Insertion of E. coli photoreactivating enzyme into V79 hamster cells

Abstract

Photoreactivating enzyme mediates the specific repair of UV light [220-300 nm, (UV)]-induced cyclobutyl pyrimidine dimers in DNA. It binds to dimer-containing DNA, and on absorption of light in the wavelength range 300-500 nm monomerizes the dimer, restoring biological activity to DNA. The specificity of the enzyme for pyrimidine dimers in DNA allows its use as an analytical tool. If UV-induced biological damage is photoreactivable, dimers are probably a major cause of that damage. Thus dimers have been implicated in producing death and mutation in prokaryotes and in simple eukaryotes. Although this photoreactivation test has great potential value in assessing the role of dimers in UV-induced damage in mammalian cells, its use in cultured mammalian cells has been limited by the dependence of photoreactivating enzyme levels on the cell species, genotype and culture medium. We have developed a method for insertion of Escherichia coli photoreactivating enzyme into mammalian cells, and show here that the inserted bacterial enzyme can mediate photoreactivation of pyrimidine dimers in V79 rodent cells.