Skip to main content
PMC home page
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1991 Apr 15;88(8):3465–3469. doi: 10.1073/pnas.88.8.3465

Exonucleolytic proofreading of leading and lagging strand DNA replication errors.

J D Roberts 1, D C Thomas 1, T A Kunkel 1
PMCID: PMC51468  PMID: 1901658

Abstract

We have asked whether exonucleolytic proofreading occurs during simian virus 40 origin-dependent, bidirectional DNA replication in extracts of human HeLa cells. In addition, we have compared the fidelity of leading and lagging strand DNA synthesis. In a fidelity assay that scores single-base substitution errors that revert a TGA codon in the lacZ alpha gene in an M13mp vector, providing an excess of a single dNTP substrate over the other three dNTP substrates in a replication reaction generates defined, strand-specific errors. Fidelity measurements with two vectors having the origin of replication on opposite sides of the opal codon demonstrate that error rates for two different A.dCTP and T.dGTP mispairs increase when deoxyguanosine monophosphate is added to replication reaction mixtures or when the concentration of deoxynucleoside triphosphates is increased. The data suggest that exonucleolytic proofreading occurs on both strands during bidirectional replication. Measurements using the two simian virus 40 origin-containing vectors suggest that base substitution error rates are similar for replication of the leading and lagging strands.

