Cellular and chromosomal hypersensitivity to DNA crosslinking agents and topoisomerase inhibitors in the radiosensitive Chinese hamster irs mutants: phenotypic similarities to ataxia telangiectasia and Fanconi's anaemia cells.

Abstract

The mutants irs1, irs2 and irs3 were previously isolated from the Chinese hamster line V79-4 on the basis of their hypersensitivity (2-3-fold) to cell inactivation by X-rays. Here the cross-sensitivities of the irs mutants to an array of chemical mutagens and topoisomerase inhibitors was determined in a differential cytotoxicity assay. Irs2 showed moderate hypersensitivity (2-3-fold) to simple alkylating agents and oxidative mutagens but was most sensitive (8-fold) to the topisomerase I inhibitor camptothecin. In contrast irs2 showed little or no increased sensitivity to four topoisomerase II inhibitors. Irs3 proved to be particularly hypersensitive to DNA crosslinking agents (5-15-fold) such as 1,3-butadiene diepoxide and mitomycin C. Irs1 was hypersensitive (3-fold or greater) to simple alkylating agents, oxidative mutagens and topoisomerase I and II inhibitors and exhibited extreme sensitivity (20-100-fold) to DNA crosslinking agents. The cellular hypersensitivities of irs2 and irs3 were reflected at the level of the chromosome. Camptothecin induced chromosomal aberrations in irs2 consisted almost exclusively of chromatid deletions and exchanges, whilst in irs3 1,3 butadiene diepoxide induced a 50-fold increase in chromatid exchanges compared with V79-4. The nature of irs2's camptothecin hypersensitivity was investigated. Analysis of the protein associated DNA single strand breaks produced by camptothecin indicated that there was no difference between V79-4 and irs2 in either the number of breaks induced or in the rate of their reversal following drug removal. In addition, levels of topoisomerase I activity in V79-4 and irs2 were indistinguishable. The data presented suggest that irs3 is likely to be defective in some aspect of DNA cross-link removal and irs2, whilst showing no gross defect in DNA strand break repair may fail to correctly respond to or repair certain types of strand breaks, possibly those associated with replicating DNA. The phenotypes of irs2 and irs3 respectively show similarities to those of cultured cells from the syndromes ataxia telangiectasia and Fanconi's anaemia.