DNA REPAIR GENE EXPRESSION IN MAMMALIAN CELLS

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The goals of this research proposal include the characterization of the DNA repair properties of xeroderma pigmentosum (XP) fibroblasts that express one or more of the E. coli uvr genes (uvrA, uvrB, uvrC) stably integrated into their chromosomes. A number of independent and unique phenotypic properties of these clones of XP fibroblast cell lines include: peptide maps of the Uvr proteins, structure of the integrated E. coli uvr genes, cellular localization of the respective Uvr proteins, enzymatic and host cell reactivation of damaged genes by complementing microinjected Uvr proteins and survival in the presence of DNA damaging agents. The effects of specific nuclear and cytoplasmic localization of individual repair phenotypes will be examined as well as the effects of varying the expressible concentrations of the Uvr proteins through methotrexate amplification of a co- transfected DHFR gene. An assay is being developed as a component of a feasibility study for measuring the DNA repair potential in man through an assessment of the ability of fresh lymphocytes to restore the activity of a specific gene inactivated, in this case, by ultraviolet light.