Full text

PDF
3465

Selected References

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

  1. Abbotts J., Loeb L. A. DNA polymerase alpha and models for proofreading. Nucleic Acids Res. 1985 Jan 11;13(1):261–274. doi: 10.1093/nar/13.1.261. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Akiyama M., Maki H., Sekiguchi M., Horiuchi T. A specific role of MutT protein: to prevent dG.dA mispairing in DNA replication. Proc Natl Acad Sci U S A. 1989 Jun;86(11):3949–3952. doi: 10.1073/pnas.86.11.3949. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bambara R. A., Jessee C. B. Properties of DNA polymerases delta and epsilon, and their roles in eukaryotic DNA replication. Biochim Biophys Acta. 1991 Jan 17;1088(1):11–24. doi: 10.1016/0167-4781(91)90147-e. [DOI] [PubMed] [Google Scholar]
  4. Bebenek K., Joyce C. M., Fitzgerald M. P., Kunkel T. A. The fidelity of DNA synthesis catalyzed by derivatives of Escherichia coli DNA polymerase I. J Biol Chem. 1990 Aug 15;265(23):13878–13887. [PubMed] [Google Scholar]
  5. Bhatnagar S. K., Bessman M. J. Studies on the mutator gene, mutT of Escherichia coli. Molecular cloning of the gene, purification of the gene product, and identification of a novel nucleoside triphosphatase. J Biol Chem. 1988 Jun 25;263(18):8953–8957. [PubMed] [Google Scholar]
  6. Burgers P. M., Bambara R. A., Campbell J. L., Chang L. M., Downey K. M., Hübscher U., Lee M. Y., Linn S. M., So A. G., Spadari S. Revised nomenclature for eukaryotic DNA polymerases. Eur J Biochem. 1990 Aug 17;191(3):617–618. doi: 10.1111/j.1432-1033.1990.tb19165.x. [DOI] [PubMed] [Google Scholar]
  7. Byrnes J. J., Downey K. M., Que B. G., Lee M. Y., Black V. L., So A. G. Selective inhibition of the 3' to 5' exonuclease activity associated with DNA polymerases: a mechanism of mutagenesis. Biochemistry. 1977 Aug 23;16(17):3740–3746. doi: 10.1021/bi00636a002. [DOI] [PubMed] [Google Scholar]
  8. Clayton L. K., Goodman M. F., Branscomb E. W., Galas D. J. Error induction and correction by mutant and wild type T4 DNA polymerases. Kinetic error discrimination mechanisms. J Biol Chem. 1979 Mar 25;254(6):1902–1912. [PubMed] [Google Scholar]
  9. Cotterill S. M., Reyland M. E., Loeb L. A., Lehman I. R. A cryptic proofreading 3'----5' exonuclease associated with the polymerase subunit of the DNA polymerase-primase from Drosophila melanogaster. Proc Natl Acad Sci U S A. 1987 Aug;84(16):5635–5639. doi: 10.1073/pnas.84.16.5635. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Crute J. J., Wahl A. F., Bambara R. A. Purification and characterization of two new high molecular weight forms of DNA polymerase delta. Biochemistry. 1986 Jan 14;25(1):26–36. doi: 10.1021/bi00349a005. [DOI] [PubMed] [Google Scholar]
  11. DiFrancesco R., Bhatnagar S. K., Brown A., Bessman M. J. The interaction of DNA polymerase III and the product of the Escherichia coli mutator gene, mutD. J Biol Chem. 1984 May 10;259(9):5567–5573. [PubMed] [Google Scholar]
  12. Edenberg H. J., Huberman J. A. Eukaryotic chromosome replication. Annu Rev Genet. 1975;9:245–284. doi: 10.1146/annurev.ge.09.120175.001333. [DOI] [PubMed] [Google Scholar]
  13. Fersht A. R. Fidelity of replication of phage phi X174 DNA by DNA polymerase III holoenzyme: spontaneous mutation by misincorporation. Proc Natl Acad Sci U S A. 1979 Oct;76(10):4946–4950. doi: 10.1073/pnas.76.10.4946. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Fersht A. R., Knill-Jones J. W. Contribution of 3' leads to 5' exonuclease activity of DNA polymerase III holoenzyme from Escherichia coli to specificity. J Mol Biol. 1983 Apr 25;165(4):669–682. doi: 10.1016/s0022-2836(83)80273-3. [DOI] [PubMed] [Google Scholar]
  15. Focher F., Ferrari E., Spadari S., Hübscher U. Do DNA polymerases delta and alpha act coordinately as leading and lagging strand replicases? FEBS Lett. 1988 Feb 29;229(1):6–10. doi: 10.1016/0014-5793(88)80786-5. [DOI] [PubMed] [Google Scholar]
  16. Hauser J., Levine A. S., Dixon K. Fidelity of DNA synthesis in a mammalian in vitro replication system. Mol Cell Biol. 1988 Aug;8(8):3267–3271. doi: 10.1128/mcb.8.8.3267. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hay R. T., DePamphilis M. L. Initiation of SV40 DNA replication in vivo: location and structure of 5' ends of DNA synthesized in the ori region. Cell. 1982 Apr;28(4):767–779. doi: 10.1016/0092-8674(82)90056-3. [DOI] [PubMed] [Google Scholar]
  18. Holmes J., Jr, Clark S., Modrich P. Strand-specific mismatch correction in nuclear extracts of human and Drosophila melanogaster cell lines. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5837–5841. doi: 10.1073/pnas.87.15.5837. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hurwitz J., Dean F. B., Kwong A. D., Lee S. H. The in vitro replication of DNA containing the SV40 origin. J Biol Chem. 1990 Oct 25;265(30):18043–18046. [PubMed] [Google Scholar]
  20. Kelly T. J. SV40 DNA replication. J Biol Chem. 1988 Dec 5;263(34):17889–17892. [PubMed] [Google Scholar]
