;17(18):5497-508 PMID: 9736627
  42. Homologous recombination, but not DNA repair, is reduced in vertebrate cells deficient in RAD52.
    Mol Cell Biol. 1998 Nov;18(11):6430-5 PMID: 9774659
  43. RuvAB acts at arrested replication forks.
    Cell. 1998 Oct 30;95(3):419-30 PMID: 9814711
  44. Human Rad51 protein can form homologous joints in the absence of net strand exchange.
    J Biol Chem. 1999 Jan 15;274(3):1248-56 PMID: 9880493
  45. Sequestration of mammalian Rad51-recombination protein into micronuclei.
    J Cell Biol. 1999 Jan 11;144(1):11-20 PMID: 9885240
  46. Sister chromatid exchanges are mediated by homologous recombination in vertebrate cells.
    Mol Cell Biol. 1999 Jul;19(7):5166-9 PMID: 10373565
  47. Targeted disruption of the Rad51 gene leads to lethality in embryonic mice.
    Proc Natl Acad Sci U S A. 1996 Jun 25;93(13):6236-40 PMID: 8692798
  48. Semidominant mutations in the yeast Rad51 protein and their relationships with the Srs2 helicase.
    Mol Cell Biol. 1996 Sep;16(9):4782-9 PMID: 8756636
  49. Yeast Rad51 recombinase mediates polar DNA strand exchange in the absence of ATP hydrolysis.
    J Biol Chem. 1996 Nov 8;271(45):27983-6 PMID: 8910403
  50. Human Rad51 protein promotes ATP-dependent homologous pairing and strand transfer reactions in vitro.
    Cell. 1996 Nov 15;87(4):757-66 PMID: 8929543
  51. A mutation in mouse rad51 results in an early embryonic lethal that is suppressed by a mutation in p53.
    Mol Cell Biol. 1996 Dec;16(12):7133-43 PMID: 8943369
  52. Distantly related sequences in the alpha- and beta-subunits of ATP synthase, myosin, kinases and other ATP-requiring enzymes and a common nucleotide binding fold.
    EMBO J. 1982;1(8):945-51 PMID: 6329717
  53. Homotypic and heterotypic protein associations control Rad51 function in double-strand break repair.
    Genes Dev. 1994 Nov 1;8(21):2552-62 PMID: 7958917
Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/MCB.19.10.6891
Published
1999-10-00
Pages
6891-7
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC84684
Subset
IM
External Links
PubMed source DOI: 10.1128/MCB.19.10.6891 PMC Full Text View journal details
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