INDUCTION KINETICS OF MUTATIONS AT 2 GENETIC-LOCI, DNA DAMAGE AND REPAIR IN CHO CELLS AFTER DIFFERENT EXPOSURE TIMES TO N-ETHYL-N-NITROSOUREA

Ouabain resistance (ouar) and 6-thioguanine resistance (6-TGr) mutation frequencies were measured in Chinese hamster ovary cells after treatment with N-ethyl-N-nitrosourea (ENU) for varying periods of time. Maximal mutation frequency at the Na+/K+ ATPase genes locus (ouar mutations) was attained within 5 min of exposure, whereas the mutation frequency at the hypoxanthine guanine phosphoribosyltransferase locus (6-TGr mutations) continued to increase up to 60 min, following the theoretical curve for exponential decay of ENU with time. Deletion of DNA single strand breaks (ssb) by alkaline elution showed that maximal levels were attained within 5 min of treatment with ENU. Fast repair of DNA ssb occurred early after exposure (> 50% repair within 10 min). Analysis of DNA ethylation products by h.p.l.c. showed initially rapid removal of O2-ethylcytosine (25% in the first hour), slow removal of 7-ethylguanine, 3-ethyladenine and 3-ethylguanine and no removal at all of O6-ethylguanine, O4-ethylthymine and ethylphosphotriesters. These time-course studies reveal different target gene responses in the fixation of DNA damage into mutations.