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. 1986 Oct;6(10):3349–3356. doi: 10.1128/mcb.6.10.3349

UV light-induced cyclobutane pyrimidine dimers are mutagenic in mammalian cells.

M Protić-Sabljić, N Tuteja, P J Munson, J Hauser, K H Kraemer, K Dixon
PMCID: PMC367080  PMID: 3540589

Abstract

We used a simian virus 40-based shuttle vector plasmid, pZ189, to determine the role of pyrimidine cyclobutane dimers in UV light-induced mutagenesis in monkey cells. The vector DNA was UV irradiated and then introduced into monkey cells by transfection. After replication, vector DNA was recovered from the cells and tested for mutations in its supF suppressor tRNA marker gene by transformation of Escherichia coli carrying a nonsense mutation in the beta-galactosidase gene. When the irradiated vector was treated with E. coli photolyase prior to transfection, pyrimidine cyclobutane dimers were removed selectively. Removal of approximately 90% of the pyrimidine cyclobutane dimers increased the biological activity of the vector by 75% and reduced its mutation frequency by 80%. Sequence analysis of 72 mutants recovered indicated that there were significantly fewer tandem double-base changes and G X C----A X T transitions (particularly at CC sites) after photoreactivation of the DNA. UV-induced photoproducts remained (although at greatly reduced levels) at all pyr-pyr sites after photoreactivation, but there was a relative increase in photoproducts at CC and TC sites and a relative decrease at TT and CT sites, presumably due to a persistence of (6-4) photoproducts at some CC and TC sites. These observations are consistent with the fact that mutations were found after photoreactivation at many sites at which only cyclobutane dimers would be expected to occur. From these results we conclude that UV-induced pyrimidine cyclobutane dimers are mutagenic in DNA replicated in monkey cells.

