Distribution of Deoxyribonucleic Acid Repair Synthesis Among Repetitive and Unique Sequences in the Human Diploid Genome

DNA repair synthesis was studied in contact-inhibited (non-S-phase) human diploid fibroblasts (WI-38) after damage primarily to pyrimidine bases (ultraviolet radiation, 254 nm) or purine bases (N-acetoxy-2-acetylaminofluorene or 7-bromomethylbenz[a]anthracene). The distribution of repair synthesis among sequences of different degrees of repetitiveness was examined by following the reassociation kinetics of sheared (450 nucleotide length), denatured DNAs labeled with [³H]thymidine during repair synthesis. DNAs labeled during replicative synthesis served as standards. The kinetics of reassociation for these DNAs were identical between C₀t 0.01 and 10,000 (mol sec)/1. and suggested that families of sequences of all degrees of repetitiveness are repairable or partially repairable after either type of damage. Reassociated DNAs were subjected to thermal elution chromatography and found to have similar elution profiles. These data suggested that the similarity of the reassociation kinetics was not the result of artifacts such as the failure of hydroxylapatite to distinguish between poorly matched and well-matched sequences. Experiments in which cells were damaged over a 100-fold dose range with N-acetoxy-2-acetylaminofluorene (1-100 μM) and reassociated to C₀t 100 revealed at most only a small shift in the distribution of repair synthesis from repeated to unique sequences. Calculations of the amount of damage at the doses used and its distribution among the DNA fragments used for these studies demonstrated that no interference with strand reassociation would be expected in these experiments. The results of the present study suggest that DNA damage in human cells, involving either purines or pyrimidines, is repairable to about the same extent in both unique and repetitive sequences.