title: Temporal order of replication of genes responsible for hypoxanthine phosphoribosyl transferase and Na+/K+ ATPase in chemically transformed human fibroblasts.
abstract: The cytotoxic and mutagenic effects of a direct perturbation of DNA during various portions of the DNA synthetic period (S phase) of a chemically induced, transformed line (Hut-11A cells) derived from diploid human skin fibroblasts were examined. The cells were synchronized by a period of growth in low serum with a subsequent blockage of the cells at the G1/S boundary by hydroxyurea. This method resulted in over 90% synchrony, although approximately 20% of the cells were noncycling. Synchronized cells were treated for each of four 2-h periods during the S phase with 5-bromodeoxyuridine (BrdU) followed by irradiation with near-ultraviolet (UV). The BrdU-plus-irradiation treatment was cytotoxic and mutagenic, while treatment with BrdU alone or irradiation alone was neither cytotoxic nor mutagenic. The cytotoxicity was dependent upon the periods of S phase during which treatment was administered. The highest lethality was observed for treatment in early to middle S phase, particularly in the first 2 h of S phase, whereas scare lethality was observed in late S phase. The BrdU-plus-irradiation treatment induced ouabain- and 6-thioguanine-resistant mutants, while BrdU alone or irradiation alone was not mutagenic. Ouabain-resistant mutants were induced during early S phase by the BrdU-plus-irradiation treatment. 6-Thioguanine-resistant mutants, however, were induced during middle to late S phase. These results suggest that a certain region or regions in the DNA of Hut-11A cells, as designated by their specific temporal relationship in the S phase, may be more sensitive to the DNA perturbation by BrdU treatment plus near-UV irradiation for cell survival and that gene(s) responsible for Na+/K+ ATPase is replicated during early S phase and gene(s) for hypoxanthine phosphoribosyl transferase is replicated during middle to late S phase.
mesh_terms: D001973:Bromodeoxyuridine / Q000494:pharmacology; D002453:Cell Cycle; D002470:Cell Survival / Q000187:drug effects / Q000528:radiation effects; D004261:DNA Replication; D005347:Fibroblasts / Q000201:enzymology*; D006801:Humans; D006918:Hydroxyurea / Q000378:metabolism; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D009154:Mutation; D010042:Ouabain / Q000494:pharmacology; D010507:Periodicity; D000254:Sodium-Potassium-Exchanging ATPase / Q000235:genetics*; D014466:Ultraviolet Rays
chemical_list: D010042:Ouabain; D007041:Hypoxanthine Phosphoribosyltransferase; D000254:Sodium-Potassium-Exchanging ATPase; D001973:Bromodeoxyuridine; D006918:Hydroxyurea