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Selected References

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  1. Bishop J. M. Cancer genes come of age. Cell. 1983 Apr;32(4):1018–1020. doi: 10.1016/0092-8674(83)90284-2. [DOI] [PubMed] [Google Scholar]
  2. Brash D. E., Franklin W. A., Sancar G. B., Sancar A., Haseltine W. A. Escherichia coli DNA photolyase reverses cyclobutane pyrimidine dimers but not pyrimidine-pyrimidone (6-4) photoproducts. J Biol Chem. 1985 Sep 25;260(21):11438–11441. [PubMed] [Google Scholar]
  3. Brash D. E., Haseltine W. A. UV-induced mutation hotspots occur at DNA damage hotspots. Nature. 1982 Jul 8;298(5870):189–192. doi: 10.1038/298189a0. [DOI] [PubMed] [Google Scholar]
  4. Calos M. P., Lebkowski J. S., Botchan M. R. High mutation frequency in DNA transfected into mammalian cells. Proc Natl Acad Sci U S A. 1983 May;80(10):3015–3019. doi: 10.1073/pnas.80.10.3015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Cornelis J. J., Su Z. Z., Ward D. C., Rommelaere J. Indirect induction of mutagenesis of intact parvovirus H-1 in mammalian cells treated with UV light or with UV-irradiated H-1 or simian virus 40. Proc Natl Acad Sci U S A. 1981 Jul;78(7):4480–4484. doi: 10.1073/pnas.78.7.4480. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Franklin W. A., Doetsch P. W., Haseltine W. A. Structural determination of the ultraviolet light-induced thymine-cytosine pyrimidine-pyrimidone (6-4) photoproduct. Nucleic Acids Res. 1985 Jul 25;13(14):5317–5325. doi: 10.1093/nar/13.14.5317. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hall J. D., Mount D. W. Mechanisms of DNA replication and mutagenesis in ultraviolet-irradiated bacteria and mammalian cells. Prog Nucleic Acid Res Mol Biol. 1981;25:53–126. doi: 10.1016/s0079-6603(08)60483-3. [DOI] [PubMed] [Google Scholar]
  8. Hauser J., Seidman M. M., Sidur K., Dixon K. Sequence specificity of point mutations induced during passage of a UV-irradiated shuttle vector plasmid in monkey cells. Mol Cell Biol. 1986 Jan;6(1):277–285. doi: 10.1128/mcb.6.1.277. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  10. Kunz B. A., Glickman B. W. The role of pyrimidine dimers as premutagenic lesions: a study of targeted vs. untargeted mutagenesis in the lacI gene of Escherichia coli. Genetics. 1984 Mar;106(3):347–364. doi: 10.1093/genetics/106.3.347. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lawrence C. W., Christensen R. B., Christensen J. R. Identity of the photoproduct that causes lacI mutations in UV-irradiated Escherichia coli. J Bacteriol. 1985 Feb;161(2):767–768. doi: 10.1128/jb.161.2.767-768.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Lebkowski J. S., Clancy S., Miller J. H., Calos M. P. The lacI shuttle: rapid analysis of the mutagenic specificity of ultraviolet light in human cells. Proc Natl Acad Sci U S A. 1985 Dec;82(24):8606–8610. doi: 10.1073/pnas.82.24.8606. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lebkowski J. S., DuBridge R. B., Antell E. A., Greisen K. S., Calos M. P. Transfected DNA is mutated in monkey, mouse, and human cells. Mol Cell Biol. 1984 Oct;4(10):1951–1960. doi: 10.1128/mcb.4.10.1951. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Loeb L. A. Apurinic sites as mutagenic intermediates. Cell. 1985 Mar;40(3):483–484. doi: 10.1016/0092-8674(85)90191-6. [DOI] [PubMed] [Google Scholar]
  15. Loeb L. A., Kunkel T. A. Fidelity of DNA synthesis. Annu Rev Biochem. 1982;51:429–457. doi: 10.1146/annurev.bi.51.070182.002241. [DOI] [PubMed] [Google Scholar]
  16. McCutchan J. H., Pagano J. S. Enchancement of the infectivity of simian virus 40 deoxyribonucleic acid with diethylaminoethyl-dextran. J Natl Cancer Inst. 1968 Aug;41(2):351–357. [PubMed] [Google Scholar]
  17. Miller J. H., Lebkowski J. S., Greisen K. S., Calos M. P. Specificity of mutations induced in transfected DNA by mammalian cells. EMBO J. 1984 Dec 20;3(13):3117–3121. doi: 10.1002/j.1460-2075.1984.tb02267.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Peden K. W., Pipas J. M., Pearson-White S., Nathans D. Isolation of mutants of an animal virus in bacteria. Science. 1980 Sep 19;209(4463):1392–1396. doi: 10.1126/science.6251547. [DOI] [PubMed] [Google Scholar]
  19. Protić-Sabljić M., Kraemer K. H. One pyrimidine dimer inactivates expression of a transfected gene in xeroderma pigmentosum cells. Proc Natl Acad Sci U S A. 1985 Oct;82(19):6622–6626. doi: 10.1073/pnas.82.19.6622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Protić-Sabljić M., Kraemer K. H. Reduced repair of non-dimer photoproducts in a gene transfected into xeroderma pigmentosum cells. Photochem Photobiol. 1986 May;43(5):509–513. doi: 10.1111/j.1751-1097.1986.tb09528.x. [DOI] [PubMed] [Google Scholar]
  21. Rabkin S. D., Moore P. D., Strauss B. S. In vitro bypass of UV-induced lesions by Escherichia coli DNA polymerase I: specificity of nucleotide incorporation. Proc Natl Acad Sci U S A. 1983 Mar;80(6):1541–1545. doi: 10.1073/pnas.80.6.1541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Razzaque A., Chakrabarti S., Joffee S., Seidman M. Mutagenesis of a shuttle vector plasmid in mammalian cells. Mol Cell Biol. 1984 Mar;4(3):435–441. doi: 10.1128/mcb.4.3.435. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Razzaque A., Mizusawa H., Seidman M. M. Rearrangement and mutagenesis of a shuttle vector plasmid after passage in mammalian cells. Proc Natl Acad Sci U S A. 1983 May;80(10):3010–3014. doi: 10.1073/pnas.80.10.3010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Robb J. A., Huebner K. Effect of cell chromosome number on simian virus 40 replication. Exp Cell Res. 1973 Sep;81(1):120–126. doi: 10.1016/0014-4827(73)90118-3. [DOI] [PubMed] [Google Scholar]
  25. Sagher D., Strauss B. Abasic sites from cytosine as termination signals for DNA synthesis. Nucleic Acids Res. 1985 Jun 25;13(12):4285–4298. doi: 10.1093/nar/13.12.4285. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Sancar A., Smith F. W., Sancar G. B. Purification of Escherichia coli DNA photolyase. J Biol Chem. 1984 May 10;259(9):6028–6032. [PubMed] [Google Scholar]
  27. Santos E., Reddy E. P., Pulciani S., Feldmann R. J., Barbacid M. Spontaneous activation of a human proto-oncogene. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4679–4683. doi: 10.1073/pnas.80.15.4679. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Seidman M. M., Dixon K., Razzaque A., Zagursky R. J., Berman M. L. A shuttle vector plasmid for studying carcinogen-induced point mutations in mammalian cells. Gene. 1985;38(1-3):233–237. doi: 10.1016/0378-1119(85)90222-7. [DOI] [PubMed] [Google Scholar]
  29. Setlow R. B. The wavelengths in sunlight effective in producing skin cancer: a theoretical analysis. Proc Natl Acad Sci U S A. 1974 Sep;71(9):3363–3366. doi: 10.1073/pnas.71.9.3363. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Shapiro M. B., Senapathy P. Automated preparation of DNA sequences for publication. Nucleic Acids Res. 1986 Jan 10;14(1):65–73. doi: 10.1093/nar/14.1.65. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Witkin E. M. Ultraviolet mutagenesis and inducible DNA repair in Escherichia coli. Bacteriol Rev. 1976 Dec;40(4):869–907. doi: 10.1128/br.40.4.869-907.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Wood R. D. Pyrimidine dimers are not the principal pre-mutagenic lesions induced in lambda phage DNA by ultraviolet light. J Mol Biol. 1985 Aug 20;184(4):577–585. doi: 10.1016/0022-2836(85)90304-3. [DOI] [PubMed] [Google Scholar]
  33. Wood R. D., Skopek T. R., Hutchinson F. Changes in DNA base sequence induced by targeted mutagenesis of lambda phage by ultraviolet light. J Mol Biol. 1984 Mar 5;173(3):273–291. doi: 10.1016/0022-2836(84)90121-9. [DOI] [PubMed] [Google Scholar]

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