Phenotype reversal in induced mutants of CHO cells: analysis of the reversed cell lines.

We have previously shown that descendants of CHO-derived hprt or aprt mutants induced by ethyl methanesulphonate usually undergo a rapid loss of the mutant phenotype during the 10 generations or so of culture in non-selective medium immediately following mutagenesis (Bradley, 1980; Bradley and Laviolette, 1989). We now present an analysis of several mutants and their descendants which have lost the mutant phenotype, or 'reversed'. The drug-resistance properties of reversed cells were generally intermediate between W.T. and mutant, and message level and enzyme-specific activity were also intermediate, correlating with the phenotype. Although this was consistent with a model of inactivation-reactivation of the target gene to explain the reversal phenomenon, the model was ruled out by Northern blot analysis of several induced mutants, which showed no correlation between level of message and tendency of the mutant to lose its phenotype. Karyotype analysis showed that three out of four reversed lines were near-tetraploid and the fourth had a substantial proportion of near-tetraploid cells. This suggests cell fusion between a mutant and a W.T. cell may explain the phenomenon. A prediction of this model, namely that mutagen treatment increases cell hybrid formation, was tested and found to be true.