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A temperature-sensitive mutant showing two defective functions.

T Marunouchi

    The Japanese Journal of Experimental Medicine
    |April 1, 1978
    PubMed
    Summary
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    This study characterizes a temperature-sensitive mutant, 13B11, from CHO-K1 cells. The mutant shows impaired DNA synthesis and cell division at elevated temperatures, suggesting a single mutation affects multiple cell cycle processes.

    Area of Science:

    • Cell Biology
    • Genetics
    • Molecular Biology

    Background:

    • Characterization of temperature-sensitive mutants is crucial for understanding cell cycle regulation.
    • CHO-K1 cells are a widely used model system in mammalian cell genetics.

    Purpose of the Study:

    • To partially characterize a novel temperature-sensitive growth mutant (13B11) derived from CHO-K1 cells.
    • To investigate the effects of non-permissive temperature on DNA synthesis and cell division in this mutant.

    Main Methods:

    • Temperature shift experiments to a non-permissive temperature (39°C).
    • Radioautography to assess DNA synthesis rates and cell populations.
    • Synchronization of cell cycles (G1 phase).
    • Analysis of DNA content using flow cytometry.

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    Main Results:

    • A shift to 39°C initially slows DNA synthesis, primarily by reducing the proportion of DNA-synthesizing cells, with partial recovery due to cells entering S phase.
    • Synchronized G1 cells show a 6-hour delay in entering S phase at 39°C, with late G1 processes being heat-sensitive.
    • Mutant cells can divide if shifted to 39°C during S phase, but not if shifted during G1, though some G1-shifted cells divide upon return to permissive temperature with hydroxyurea.
    • Accumulation of nuclei with nearly 4C DNA content suggests a defect in cell division processes.

    Conclusions:

    • The 13B11 mutant exhibits temperature-sensitive defects in both DNA synthesis and cell division.
    • These dual defects likely result from a single mutation, given the low frequency of spontaneous revertants.
    • This mutant provides a valuable tool for dissecting cell cycle control mechanisms in mammalian cells.