Wavelength dependence of ultraviolet-induced DNA damage distribution: involvement of direct or indirect mechanisms and possible artefacts.
DNA damage profiles have been established in plasmid DNA using purified DNA repair enzymes and a plasmid relaxation assay, following exposure to UVC, UVB, UVA or simulated sunlight (SSL). Cyclobutane pyrimidine dimers (CPDs) are revealed as T4 endonuclease V-sensitive sites, oxidation products at purine and pyrimidine as Fpg- and Nth-sensitive sites, and abasic sites are detected by Nfo protein from Escherichia coli. CPDs are readily detected after UVA exposure, though produced 10(3) and 10(5) times less efficiently than by UVB or UVC, respectively. We demonstrate that CPDs are induced by UVA radiation and not by contaminating UVB wavelengths. Furthermore, they are produced at doses compatible with human exposure and are likely to contribute to the mutagenic specificity of UVA [E. Sage et al., Proc. Natl. Acad. Sci. USA 93 (1996) 176-180]. Oxidative damage is induced with a linear dose dependence, for each region of the solar spectrum, with the exception of oxidized pyrimidine and abasic sites, which are not detectable after UVB irradiation. The distribution of the different classes of photolesions varies markedly, depending on wavelengths. However, the unexpectedly high yield of oxidative lesions, as compared to CPDs, by UVA and SSL led us to investigate their production mechanism. An artificial formation of hydroxyl radicals is observed, which depends on the material of the sample holder used for UVA irradiation and is specific for long UV wavelengths. Our study sheds light on a possible artefact in the production of oxidative damage by UVA radiation. Meanwhile, after eliminating some potential sources of the artefact ratios of CPDs to oxidized purine of three and five upon irradiation with UVA and SSL, respectively, are still observed, whereas these ratios are about 140 and 200 after UVC and UVB irradiation.
Kuluncsics
Z
Z
CNRS UMR 218, CEA LRC-1, Institut Curie, Paris, France.
Perdiz
D
D
Brulay
E
E
Muel
B
B
Sage
E
E
D016477:Artifacts; D017122:Bacteriophage T4 / Q000201:enzymology; D019757:Carbon-Oxygen Lyases / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase; D043224:Deoxyribonuclease (Pyrimidine Dimer); D043223:Deoxyribonuclease IV (Phage T4-Induced); D004706:Endodeoxyribonucleases / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D029968:Escherichia coli Proteins; D006801:Humans; D017665:Hydroxyl Radical / Q000032:analysis; D016296:Mutagenesis; D010957:Plasmids / Q000528:radiation effects*; D013050:Spectrometry, Fluorescence / Q000379:methods; D013472:Sunlight; D014466:Ultraviolet Rays; D014764:Viral Proteins
D029968:Escherichia coli Proteins; D014764:Viral Proteins; D017665:Hydroxyl Radical; D004706:Endodeoxyribonucleases; D043223:Deoxyribonuclease IV (Phage T4-Induced); C468972:endonuclease IV, E coli; C470698:endonuclease V, phage T4; D043224:Deoxyribonuclease (Pyrimidine Dimer); C106403:NTH protein, E coli; D019757:Carbon-Oxygen Lyases; D043603:DNA-(Apurinic or Apyrimidinic Site) Lyase