  21. Kunkel T. A. Exonucleolytic proofreading. Cell. 1988 Jun 17;53(6):837–840. doi: 10.1016/s0092-8674(88)90189-4. [DOI] [PubMed] [Google Scholar]
  22. Kunkel T. A., Loeb L. A. On the fidelity of DNA replication. Effect of divalent metal ion activators and deoxyrionucleoside triphosphate pools on in vitro mutagenesis. J Biol Chem. 1979 Jul 10;254(13):5718–5725. [PubMed] [Google Scholar]
  23. Kunkel T. A., Roberts J. D., Zakour R. A. Rapid and efficient site-specific mutagenesis without phenotypic selection. Methods Enzymol. 1987;154:367–382. doi: 10.1016/0076-6879(87)54085-x. [DOI] [PubMed] [Google Scholar]
  24. Kunkel T. A., Sabatino R. D., Bambara R. A. Exonucleolytic proofreading by calf thymus DNA polymerase delta. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4865–4869. doi: 10.1073/pnas.84.14.4865. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kunkel T. A., Schaaper R. M., Beckman R. A., Loeb L. A. On the fidelity of DNA replication. Effect of the next nucleotide on proofreading. J Biol Chem. 1981 Oct 10;256(19):9883–9889. [PubMed] [Google Scholar]
  26. Li J. J., Kelly T. J. Simian virus 40 DNA replication in vitro. Proc Natl Acad Sci U S A. 1984 Nov;81(22):6973–6977. doi: 10.1073/pnas.81.22.6973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Li J. J., Kelly T. J. Simian virus 40 DNA replication in vitro: specificity of initiation and evidence for bidirectional replication. Mol Cell Biol. 1985 Jun;5(6):1238–1246. doi: 10.1128/mcb.5.6.1238. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Morrison A., Araki H., Clark A. B., Hamatake R. K., Sugino A. A third essential DNA polymerase in S. cerevisiae. Cell. 1990 Sep 21;62(6):1143–1151. doi: 10.1016/0092-8674(90)90391-q. [DOI] [PubMed] [Google Scholar]
  29. Ninio J. Kinetic amplification of enzyme discrimination. Biochimie. 1975;57(5):587–595. doi: 10.1016/s0300-9084(75)80139-8. [DOI] [PubMed] [Google Scholar]
  30. Perrino F. W., Loeb L. A. Hydrolysis of 3'-terminal mispairs in vitro by the 3'----5' exonuclease of DNA polymerase delta permits subsequent extension by DNA polymerase alpha. Biochemistry. 1990 Jun 5;29(22):5226–5231. doi: 10.1021/bi00474a002. [DOI] [PubMed] [Google Scholar]
  31. Prelich G., Stillman B. Coordinated leading and lagging strand synthesis during SV40 DNA replication in vitro requires PCNA. Cell. 1988 Apr 8;53(1):117–126. doi: 10.1016/0092-8674(88)90493-x. [DOI] [PubMed] [Google Scholar]
  32. Que B. G., Downey K. M., So A. G. Mechanisms of selective inhibition of 3' to 5' exonuclease activity of Escherichia coli DNA polymerase I by nucleoside 5'-monophosphates. Biochemistry. 1978 May 2;17(9):1603–1606. doi: 10.1021/bi00602a004. [DOI] [PubMed] [Google Scholar]
  33. Roberts J. D., Kunkel T. A. Fidelity of a human cell DNA replication complex. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7064–7068. doi: 10.1073/pnas.85.19.7064. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Schaaper R. M., Dunn R. L. Escherichia coli mutT mutator effect during in vitro DNA synthesis. Enhanced A.G replicational errors. J Biol Chem. 1987 Dec 5;262(34):16267–16270. [PubMed] [Google Scholar]
  35. Scheuermann R. H., Echols H. A separate editing exonuclease for DNA replication: the epsilon subunit of Escherichia coli DNA polymerase III holoenzyme. Proc Natl Acad Sci U S A. 1984 Dec;81(24):7747–7751. doi: 10.1073/pnas.81.24.7747. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Stillman B. Initiation of eukaryotic DNA replication in vitro. Annu Rev Cell Biol. 1989;5:197–245. doi: 10.1146/annurev.cb.05.110189.001213. [DOI] [PubMed] [Google Scholar]
  37. Thomas D. C., Roberts J. D., Kunkel T. A. Heteroduplex repair in extracts of human HeLa cells. J Biol Chem. 1991 Feb 25;266(6):3744–3751. [PubMed] [Google Scholar]
  38. Thummel C., Tjian R., Hu S. L., Grodzicker T. Translational control of SV40 T antigen expressed from the adenovirus late promoter. Cell. 1983 Jun;33(2):455–464. doi: 10.1016/0092-8674(83)90427-0. [DOI] [PubMed] [Google Scholar]
  39. Tsurimoto T., Melendy T., Stillman B. Sequential initiation of lagging and leading strand synthesis by two different polymerase complexes at the SV40 DNA replication origin. Nature. 1990 Aug 9;346(6284):534–539. doi: 10.1038/346534a0. [DOI] [PubMed] [Google Scholar]
  40. Wold M. S., Kelly T. Purification and characterization of replication protein A, a cellular protein required for in vitro replication of simian virus 40 DNA. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2523–2527. doi: 10.1073/pnas.85.8.2523. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Wold M. S., Weinberg D. H., Virshup D. M., Li J. J., Kelly T. J. Identification of cellular proteins required for simian virus 40 DNA replication. J Biol Chem. 1989 Feb 15;264(5):2801–2809. [PubMed] [Google Scholar]
  42. Wu C. I., Maeda N. Inequality in mutation rates of the two strands of DNA. Nature. 1987 May 14;327(6118):169–170. doi: 10.1038/327169a0. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES