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. 1988 Apr 25;16(8):3327–3340. doi: 10.1093/nar/16.8.3327

UV-induced photoproducts of 5-methylcytosine in a DNA sequence context.

T Barna 1, J Malinowski 1, P Holton 1, M Ruchirawat 1, F F Becker 1, J N Lapeyre 1
PMCID: PMC336497  PMID: 3375057

Abstract

In order to detect possible m5C photoproducts, highly purified rat liver DNA-cytosine methyltransferase was used to specifically generate m5C with a radioactive methyl group. When these DNAs were subjected to a large dose (10 kJ/m2) of 254 nm or 302 nm ultraviolet light (UVB) to enhance the yield, two labeled photoproducts were detected and isolated by reverse phase HPLC after formic acid hydrolysis. Further studies using acetone as a triplet state sensitizer and UVB irradiation suggested that photoproduct II was activated via a triplet state while the more polar photoproduct I was not. Photoreversion of the purified photoproducts with 10 kJ/m2 254 nm light demonstrated the following reactions: Photoproduct I regenerated m5C, while photoproduct II is split and regenerated m5C and photoproduct I. These results suggest that photoproduct I is monomeric while photoproduct II dimeric, and from the latter's elution position possibly a cyclobutyl type dimer arising from a reaction with an adjacent cytosine. Using d[TTG] and d[Cm5CG] as models of typical sequences, irradiation with 10 kJ/m2 254 nm or 302 nm, respectively, gave rise to a small component having altered mobility in sequencing gels. The altered mobility trinucleotides were resistant to degradation by PI and micrococcal nucleases as expected from photodimerization of the pyrimidine bases. Furthermore, oligonucleotide substrates containing m5C were synthesized and shown to be susceptible to T4 endonuclease v action at locations consistent with d[Cm5C] photodimer formation when irradiated in the UVB range.

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Selected References

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  1. Banaszuk A. M., Deugau K. V., Sherwood J., Michalak M., Glick B. R. An efficient method for the sequence analysis of oligodeoxyribonucleotides. Anal Biochem. 1983 Feb 1;128(2):281–286. doi: 10.1016/0003-2697(83)90376-7. [DOI] [PubMed] [Google Scholar]
  2. Becker F. F., Holton P., Ruchirawat M., Lapeyre J. N. Perturbation of maintenance and de novo DNA methylation in vitro by UVB (280-340 nm)-induced pyrimidine photodimers. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6055–6059. doi: 10.1073/pnas.82.18.6055. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Browne M. J., Turnbull J. F., McKay E. L., Adams R. L., Burdon R. H. The sequence specificity of a mammalian DNA methylase. Nucleic Acids Res. 1977 Apr;4(4):1039–1045. doi: 10.1093/nar/4.4.1039. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Busslinger M., Hurst J., Flavell R. A. DNA methylation and the regulation of globin gene expression. Cell. 1983 Aug;34(1):197–206. doi: 10.1016/0092-8674(83)90150-2. [DOI] [PubMed] [Google Scholar]
  5. Christman J. K., Price P., Pedrinan L., Acs G. Correlation between hypomethylation of DNA and expression of globin genes in Friend erythroleukemia cells. Eur J Biochem. 1977 Nov 15;81(1):53–61. doi: 10.1111/j.1432-1033.1977.tb11926.x. [DOI] [PubMed] [Google Scholar]
  6. Doerfler W. DNA methylation and gene activity. Annu Rev Biochem. 1983;52:93–124. doi: 10.1146/annurev.bi.52.070183.000521. [DOI] [PubMed] [Google Scholar]
  7. Duker N. J., Merkel G. W. Inhibition of enzymic incision of thymine dimers by covalently bound 2-[N-[(deoxyguanosin-8-yl)acetyl]amino]fluorene in deoxyribonucleic acid. Biochemistry. 1985 Jan 15;24(2):408–412. doi: 10.1021/bi00323a025. [DOI] [PubMed] [Google Scholar]
  8. Ellison M. J., Childs J. D. Pyrimidine dimers induced in Escherichia coli DNA by ultraviolet radiation present in sunlight. Photochem Photobiol. 1981 Oct;34(4):465–469. [PubMed] [Google Scholar]
  9. Ford J. P., Coca-Prados M., Hsu M. T. Enzymatic analysis of 5-methylcytosine content in eukaryotic DNA. Study of intracellular Simian Virus 40 DNA. J Biol Chem. 1980 Aug 25;255(16):7544–7547. [PubMed] [Google Scholar]
  10. Franklin W. A., Lo K. M., Haseltine W. A. Alkaline lability of fluorescent photoproducts produced in ultraviolet light-irradiated DNA. J Biol Chem. 1982 Nov 25;257(22):13535–13543. [PubMed] [Google Scholar]
  11. Gallagher P. E., Duker N. J. Detection of UV purine photoproducts in a defined sequence of human DNA. Mol Cell Biol. 1986 Feb;6(2):707–709. doi: 10.1128/mcb.6.2.707. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Gill J. E. Fluorescence of 5-methylcytosine. Photochem Photobiol. 1970 Apr;11(4):259–269. doi: 10.1111/j.1751-1097.1970.tb05994.x. [DOI] [PubMed] [Google Scholar]
  13. Gordon L. K., Haseltine W. A. Comparison of the cleavage of pyrimidine dimers by the bacteriophage T4 and Micrococcus luteus UV-specific endonucleases. J Biol Chem. 1980 Dec 25;255(24):12047–12050. [PubMed] [Google Scholar]
  14. Hart R. W., Setlow R. B., Woodhead A. D. Evidence that pyrimidine dimers in DNA can give rise to tumors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5574–5578. doi: 10.1073/pnas.74.12.5574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Haseltine W. A., Gordon L. K., Lindan C. P., Grafstrom R. H., Shaper N. L., Grossman L. Cleavage of pyrimidine dimers in specific DNA sequences by a pyrimidine dimer DNA-glycosylase of M. luteus. Nature. 1980 Jun 26;285(5767):634–641. doi: 10.1038/285634a0. [DOI] [PubMed] [Google Scholar]
  16. Kastan M. B., Gowans B. J., Lieberman M. W. Methylation of deoxycytidine incorporated by excision-repair synthesis of DNA. Cell. 1982 Sep;30(2):509–516. doi: 10.1016/0092-8674(82)90248-3. [DOI] [PubMed] [Google Scholar]
  17. Keshet I., Yisraeli J., Cedar H. Effect of regional DNA methylation on gene expression. Proc Natl Acad Sci U S A. 1985 May;82(9):2560–2564. doi: 10.1073/pnas.82.9.2560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Klocker H., Auer B., Burtscher H. J., Hofmann J., Hirsch-Kauffmann M., Schweiger M. A sensitive radioimmuno assay for thymine dimers. Mol Gen Genet. 1982;186(4):475–477. doi: 10.1007/BF00337950. [DOI] [PubMed] [Google Scholar]
  19. Lamola A. A., Guéron M., Yamane T., Eisinger J., Shulman R. G. Triplet state of DNA. J Chem Phys. 1967 Oct 1;47(7):2210–2217. doi: 10.1063/1.1703293. [DOI] [PubMed] [Google Scholar]
  20. Lapeyre J. N., Becker F. F. 5-Methylcytosine content of nuclear DNA during chemical hepatocarcinogenesis and in carcinomas which result. Biochem Biophys Res Commun. 1979 Apr 13;87(3):698–705. doi: 10.1016/0006-291x(79)92015-1. [DOI] [PubMed] [Google Scholar]
  21. Lieberman M. W., Beach L. R., Palmiter R. D. Ultraviolet radiation-induced metallothionein-I gene activation is associated with extensive DNA demethylation. Cell. 1983 Nov;35(1):207–214. doi: 10.1016/0092-8674(83)90223-4. [DOI] [PubMed] [Google Scholar]
  22. Low M., Read E. L., Borek E. Methylation of DNA in HeLa cells after ultraviolet irradiation. Int J Radiat Oncol Biol Phys. 1976 Jan-Feb;1(3-4):289–294. doi: 10.1016/0360-3016(76)90053-5. [DOI] [PubMed] [Google Scholar]
  23. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  24. Meistrich M. L., Lamola A. A., Gabbay E. Sensitized photoinactivation of bacteriophage T4. Photochem Photobiol. 1970 Mar;11(3):169–178. doi: 10.1111/j.1751-1097.1970.tb05985.x. [DOI] [PubMed] [Google Scholar]
  25. Messing J., Vieira J. A new pair of M13 vectors for selecting either DNA strand of double-digest restriction fragments. Gene. 1982 Oct;19(3):269–276. doi: 10.1016/0378-1119(82)90016-6. [DOI] [PubMed] [Google Scholar]
  26. Riggs A. D., Jones P. A. 5-methylcytosine, gene regulation, and cancer. Adv Cancer Res. 1983;40:1–30. doi: 10.1016/s0065-230x(08)60678-8. [DOI] [PubMed] [Google Scholar]
  27. Ruchirawat M., Noshari J., Lapeyre J. N. Kinetic mechanisms and interaction of rat liver DNA methyltransferase with defined DNA substrates. Mol Cell Biochem. 1987 Jul;76(1):45–54. doi: 10.1007/BF00219397. [DOI] [PubMed] [Google Scholar]
  28. Simon D., Stuhlmann H., Jähner D., Wagner H., Werner E., Jaenisch R. Retrovirus genomes methylated by mammalian but not bacterial methylase are non-infectious. Nature. 1983 Jul 21;304(5923):275–277. doi: 10.1038/304275a0. [DOI] [PubMed] [Google Scholar]
  29. Vanyushin B. F., Mazin A. L., Vasilyev V. K., Belozersky A. N. The content of 5-methylcytosine in animal DNA: the species and tissue specificity. Biochim Biophys Acta. 1973 Mar 28;299(3):397–403. doi: 10.1016/0005-2787(73)90264-5. [DOI] [PubMed] [Google Scholar]
  30. van der Leun J. C. UV-carcinogenesis. Photochem Photobiol. 1984 Jun;39(6):861–868. doi: 10.1111/j.1751-1097.1984.tb08872.x. [DOI] [PubMed] [Google Scholar]

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