Alkali‐labile Photolesions Mapping to Purine Sites in Ultraviolet‐irradiated Dna

Abstract

Gel sequencing experiments with the 5'- and 3'-end-labeled oligonucleotides d(A3GA4GA5GA6GA3G) and d(AT)10 have demonstrated that dimeric adenine photoproducts and thymine-adenine photoadducts constitute alkali-labile lesions in UV-irradiated DNA. On treatment with hot piperidine, DNA strand breakage occurs predominantly at the sites of 5'-adenines in the dimeric photoproducts and of 3'-adenines in the thymine-adenine photoadducts. With 5'-end-labeled oligonucleotides of mixed sequence, major UV-induced loci for alkaline cleavage map to purine bases flanked on their 5'-side by two pyrimidines. This behavior does not arise from enhanced photoreactivity of purines in this sequence context as has been inferred from photofootprinting studies. Instead, as shown by 3'-labeling and selective substitution with 5-methylcytosine, it results from the anomalous electrophoretic mobility of 5'-end-labeled fragments produced by alkaline cleavage of DNA at adjacent pyrimidine (6-4) pyrimidone photoproducts.

Ultraviolet Radiation Damage in DNA

“…Accordingly, it was inferred [56] that purine bases in 5'.PyPyPu.3' triplets possess enhanced photoreactivity. We have subsequently shown [57], through experiments with 3'-end-labelled and 5-methylcytosine-substituted oligonucleotides, that this is not the case. The apparent cleavage at purine residues, which can also be induced by treatment with hot piperidine, is an artefact arising from the anomalous electrophoretic mobility of S'-endlabelled fragments produced by chemical cleavage of the DNA at adjacent (6-4) photoproducts.…”

Quantification of DNA damage and repair in amino acid auxotrophs and UV‐sensitive mutants of Aspergillus nidulans using an ELISA

“…UV radiation produces a variety of photo° pr. ,ducts many of which (including pyrimidine dimers and 64 ph ~toproducts) can be classified as alkali-labile lesions [19] and all lough they are chemically stable under normal physiological co,~ditions their decomposition under alkaline conditions leads to the cleavage of the DNA backbone through the formation of ~basic sites [20]. Other UV-induced DNA lesions known to ac: as alkali-labile sites include thymine glycols [21] and 8-hy lroxyguanine [22].…”

Synergistic cytotoxicity and DNA strand breaks in cells and plasmid DNA exposed to uranyl acetate and ultraviolet radiation

“…Post-treatment exposure to warm piperidine was used to analyze the extent to which any photoactivated UA-induced lesions, or UV-induced alkali-labile lesions, for example pyrimidine(6-4)pyrimidone photoproduct ([6-4]PP), Dewar isomers, adenine and thymine products, and other abasic sites, could be converted to SSB (Hejmadi et al, 1994). For reactions that received subsequent heat and piperidine exposures (Fig.…”

“…Using methods we have previously developed (Wilson et al, 2014), pBR322 plasmid DNA was exposed to DU as UA followed by exposure to UVB (302 nm) or UVA radiation (368 nm). The generation of DNA SSB was measured, and the presence of DNA adducts and AP sites were probed by posttreatment exposures to heat and piperidine, which convert these lesions to SSB (Hejmadi et al, 1994). Chinese hamster ovary (CHO) cells, either proficient in DNA repair, deficient in base excision repair (BER), or deficient in nucleotide excision repair (NER), and human keratinocyte HaCaT cells were exposed to UA and UVB radiation to measure the effect of DNA repair proficiency on cell survival.…”