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In order to study the mechanism of induction of mutations and chromosome aberrations by ionizing radiations, it is particularly useful to have available radiation-sensitive mutants. While several X-ray-sensitive rodent cell lines are available, they have been selected rather nonspecifically. It was determined that selection for resistance to the DNA replication/repair inhibitor, 1-beta-D-arabinofuranosylcytosine (ara-C), would permit production of a set of X-ray-sensitive mutant cell lines that would be defective in the resynthesis step of excision or recombination repair. Such mutant cells could also be used for the isolation and characterization of human DNA repair genes. In particular, it was predicted that the repair gene defective in individuals with ataxia telangiectasia (AT) might be amenable to study with ara-C-resistant (X-ray-sensitive) mutants, since additional studies, presented here, have shown that AT cells are resistant to ara-C. In the long term, it is hoped that determining the specific defect in AT might lead to an understanding of the possible role of defective repair in tumor induction and/or progression. The general approach used to isolate ara-C-resistant Chinese hamster ovary cell mutants was to treat cells with ethyl methanesulfonate and select in increasing concentrations of ara-C. Although several mutants were isolated, one in particular, Ara-CR213, has been studied most extensively. It was selected largely because it shows the greatest sensitivity to X-rays. Ara-CR213 cells were hypersensitive to the killing effect of X-rays with an LD10 of 2.5 Gy as compared to the wild-type cells that had an LD10 of 6 Gy. The mutant showed an increased frequency of X-ray-induced chromosomal aberrations in the G1 and G2 stages of the cell cycle compared to wild-type frequencies. There was no increase in sister chromatid exchange levels. All of these observations in Ara-CR213 are very similar to those made with AT cells in our and other laboratories. Even more important, complementation analysis of Ara-CR213 x AT hybrid cells indicated that the gene responsible for X-ray sensitivity of AT is also mutated in Ara-CR213 cells. Thus, Ara-CR213 appears to have a mutant phenotype and probably genotype that is very similar to, if not exactly the same as, those of AT. This makes it quite different from other X-ray-sensitive cells that have been isolated in other laboratories.

A detailed cytogenetic study of maturing mouse oocyte radiosensitivity was performed. Oocytes were collected at various intervals ranging from 1.5 days to 28.5 days after irradiation with 50, 100, 200, and 300R of acute X-rays. The observed sensitivity to chromatid aberration induction varied greatly over this time span. Sensitivity was lowest at the shortest time interval before ovulation and gradually increased up to 9.5 days; it then remained constant until insufficient numbers of oocytes could be collected. The data were analyzed in three ways. First, the data from all time intervals at each dose were pooled; second the data from the least sensitive time intervals, at each dose, were pooled, and third, the data from the period of uniform sensitivity, at each dose, were pooled. Dose-response regression analyses were done on these pooled data and the best fits obtained were to the models Y= a + bD + cD2 and Y= a + cD2 for both deletions and interchanges. This result is interpreted as indicating that the aberrations result from a predominantly two-track process. The cytogenetic data were compared to specific-locus mutation induction data in comparable oocyte stages, and qualitative similarity in dosei esponse characteristics were observed. This similarity is interpreted to mean that both events result from the same mechanism, and that the large dose-rate effect, observed for both events, is a reflection of the two-track component in the dose-response curves.

Split-dose experiments were done on maturing dictyate oocytes to determine if the magnitude of the first dose influenced the "rejoining time" of radiation-induced chromosomal lesions. A total dose of 400r was split into various combinations with varying fractionation intervals. The data derived from analyzing interchanges indicate that there is no difference in the rejoining time whether the first dose was 100, 200, or 300r. It thus appears that the radiation dose in the ranges studied does not significantly alter the rate of repair of the chromosomal lesions. This conclusion is contrary to that which has been propounded to explain the nonlinear dose curves obtained for specific locus mutations. Chronic 60Co Υ-ray exposures were given to female mice over an 8-day period. The exposures were delivered during the period of peak sensitivity, i.e., 8-16 days prior to ovulation. The doses given were 117, 240, 348, and 483r. The aberration yields observed were dramatically lower than for comparable doses of acute X rays even when the RBE of Υ rays compared with X rays is taken into account. The large drop in yields at the low dose rates is interpreted as resulting from a large two-track component in the acute curve, and as being independent of effects on repair systems.