Analysis of Glucocorticoid-resistant Human Leukemic Cells by Somatic Cell Hybridization1

Glucocorticoid-resistant mutants isolated from the glucocorticoid-sensitive human leukemic cell line CEM-C7 can be divided into three phenotypes: (a) those with almost no glucocorticoidbinding activity (r~); (b) those whose steroid-receptor complexes are unstable during attempted activation but are stabilized by the presence of sodium molybdate (acf^molybdate-sensitive); and (c) those whose steroid-receptor complexes are unstable during attempted activation but are insensitive to the presence of molybdate (acr'imolybdate-resistant). To determine if these phenotypes represent different mutations within the glucocorticoid receptor locus itself or reflect alterations in other compo nents modifying receptor function, somatic hybrids were con structed between wild-type cells and all three classes of resistant mutants, as well as between various classes of resistant mu tants. Hybrids were analyzed for chromosome content, steroidinduced growth inhibition, induction of the enzyme glutamine synthetase, and glucocorticoid receptor content. Hybrids con structed between wild-type cells and any of the three classes of resistant cells were growth-inhibited in the presence of dexamethasone, displayed normal levels of glutamine synthetase induction, and contained a quantity of glucocorticoid receptor approximately equal to the sum of the glucocorticoid receptor concentrations of the parental cell lines. Hybrids constructed between various classes of resistant cells were not growthinhibited by dexamethasone, displayed no glutamine synthetase induction, and also contained the sum of the glucocorticoid receptor concentration of the individual parents. Thus, each phenotype is recessive, and there is no complementation be tween phenotypes. We conclude that the three phenotypes are the result of different mutations within the glucocorticoid receptor locus itself and do not represent the presence of dominant receptor-inactivating factors or the absence of positive regulatory components.