Oxidative DNA base damage induced by singlet oxygen and photosensitization: recognition by repair endonucleases and mutagenicity

We have analyzed the recognition by various repair endonucleases of DNA bas e modifications induced by three oxidants, viz. [4-(tert-butyldioxycarbonyl )benzyl] triethylammonium chloride (BCBT), a photo chemical source of tert- butoxyl radicals, disodium salt of 1,4-etheno-2,3-benzodioxin-1,4-dipropano ic acid (NDPO2), a chemical source of singlet oxygen, and riboflavin, a typ e-I photosensitizer. The base modifications induced by BCBT, which were pre viously shown to be mostly 7,8-dihydro-8-oxoguanine (8-oxoGua) residues, we re recognized by Fpg and Ogg1 proteins, but not by endonuclease IIII, Ntg1 and Ntg2 proteins. In the case of singlet oxygen induced damage, 8-oxoGua a ccounted for only 35% of the base modifications recognized by Fpg protein. The remaining Fpg-sensitive modifications were not recognized by Ogg1 prote in and relatively poor by endonuclease III, but they were relatively good s ubstrates of Ntg1 and Ntg2. In the case of the damage induced by photoexcit ed riboflavin, the fraction of Fpg-sensitive base modifications identified as 8-oxoGua was only 23%. In contrast to the damage induced by singlet oxyg en, the remaining lesions were not only recognized by Ntg1 and Ntg2 protein s and (relatively poor) by endonuclease III, but also by Ogg1 protein. The analysis of the mutations observed after transfection of modified plasmid p SV2gpt into Escherichia coli revealed that all agents induced near exclusiv ely GC --> TA and GC --> CG transversions, the numbers of which were correl ated with the numbers of 8-oxoGua residues and Ntg-sensitive modifications, respectively. In conclusion, both singlet oxygen and the type-I photosensi tizer riboflavin induce predominantly oxidative guanine modifications other than 8-oxoGua, which most probably give rise to GC --> CG transversions an d in which eukaryotic cells are substrates of Ntg1 and Ntg2 proteins. (C) 2 000 Elsevier Science B.V. All rights reserved.