Cytological characterization of Chinese hamster ovary X-ray-sensitive mutant cells, xrs 5 and xrs 6: II. Induction of sister-chromatid exchanges and chromosomal aberrations by X-rays and UV-irradiation and their modulation by inhibitors of poly(ADP-ribose) synthetase and α-polymerase

Abstract

The cell killing and induction of sister-chromatid exchanges (SCEs) by X-rays and short-wave ultraviolet (UV) irradiation in combination with inhibitors of DNA repair, 3-aminobenzamide (3AB), cytosine arabinoside (ara-C) or aphidicolin (APC) were studied in wild-type CHO-K1 and two X-ray-sensitive mutants, xrs 5 and xrs 6 cells. The spontaneous frequency of SCEs was similar in the mutants and the wild-type CHO-K1 cells (8.4–10.3 SCEs/cell). Though X-rays are known to be poor inducers of SCEs, a dose-dependent increase in the frequency of SCEs in xrs 6 cells (doubling at 150 rad) was found in comparison to a small increase in xrs 5 and no increase in wild-type CHO-K1 cells. 3AB, an inhibitor of poly(ADP-ribose) synthetase increased the spontaneous frequency of SCEs in all the cell types. 3AB did not potentiate the X-ray-induced frequency of SCEs in any of the cell lines. Ara-C, an inhibitor of DNA polymerase α, increased the frequency of SCEs in all the cell lines. In combined treatment with X-rays, ara-C had no synergistic effect in xrs 5 and xrs 6 cells, but the frequency of SCEs increased in X-irradiated wild-type CHO-K1 cells post-treated with ara-C. For the induced frequency of SCEs, CHO-K1 cells terated with X-rays plus ara-C behaved like xrs 6 cells treated with X-rays alone, suggesting a possible defect in DNA base damage repair in xrs 6 cells, in addition to the known defective repair of DNA double-strand breaks (DSBs). Survival experiments revealed higher sensitivity of xrs 5 and xrs 6 mutant cells to the cell killing effect of X-rays in S-phase when compared to wild-type CHO-K1 cells. The mutants responded with lesser sensitivity to cell killing effect of ara-C and APC than CHO-K1 cells, the relative sensitivity to ara-C or APC being CHO-K1 > xrs 5 > xrs 6 cells. When X-irradiation was coupled with ara-C, the results obtained for survival were similar to those of the SCE test, i.e., unlike wild-type CHO-K1, no synergistic effect was observed in xrs 5 or xrs 6 cells. After UV-irradiation, the frequency of SCEs increased similarly in wild-type CHO-K1 and xrs 6 cells, but xrs 5 cells responded with lower frequency of SCEs. When UV-irradiation was followed by post-treatment with ara-C, a synergistic effect was obtained in CHO-K1 cells, but not in xrs 5 or xrs 6 cells. UV alone in G 1-stage increased the frequencies of chromatid-type of aberrations in all cell lines, but the response was more pronounced in mutants xrs 5 and xrs 6 when compared to wild-type CHO-K1 cells. UV generally does not induce chromosome-type aberrations in G 1. It induced chromosome-type aberrations when coupled with ara-C in CHO-K1 ( P < 0.01) and in xrs 5 ( P ≤ 0.05), but no induction of chromosome-type aberrations was obtained in xrs 6 cells. On the basis of these findings, the mechanism of involvement of specific type of DNA lesion responsible for formation of SCE and chromosomal aberration in X-or UV-irradiated cells is discussed.