The budding yeast Saccharomyces cerevisiae has been exploited to investigate the cytotoxic mechanisms of drugs that target DNA topoisomerases. This model organism has been used to establish eukaryotic DNA topoisomerase I or II as the cellular target of specific antineoplastic agents, to define mutations in these enzymes that confer drug resistance and to elucidate the cellular factors that modulate cell sensitivity to DNA topoisomerase-targeted drugs. These findings have provided valuable insights into the critical activities of these enzymes and how perturbing their functions produces DNA damage and cell death.