Topoisomerase I (Top1) removes
DNA torsional stress by nicking and resealing one strand of
DNA, and is essential in higher eukaryotes. The
enzyme is frequently overproduced in
tumors and is the sole target of the chemotherapeutic drug
camptothecin (
CPT) and its clinical derivatives.
CPT stabilizes the covalent Top1-DNA cleavage intermediate, which leads to toxic double-strand breaks (DSBs) when encountered by a replication fork. In the current study, we examined genetic instability associated with
CPT treatment or with Top1 overexpression in the yeast Saccharomyces cerevisiae. Two types of instability were monitored: Top1-dependent deletions in haploid strains, which do not require processing into a
DSB, and instability at the repetitive
ribosomal DNA (
rDNA) locus in diploid strains, which reflects
DSB formation. Three 2-bp deletion hotspots were examined and mutations at each were elevated either when a wild-type strain was treated with
CPT or when TOP1 was overexpressed, with the mutation frequency correlating with the level of TOP1 overexpression. Under both conditions, deletions at novel positions were enriched.
rDNA stability was examined by measuring loss-of-heterozygosity and as was observed previously upon
CPT treatment of a wild-type strain, Top1 overexpression destabilized
rDNA. We conclude that too much, as well as too little of Top1 is detrimental to eukaryotic genomes, and that
CPT has destabilizing effects that extend beyond those associated with
DSB formation.