Characterization of a mammalian mutant with a camptothecin-resistant DNA topoisomerase I

DNA topoisomerase I was purified to near homogeneity from a clonal line of human lymphoblastic leukemia cells, RPMI 8402, that is resistant to camptothecin, a cytotoxic alkaloid from Camptotheca acuminata, and compared with that of the parent wild-type cells. As assayed by relaxation of the supercoi...

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Bibliographic Details
Published in:Proceedings of the National Academy of Sciences - PNAS 1987-08, Vol.84 (16), p.5565-5569
Main Authors: Andoh, T, Ishii, K, Suzuki, Y, Ikegami, Y, Kusunoki, Y, Takemoto, Y, Okada, K
Format: Article
Language:English
Subjects:
acute lymphatic leukemia
ALCALOIDE
ALCALOIDES
ALKALOIDS
ANTI-TUMOR AGENTS
antitumor agents
Biological and medical sciences
CAMPTOTHECA ACUMINATA
camptothecin
Camptothecin - pharmacology
Cell growth
Cell Line
Cell lines
Cell physiology
CELLS
CELLULE
CELULAS
Chromatin
DNA
DNA topoisomerase
DNA Topoisomerases, Type I - metabolism
DNA, Superhelical - drug effects
Drug Resistance
ENZIMAS
ENZYME
ENZYMES
Fundamental and applied biological sciences. Psychology
Gels
HeLa cells
Humans
Immunosorbent Techniques
Leukemia, Lymphoid - enzymology
MAMIFEROS
MAMMALS
MAMMIFERE
man
Miscellaneous
Molecular and cellular biology
MUTANT
MUTANTES
MUTANTS
Mutation
Nucleic Acid Conformation - drug effects
Nucleic acids
Peptide Mapping
PLANT ALKALOIDS
Plasmids
RESISTANCE AUX FACTEURS NUISIBLES
RESISTANCE TO INJURIOUS FACTORS
RESISTENCIA A AGENTES DANINOS
RNA
TOXICIDAD
TOXICITE
TOXICITY
UMBELLIFLORAE
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Summary:DNA topoisomerase I was purified to near homogeneity from a clonal line of human lymphoblastic leukemia cells, RPMI 8402, that is resistant to camptothecin, a cytotoxic alkaloid from Camptotheca acuminata, and compared with that of the parent wild-type cells. As assayed by relaxation of the supercoiled plasmid DNA and by formation of enzyme-linked DNA breaks, the purified enzyme from the resistant cells was shown to be >125-fold as resistant to camptothecin as the wild-type enzyme, comparable to a cellular resistance index of about 300. Therefore, the cellular resistance appears to be due to the resistance of the enzyme. The amount of the immunoreactive enzyme protein in whole extract appeared to be reduced to less than half that of the wild-type enzyme. These results establish that DNA topoisomerase I is the cellular target of camptothecin and that DNA topoisomerase I is essential for the survival of mammalian cells.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.84.16.5565