DNA lesions, chromosomal aberrations and G2 delay in CHO cells cultured in medium containing bromo- or...

Experiments have been performed to investigate whether BrdUrd- and CldUrd-substituted DNA contains lesions causing a delay in cell-cycle progression and induction of chromosomal aberrations. The presence of lesions has been determined directly by alkaline sucrose gradient and nucleoid sedimentation analysis and indirectly by screening for induced chromosomal aberrations. The influence of inhibitors of DNA repair (caffeine and 3-aminobenzamide) or DNA synthesis (hydroxyurea) on the frequencies of such aberrations has been estimated. It is found that BrdUrd and CldUrd are cytotoxic when present in DNA. No randomly located DNA breaks could be detected under neutral conditions, but BrdUrd-substituted DNA was found to contain numerous alkali-labile sites. CldUrd at high concentrations causes G2 delay, similar to the action of known DNA-damaging agents. The extent of delay depends on the pattern of incorporation of the analogue, i.e., incorporation for two cell cycles causes the longest delay, growth for 12 h in CldUrd followed by 12 h in dThd-containing medium causes a lesser delay and the delay is not significant when the cells are incubated in the analogue for only 12 h prior to fixation. Numerous chromatid type aberrations are present in cells incubated at the highest CldUrd concentration, and their induction follows the pattern of induction of G2 delay, indicating the sharing of a common lesion. Caffeine, 3-aminobenzamide and hydroxyurea increase the number of chromosomal aberrations when added 2 h before fixation. The significance of these results is discussed.