EVIDENCE FOR COINCIDENT MUTATIONS IN HUMAN LYMPHOBLAST CLONES SELECTED FOR FUNCTIONAL LOSS OF A THYMIDINE KINASE GENE

A mitotic "recombination-competent state" inducible by x-irradiation is thought to exist in yeast. We sought evidence for such a process in mammalian cells by examining the occurrence of mutations at unlinked loci in clones derived from a human lymphoblast cell line. A total of 169 independent clones that arose spontaneously or after exposure to x-rays or ethyl methanesulfonate were selected for new somatic mutations at the thymidine kinase gene on chromosome 17q. They were subsequently screened for coincident mutations by use of variable-number-of-tandem-repeat probes located on different chromosomes. Three coincident mutations were positively identified by Southern analysis on chromosomes 7 and 14; they included one that produced a new allele and two that caused loss of allele heterozygosity. Densitometric analysis of the latter two indicated the presence of two copies of the remaining allele. Several possible coincident genetic events were also observed on chromosome 17. These findings revealed a coincident mutant fraction of about 10(-2)/cell, whereas the expected mutation fraction at these loci is less than 10(-4)/cell. These results may thus provide the first molecular evidence that a "global" mutational process capable of inducing genetic instability exists in mammalian cells.