Incorporation of two deoxycytidine oxidation products into cellular DNA

Abstract

The oxidation of cytosine in DNA by free radicals and other oxidants leads to an assortment of products including pyrimidine ring 5,6-saturated, 5,6-unsaturated, contraction, and fragmentation products. The formation of these products in cellular DNA may explain in part the preponderance of C to T transitions induced spontaneously and by H2O2 or ionizing radiation. Our studies have focused on the biological effects of two major 5,6-unsaturated oxidation products of cytosine: 5- hydroxycytosine and 5-hydroxyuracil. In the present work, we have attempted to study the repair of these two lesions by specifically incorporating them into cellular DNA upon incubation of cells with 5-hydroxy-2'deoxycytidine and 5-hydroxy-2'-deoxyuridine. Incubation of mouse L1210 cells with 250 M 5-hydroxy-2'-deoxycytidine led to the incorporation of this lesion to a level 20 times higher (43 lesions/10(5) cytosines) than base-line levels; however, there was no evidence for its repair following a 15-h chase. In contrast, we did not observe any significant incorporation of 5-hydroxy-2'-deoxyuridine into the DNA of L1210 cells but did observe an unidentified product, presumably an oxidation product. This unidentified pyrimidine was incorporated at a very high level (about 2000 lesions/10(5) cytosine residues) and then partially repaired in chase experiments.