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.

Carbon Radioisotopes↗

PubMed1974

DNA repair in human/embryonic chick heterokaryons. Ability of each species to aid the other in the removal of ultraviolet-induced damage.

Cultured human and embryonic chick fibroblasts possess different enzyme-mediated processes to repair cyclobutyl pyrimidine dimers induced in their deoxyribonucleic acid (DNA) by ultraviolet (UV) radiation. While dimers are corrected in human cells by excision repair, a photoenzymatic repair process exists in embryonic chick cells for the removal of these potentially deleterious UV photoproducts. We have utilized a sensitive enzymatic assay to monitor the disappearance, i.e. repair, of dimer-containing sites in fused populations of human and chick cells primarily consisting of multinucleate human/chick heterokaryons. Fused cultures were constructed such that UV photoproducts were present only in chick DNA when evaluating excision repair and only in human DNA when evaluating photoenzymatic repair. Based on the kinetics of site removal observed in these cultures we are led to conclude the following: Within heterokaryons per se the photoreactivating enzyme derived from chick nuclei and at least one excision-repair enzyme (presumably a UV endonuclease) derived from human nuclei act on UV-damaged DNA in foreign nuclei with an efficiency equal to that displayed toward their own nuclear DNA. Hence, after cell fusion these chick and human repair enzymes are apparently able to diffuse into foreign nuclei and once therein competently attack UV-irradiated DNA independently of its origin. In harmony with the situation in nonfused parental cultures, in heterokaryons the chick photoenzymatic repair process rapidly removed all dimer-containing sites from human DNA including the residual fraction normally acted upon slowly by the human excision-repair process.

Animals↗

PubMed1974

Human antigen and enzyme markers in man-Chinese hamster somatic cell hybrids: evidence for synteny between the HL-A, PGM3, ME1, and IPO-B loci.

In man-Chinese hamster somatic cell hybrids the segregation of the loci for 27 human enzyme markers and the species-specific surface antigens, including the HL-A histocompatibility antigens, was studied. The results show a synteny of the human loci for phosphoglucomutase 3, cytoplasmic malic enzyme, tetrameric indophenol oxidase, and HL-A. Furthermore, evidence is presented that the loci for the human species-specific antigens are distributed over several chromosomes.

Animals↗

PubMed1974