We have examined the roles of eukaryotic
DNA topoisomerases I and II in DNA replication by the use of a set of four isogenic strains of Saccharomyces cerevisiae that are TOP1+ TOP2+, TOP1+ top2 ts, delta top1 TOP2+, and delta top1 top2 ts. Cells synchronized by treatment with the
alpha-mating factor, or by cycles of feeding and
starvation, were released from cell-cycle arrest, and the size distribution of
DNA chains that were synthesized after the cells reentered the S-phase was determined as a function of time. The results indicate that synthesis of short
DNA chains several thousand
nucleotides in length can initiate in the absence of both topoisomerases, but their further elongation requires at least one of the two topoisomerases. Inactivation of
DNA topoisomerase II does not alter significantly the time dependence of the patterns of nascent
DNA chain synthesis, which is consistent with the notion that the requirement of this
enzyme for viability is due to its essential role during mitosis, when pairs of intertwined newly replicated chromosomes are being segregated. The absence of
DNA topoisomerase I leads to a temporary delay in the extension of the short
DNA chains; this delay in chain elongation is also reflected in the rate of total
DNA synthesis in the delta top1 mutant during the early S-phase. Thus, in wild-type cells,
DNA topoisomerase I is probably the major replication swivel. The patterns of
DNA synthesis in asynchronously grown delta top1 top2 ts cells at permissive and non-permissive temperatures are also consistent with the above conclusions.