Complementation of a yeast top2ts mutation by a cDNA encoding rat DNA topoisomerase II alpha.
A series of yeast expression plasmids which comprise segments of the cDNA sequences encoding rat topo II alpha have been constructed. The transcription of these constructs is under the control of the yeast GAL1 promoter. Galactose-dependent expression of the cloned rat topo II alpha cDNA complemented a yeast top2ts mutation, as well as a deletion mutation at the yeast TOP2 locus. Truncation of 12 N-terminal amino acids and/or 158 C-terminal amino acids of rat topo II alpha had no effect on its ability functionally to substitute for top2ts. Moreover, a cDNA construct with mutated putative leucine zipper domain (amino acids 993-1013) retained the complementation activity. These observations suggest that transformants capable of conditional topo II alpha expression can be exploited as a useful model system for studies on the structure-function relationships of wild-type and mutated topo II alpha, as well as the interplay of potential antitumor drugs with the enzyme.
Yoon
J H
JH
Department of Molecular Biology, Seoul National University, Republic of Korea.
Park
S H
SH
Cho
H A
HA
Seong
R H
RH
Hong
S H
SH
Park
S D
SD
D000818:Animals; D000951:Antigens, Neoplasm; D015139:Blotting, Southern; D004250:DNA Topoisomerases, Type II / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D018076:DNA, Complementary / Q000235:genetics; D004268:DNA-Binding Proteins; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D016350:Leucine Zippers; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010641:Phenotype; D011401:Promoter Regions, Genetic; D051381:Rats; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*; D017384:Sequence Deletion; D013696:Temperature; D014170:Transformation, Genetic
D000951:Antigens, Neoplasm; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D007527:Isoenzymes; D004250:DNA Topoisomerases, Type II