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A selection system specific for the Thy mutator phenotype

M Meuth, O Gonçalves, P Thom

    Somatic Cell Genetics
    |July 1, 1982
    PubMed
    Summary
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    Thy- mutants exhibit resistance to arabinosyl cytosine (araC) and 5-fluorouracil (5FU). These mutants show altered nucleotide levels and increased mutation rates, suggesting a single mutation causes the thymidine auxotrophy phenotype in Chinese hamster ovary cells.

    Area of Science:

    • Biochemistry
    • Genetics
    • Cell Biology

    Background:

    • Thy- mutants display resistance to arabinosyl cytosine (araC) and cross-resistance to 5-fluorouracil (5FU).
    • Understanding the genetic basis of drug resistance in mammalian cells is crucial for developing targeted therapies.

    Purpose of the Study:

    • To characterize Chinese hamster ovary (CHO) cell clones resistant to both araC and 5FU.
    • To determine if these resistant clones are identical to thy- mutants and to investigate the underlying genetic and biochemical alterations.

    Main Methods:

    • Exposure of CHO cells to a selection system using both araC and 5FU.
    • Analysis of resistant clones for thymidine auxotrophy, nucleotide pool levels (dCTP, dTTP, dATP), and spontaneous mutation rates.
    • Genetic characterization to assess reversion frequency and mutation rates for independent markers.

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    Main Results:

    • Isolated resistant clones were identical to thy- mutants, exhibiting auxotrophy for thymidine with high reversion frequencies.
    • Resistant cells showed elevated dCTP levels, while dTTP and dATP levels remained unchanged compared to wild-type cells.
    • All tested resistant clones displayed a 7- to 50-fold higher spontaneous mutation rate than wild-type CHO cells.

    Conclusions:

    • The thy- phenotype in CHO cells is likely caused by a single mutation.
    • The combined resistance to araC and 5FU, altered nucleotide pools, and increased mutation rate are characteristic of thy- mutants.