Covalent Labeling of Rat Thymocyte and Human Lymphoid Glucocorticoid Receptor1

Lymphoid cells contain specific receptors for glucocorticoids. We have used [3H]dexamethasone-21-mesylate to label covalently glucocorticoid receptors in rat thymic lymphocytes and in neoplastic cells obtained from patients with acute lymphoblastic leukemia and malignant lymphoma. The covalently labeled glu cocorticoid receptors were identified by polyacrylamide gel electrophoresis (in the presence of 0.1% sodium dodecyl sulfate). In cytosolic fractions prepared from rat thymic lymphocytes, [3H]dexamethasone-21 -mesylate labels a protein (M, a95,000) which was identified as the glucocorticoid receptor by the following criteria: (a) labeling of this moiety is inhibited by treatment with a 100-fold molar excess of glucocorticoids, such as dexamethasone and triamcinolone acetonide; and (b) the covalently labeled M, s95,000 protein is activated (by heating at 20°for 30 min) to a form that binds to DNA-cellulose. When intact thymocytes are treated with [3H]dexamethasone-21-mesylate, an M, s95,000 moiety is also labeled covalently. Approximately 35% of the glucocorticoid receptors can be labeled covalently when intact thymocytes are treated with 100 nM [3H]dexamethasone-21mesylate for 30 min at 4°. Neoplastic cells from acute lymphoblastic leukemia and malig nant lymphoma were treated with [3H]dexamethasone-21-me sylate. In all samples, an M, s95,000 moiety was labeled cova lently; labeling was inhibited by excess glucocorticoid. Smaller moieties were also identified by competition experiments; these may represent proteolytic fragments of the M, s95,000 receptor. Thus, in rat and human lymphoid cells, [3H]dexamethasone-21mesylate can be used to label covalently the glucocorticoid receptor.