title: Differential susceptibility to carcinogen-induced amplification of SV40 and dhfr sequences in SV40-transformed human keratinocytes.
abstract: Gene amplification contributes to carcinogenesis by enhancing proto-oncogene activity and causing chromosomal instability. The ease of detecting amplified tumor-virus sequences has encouraged use of this system as a surrogate for studying the molecular events involved in endogenous gene amplification. We report here a new system for studying carcinogen-induced amplification of both endogenous and viral sequences in the SV40-transformed human keratinocyte line AG06. Treatment with carcinogens induced a transient dose-dependent amplification of the integrated SV40 sequences. The amplified sequences appeared in the extrachromosomal fraction. Treatment of these cells with carcinogens prior to methotrexate (MTX) selection increased the frequency of MTX-resistant colonies, 67% of which exhibited dihydrofolate reductase (dhfr) amplification. The abilities of five carcinogens with different DNA-damaging activities (the DNA-damaging agents N-methyl-N-nitro-N-nitrosoguanidine, mitomycin C (MMC), ultraviolet light C, and X-rays and the non-DNA-damaging agent arsenite) to induce SV40 and dhfr amplification at concentrations that result in 50% clonal survival were compared. All four DNA-damaging carcinogens (as well as growth arrest) were able to elicit some SV40 amplification, but responses varied markedly, from 1.8-fold for X-rays to sevenfold to eightfold for MMC. There was no correlation between the ability to elicit the two amplification responses. Arsenite, which did not induce SV40 amplification, was the best inducer of MTX resistance. These results point to different controls involved in the induction of viral and dhfr amplification. The signal for amplification of viral genes may be triggered by DNA damage and growth arrest, whereas amplification of dhfr, and perhaps other endogenous sequences, seems to be triggered by other signals as well.
authors.affiliation: Nelson Institute of Environmental Medicine, NYU Medical Center, New York 10016.
authors.forename: T G
authors.initials: TG
authors.lastname: Rossman
authors.forename: D
authors.initials: D
authors.lastname: Wolosin
mesh_terms: D016590:Aphidicolin / Q000494:pharmacology; D002273:Carcinogens / Q000494:pharmacology*; D002461:Cell Line, Transformed; D002472:Cell Transformation, Viral / Q000187:drug effects*; D003513:Cycloheximide / Q000494:pharmacology; D004351:Drug Resistance / Q000235:genetics; D005784:Gene Amplification / Q000187:drug effects* / Q000528:radiation effects; D005814:Genes, Viral; D006801:Humans; D006918:Hydroxyurea / Q000494:pharmacology; D015603:Keratinocytes / Q000382:microbiology*; D008727:Methotrexate / Q000494:pharmacology; D016685:Mitomycin / Q000494:pharmacology; D000090063:Proto-Oncogene Mas; D013539:Simian virus 40 / Q000235:genetics*; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D013997:Time Factors; D014466:Ultraviolet Rays / Q000009:adverse effects; D014965:X-Rays / Q000009:adverse effects
chemical_list: D002273:Carcinogens; C000715470:MAS1 protein, human; D000090063:Proto-Oncogene Mas; D016590:Aphidicolin; D016685:Mitomycin; D003513:Cycloheximide; D013762:Tetrahydrofolate Dehydrogenase; D006918:Hydroxyurea; D008727:Methotrexate