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Eukaryotic polymerases ι and ζ act sequentially to bypass DNA lesions

Nature volume 406, pages 1015–1019 (2000)Cite this article

Abstract

DNA lesions can often block DNA replication, so cells possess specialized low-fidelity, and often error-prone, DNA polymerases that can bypass such lesions and promote replication of damaged DNA1. The Saccharomyces cerevisiae RAD30 and human hRAD30A encode Polη, which bypasses a cis–syn thymine–thymine dimer efficiently and accurately2,3,4,5,6,7. Here we show that a related human gene, hRAD30B8, encodes the DNA polymerase Polι, which misincorporates deoxynucleotides at a high rate. To bypass damage, Polι specifically incorporates deoxynucleotides opposite highly distorting or non-instructional DNA lesions. This action is combined with that of DNA polymerase Polζ, which is essential for damage-induced mutagenesis, to complete the lesion bypass. Polζ is very inefficient in inserting deoxynucleotides opposite DNA lesions, but readily extends from such deoxynucleotides once they have been inserted. Thus, in a new model for mutagenic bypass of DNA lesions in eukaryotes, the two DNA polymerases act sequentially: Polι incorporates deoxynucleotides opposite DNA lesions, and Polζ functions as a mispair extender.

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Figure 1: Purification and activity of human Rad30B protein.
Figure 2: Deoxynucleotide incorporation by Polι on undamaged and damaged DNA and its role in damage bypass.
Figure 3: Comparison of Polζ mispair extension and mispair insertion efficiencies.

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References

  1. Johnson, R. E., Washington, M. T., Prakash, S. & Prakash, L. Bridging the gap: a family of novel DNA polymerases that replicate faulty DNA. Proc. Natl Acad. Sci. USA 96, 12224 –12226 (1999).

    Article  ADS  CAS  Google Scholar 

  2. Johnson, R. E., Prakash, S. & Prakash, L. Efficient bypass of a thymine-thymine dimer by yeast DNA polymerase, Polη. Science 283, 1001 –1004 (1999).

    Article  ADS  CAS  Google Scholar 

  3. Johnson, R. E., Kondratick, C. M., Prakash, S. & Prakash, L. hRAD30 mutations in the variant form of xeroderm pigmentosum. Science 285, 263–265 ( 1999).

    Article  CAS  Google Scholar 

  4. Masutani, C. et al. The XPV (xeroderma pigmentosum variant) gene encodes human DNA polymerase η. Nature 399, 700–704 (1999).

    Article  ADS  CAS  Google Scholar 

  5. Washington, M. T., Johnson, R. E., Prakash, S. & Prakash, L. Accuracy of thymine-thymine dimer bypass by Saccharomyces cerevisiae DNA polymerase η. Proc. Natl Acad. Sci. USA 97, 3094–3099 (2000).

    ADS  CAS  PubMed  Google Scholar 

  6. Johnson, R. E., Washington, M. T., Prakash, S. & Prakash, L. Fidelity of human DNA polymerase η. J. Biol. Chem. 275, 7447–7450 (2000).

    Article  CAS  Google Scholar 

  7. Washington, M. T., Johnson, R. E., Prakash, S. & Prakash, L. Fidelity and processivity of Saccharomyces cerevisiae DNA polymerase η. J. Biol. Chem. 274, 36835– 36838 (1999).

    Article  CAS  Google Scholar 

  8. McDonald, J. P. et al. Novel human and mouse homologs of Saccharomyces cerevisiae DNA polymerase η. Genomics 60, 20–30 (1999).

    Article  CAS  Google Scholar 

  9. Goodman, M. F., Creighton, S., Bloom, L. B. & Petruska, J. Biochemical basis of DNA replication fidelity. Crit. Rev. Biochem. Mol. Biol. 28, 83–126 ( 1993).

    Article  CAS  Google Scholar 

  10. Creighton, S., Bloom, L. B. & Goodman, M. F. Gel fidelity assay measuring nucleotide misinsertion, exonucleolytic proofreading, and lesion bypass efficiencies. Methods Enzymol. 262, 232–256 ( 1995).

