Topoisomerase I (Top1) is the specific target of the anticancer drug camptothecin (CPT) that interferes with enzyme activity promoting Top1-mediated DNA breaks and inhibition of DNA and RNA synthesis. To define the specific transcriptional response to CPT, we have determined the CPT-altered transcription profiles in yeast by using a relatively low concentration of the drug. CPT could alter global expression profiles only if a catalytically active Top1p was expressed in the cell, demonstrating that drug interference with Top1 was the sole trigger of the response. A total of 95 genes showed a statistically-significant alterations. Gene Ontology term analyses suggested that the cell response was mainly to the inhibition of nucleic acid synthesis and cell cycle progression. Promoter sequence analyses of the 22 up-regulated genes and expression studies in gene-deleted strains showed that the transcription factors, Swi4p and Mbp1p, mediate at least partially the transcriptional response to CPT. The MBP1 gene deletion abrogates a transient cell growth delay caused by CPT whereas the SWI4 gene deletion increases yeast resistance to CPT. Thus, the findings show that yeast cells have a highly selective and sensitive transcriptional response to CPT depending on SWI4 and MBP1 genes suggesting a complex regulation of cell cycle progression by the two factors in the presence of CPT.