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Aphidicolin hypersensitive mutant of chinese hamster V79 fibroblasts that underproduces DNA polymerase-α antigen

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Somatic Cell and Molecular Genetics

Abstract

Aphidicolin is a specific inhibitor of DNA polymerase-α and -δ from eukaryotic cells. Because of the specificity of this inhibitor, it is potentially a useful probe for the detailed studies of the function of these polymerases. DNA polymerase-α mutants isolated on the basis of resistance to aphidicolin have been described. We have isolated four variants that exhibit hypersensitivities to aphidicolin (Aphhs) from Chinese hamster V79/743X fibroblasts. These variants are designated aphhs-1, aphhs-2, aphhs-3 and aphhs-4. We reported here results of studies involving immunochemical characterization. The Aphhs phenotype in all mutants was stable for at least 30 days in the absence of selection pressure. The dCTP pools in the 743X and Aphhs cell lines were not significantly different. The level of total DNA polymerase activity in the crude extract from aphhs-2 cells was 30% of that observed in the parental 743X clone. We developed a method to quantitate DNA polymerase-α antigen at single cells in situ using monoclonal antibody SJK 132-20 and fluorescence pseudocolor image. We found that the antigen of DNA polymerase-α in aphhs-2 was 30–50% of that in the parental 743X cells. The underproduction of the antigen of DNA polymerase-α provides a basis for the observed Aphhs phenotype. Possible mechanisms for the underproduction of DNA polymerase-α in aphhs-2 clone are presented.

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Authors and Affiliations

  1. Department of Environmental Health Sciences, Case Western Reserve University, School of Medicine, 44106, Cleveland, Ohio

    Philip K. Liu, Brian Goudreau & Griffith S. Hsu

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  1. Philip K. Liu
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  2. Brian Goudreau
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  3. Griffith S. Hsu
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Liu, P.K., Goudreau, B. & Hsu, G.S. Aphidicolin hypersensitive mutant of chinese hamster V79 fibroblasts that underproduces DNA polymerase-α antigen. Somat Cell Mol Genet 15, 331–344 (1989). https://doi.org/10.1007/BF01534972

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  • DOI: https://doi.org/10.1007/BF01534972

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