    Article  CAS  Google Scholar 

  11. Nelson, J. R., Lawrence, C. W. & Hinkle, D. C. Thymine-thymine dimer bypass by yeast DNA polymerase ζ. Science 272, 1646–1649 (1996).

    Article  ADS  CAS  Google Scholar 

  12. Gibbs, P. E. M., McGregor, W. G., Maher, V. M., Nisson, P. & Lawrence, C. W. A human homolog of the Saccharomyces cerevisiae REV3 gene, which encodes the catalytic subunit of DNA polyerase ζ. Proc. Natl Acad. Sci. USA 95, 6876– 6880 (1998).

    Article  ADS  CAS  Google Scholar 

  13. Thomas, D. C. et al. Fidelity of mammalian DNA replication and replicative DNA polymerases. Biochem. 30, 11751– 11759 (1991).

    Article  CAS  Google Scholar 

  14. Mendelman, L. V., Petruska, J. & Goodman, M. F. Base mispair extension kinetics. Comparison of DNA polymerase α and reverse transcriptase. J. Biol. Chem. 265, 2338–2346 (1990).

    CAS  PubMed  Google Scholar 

  15. Gibbs, P. E. M., Kilbey, B. J., Banerjee, S. K. & Lawrence, C. W. The frequency and accuracy of replication past a thymine-thymine cyclobutane dimer are very different in Saccharomyces cerevisiae and Escherichia coli. J. Bacteriol. 175, 2607– 2612 (1993).

    Article  CAS  Google Scholar 

  16. Gibbs, P. E. M., Borden, A. & Lawrence, C. W. The T–T pyrimidine (6-4) pyrimidinone UV photoproduct is much less mutagenic in yeast than in Escherichia coli. Nucleic Acids Res. 23, 1919–1922 (1995).

    Article  CAS  Google Scholar 

  17. Ciarrocchi, G. & Pedrini, A. M. Determination of pyrimidine dimer unwinding angle by measurement of DNA electrophoretic mobility. J. Mol. Biol. 155, 177– 183 (1982).

    Article  CAS  Google Scholar 

  18. Kim, J.-K., Patel, D. & Choi, B.-S. Contrasting structural impacts induced by cis–syn cyclobutane dimer and (6-4) adduct in DNA duplex decamers: implication in mutagenesis and repair activity. Photochem. Photobiol. 62, 44–50 (1995).

    Article  CAS  Google Scholar 

  19. Kim, J.-K. & Choi, B.-S. The solution structure of DNA duplex-decamer containing the (6-4) photoproduct of thymidyl(3′→5′)thymidine by NMR and relaxation matrix refinement. Eur. J. Biochem. 228, 849–854 (1995).

    Article  CAS  Google Scholar 

  20. Wong, I., Patel, S. S. & Johnson, K. A. An induced-fit kinetic mechanism for DNA replication fidelity: direct measurement by single-turnover kinetics. Biochemistry 30, 526–537 ( 1991).

    Article  CAS  Google Scholar 

  21. Huang, M.-M., Arnheim, N. & Goodman, M. F. Extension of base mispairs by Taq DNA polymerase: implications for single nucleotide discrimination in PCR. Nucleic Acids Res. 20, 4567–4573 (1992).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank D. Hinkle for plasmids pGST-Rev3 and pRev7 and R. Hodge for providing the T–T dimer- and (6-4) photoproduct-containing DNAs, the construction of which was supported by a National Institute of Environmental Health Science (NIEHS) Centre Grant. This work was supported by a grant from the NIH.

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Author notes

  1. Robert E. Johnson, M. Todd Washington and Lajos Haracska: These authors contributed equally to this work.

  2. Correspondence and requests for materials should be addressed to L.P. .

Authors
  1. Robert E. Johnson
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  2. M. Todd Washington
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  3. Lajos Haracska
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  4. Satya Prakash
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  5. Louise Prakash
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Sealy Centre for Molecular Science, University of Texas Medical Branch at Galveston, 6.104 Medical Research Building, 11th and Mechanic Streets, Galveston, Texas 77555-1061, USA

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Johnson, R., Washington, M., Haracska, L. et al. Eukaryotic polymerases ι and ζ act sequentially to bypass DNA lesions. Nature 406, 1015–1019 (2000). https://doi.org/10.1038/35023030

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