Expression of human DNA topoisomerase I in yeast cells lacking yeast DNA topoisomerase I: restoration of sensitivity of the cells to the antitumor drug camptothecin.
Yeast Saccharomyces cerevisiae strains that are permeable to the antitumor alkaloid camptothecin are killed by the drug if they express DNA topoisomerase I, the cellular target of the drug (J. Nitiss and J.C. Wang, Proc. Natl. Acad. Sci. USA, 85: 7501-7505, 1988). We show that in a yeast strain permeable to camptothecin but lacking DNA topoisomerase I, sensitivity to the drug was restored upon expression of human DNA topoisomerase I from a plasmid-borne human complementary DNA clone. When the human enzyme was expressed from a galactose-inducible, glucose-repressible yeast promoter, PGAL1, sensitivity to camptothecin was observed in the presence of galactose but not in the presence of glucose. Expression of human DNA topoisomerase I in Escherichia coli was also demonstrated by the complementation of a conditional lethal E. coli DNA topoisomerase I mutant. These systems can be used in the study of human DNA topoisomerase I-camptothecin interactions and in the screening of additional therapeutics targeting the enzyme.
Bjornsti
M A
MA
Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
Benedetti
P
P
Viglianti
G A
GA
Wang
J C
JC
D001483:Base Sequence; D002166:Camptothecin / Q000378:metabolism / Q000494:pharmacology*; D004264:DNA Topoisomerases, Type I / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005796:Genes; D005816:Genetic Complementation Test; D006367:HeLa Cells / Q000201:enzymology; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D010957:Plasmids; D011994:Recombinant Proteins / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*
D011994:Recombinant Proteins; D004264:DNA Topoisomerases, Type I; D002166:Camptothecin