Photoreactivation and excision repair of ultraviolet radiation-injured DNA in primary embryonic chick cells.
Primary embryonic chick cells have been evaluated on the basis of their capacity to repair photochemical lesions produced in the deoxyribonucleic acid (DNA) by ultraviolet (UV) radiation. The fate of one prominent class of UV photoproducts, cyclobutane pyrimidine dimers, was monitored by an in vitro enzymatic assay. UV-irradiated cultures were incubated for prescribed times after which their damaged, radioactive-labeled DNA was extracted and exposed to a purified UV endonuclease selectively active toward sites altered by dimer formation. Single-strand scissions specifically introduced by the enzyme treatment and, therefore, the dimer-containing sites remaining in the DNA were quantified retrospectively by velocity sedimentation in alkaline sucrose. When the chick fibroblasts were incubated in black light, essentially all nuclease-susceptible sites rapidly disappeared from the UV-damaged DNA. In sharp contrast, incubation of the irradiated cultures in total darkness severely impeded the metabolic machinery responsible for site elimination. A substantial amount of UV-stimulated DNA repair synthesis was also detected in the chick cells by conventional techniques involving isopyknic centrifugation and autoradiography. However, the UV photoproducts triggering this indicator of excision repair were probably not dimers since incubation of the irradiated cultures in the light rather than in the dark did not lead to a diminution in the extent of repair synthesis. By these criteria of DNA repair, it appears that embryonic chick cells primarily rely on a highly proficient, light-requiring mechanism, presumably enzymatic photoreactivation, for dimer elimination but also possess a light-independent, excision-type process to cope with other, as yet unidentified, photochemical defects.
D002250:Carbon Radioisotopes; D002455:Cell Division / Q000528:radiation effects; D002478:Cells, Cultured; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000096:biosynthesis / Q000528:radiation effects*; D004260:DNA Repair / Q000528:radiation effects*; D004307:Dose-Response Relationship, Radiation; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D008938:Mitosis / Q000528:radiation effects; D011830:Radiation Effects; D013194:Staining and Labeling; D013936:Thymidine / Q000378:metabolism; D013997:Time Factors; D014316:Tritium; D014466:Ultraviolet Rays
D002250:Carbon Radioisotopes; D014316:Tritium; D004247:DNA; D013936:Thymidine