Functional complementation studies with X-ray-sensitive mutants of Chinese hamster cells closely resembling ataxia-telangiectasia cells

Abstract

In order to isolate a human gene complementing the defect in A-T-like hamster cell mutants, the mutants were used as recipients for genomic DNA transfection, using either HeLa chromosomal DNA or DNA from a human cosmid library. Three primary transformants with an intermediate X-ray sensitivity and almost normal sensitivity to MMS, but retaining radioresistant DNA synthesis (RDS), were obtained. To identify the human chromosome that complements the defect in the A-T-like mutants, and to assess the degree of complementation for survival and RDS, microcell-mediated chromosome transfer was used. At least 20 independent hybrid clones between the mutant and each one of the human chromosomes 1, 2, 4, 5, 15, 17 or 18 were isolated. All hybrid clones remained X-ray sensitive, except one with chromosome 4, and another with chromosome 15, both showing an intermediate X-ray sensitivity. By using in situ hybridization we found that this partial correction was due to the presence of a mouse chromosome. In these two hybrids containing the mouse chromosome together with human chromosome 4 or 15, RDS was fully complemented only in the hybrid with chromosome 4 but not in the one containing chromosome 15, suggesting that RDS and X-ray sensitivity may be complemented independently.