Identification of transformed glucocorticoid receptor from dexamethasone resistant melanoma

Abstract

We have examined properties of glucocorticoid-receptor complexes which might account for dexamethasone induced alterations in the growth and morphology of melanoma target cells. We have used cultured cells and solid tumors derived from the dexamethasone-sensilive RPMI 3460 Syrian hamster melanoma cell line together with clonal variants which are either more sensitive (clone 6) or resistant (clone 5) to the growth inhibiting effects of dexamethasone. Although differing markedly in their response to corticoids. each of the cell lines contains significant quantities of glucocorticoid receptor. The present studies were designed to determine if differences in the transformability (nuclear binding ability) of the glucocorticoid receptors from these target cells could account for their sensitivity or resistance to glucocorticoids. Cytosolic glucocorticoid-receptor complexes were analyzed by DEAE-cellulose chromatography to identify and quantitate the relative amounts of native (unactivated) and transformed (activated-nuclear binding) receptors present. We could readily separate these two major forms of the glucocorticoid-receptor complex in cytosols from each of the melanoma cell lines and solid tumors examined. The identity of these receptor complexes as native or transformed was confirmed using both ATP-agarose and isolated nuclei; only transformed receptor complexes are bound. When cytosol is prepared in the presence of 10m M sodium molybdate, only native receptor is present. After molybdate is removed, native glucocorticoid receptor is readily transformed by increased ionic strength. We conclude that resistance to dexamethasone-induccd changes in growth observed in resistant clone 5 cells and solid tumors cannot be attributed to an inability of the receptor they contain to exist as a stable, transformed complex.