ABSTRACT
Low level cytotoxic stress greatly accelerates the loss of unstably amplified dihydrofolate reductase (dhfr) genes from methotrexate-resistant mouse cell lines. To understand this drug-induced loss of amplified genes, the highly methotrexate-resistant mouse R500 cell line was flow sorted into two subpopulations with higher and lower average dhfr gene dosage respectively. The subpopulation with higher levels of gene amplification was much more sensitive to low level cytotoxic stress as judged by both cloning efficiency and growth in the presence of low concentrations of cytotoxic drugs. These results suggest that high levels of gene amplification can confer hypersensitivity to cytotoxic stressors such as anticancer drugs.
Subject(s)
Hypersensitivity/genetics , Stress, Physiological/genetics , Tetrahydrofolate Dehydrogenase/genetics , Animals , Cell Division , Cell Separation , Cells, Cultured , Clone Cells/physiology , Drug Resistance , Flow Cytometry , Gene Amplification/genetics , Hypersensitivity/enzymology , Methotrexate/pharmacology , Mice , Stress, Physiological/enzymologyABSTRACT
Caffeine was found to inhibit both type I and type II topoisomerases in vivo as judged by its effects on replicating simian virus 40 (SV40) chromosomes. The study was facilitated by the use of a rapid filter assay for the detection and characterization of topoisomerase inhibitors. The assay, which requires neither purified enzymes nor substrates, was able to identify both antagonists and poisons of type I and type II topoisomerases.
Subject(s)
Caffeine/pharmacology , Topoisomerase I Inhibitors , Topoisomerase II Inhibitors , Animals , Camptothecin/pharmacology , Etoposide/pharmacology , Haplorhini , Teniposide/pharmacologyABSTRACT
We have found that type II topoisomerase inhibitors have two effects on replicating simian virus 40 genomes in vivo: production of catenated dimers and slowed replication of the last 5% of the genome. This suggests that type II topoisomerase simultaneously decatenates and facilitates replication fork movement at this stage of DNA replication. On the basis of this observation, a detailed model is proposed for the roles of topoisomerases I and II in simian virus 40 DNA replication.
