Kinetic Pulse-Chase Labeling Study of the Glucocorticoid Receptor in Mouse Lymphoma Cells

A kinetic pulse-chase labeling technique was used to measure the intracellular half-life of the glucocorticoid receptor in S49 mouse lymphoma cells. Cells were pulse-labeled with [35S]methionine for 30 min and then cultured in the presence of unlabeled methionine (chase). Labeled receptors were quantitated at periodic time points during the chase by immunoadsorption to protein A-Sepharose using the BuGR2 monoclonal antireceptor antibody. The decay of labeled receptors during the chase was linear on a semilog plot, consistent with first order kinetics. Receptor half-life was 9 h when cells were cultured in either phenol red-containing medium supplemented with fetal calf serum or in phenol red free-medium supplemented with charcoal extracted serum, indicating that endogenous steroids do not affect receptor half-life. Receptor half-life was also unchanged when cells were cultured in the presence of 0.1 microM dexamethasone, a glucocorticoid hormone, or 0.1 microM RU486 (11 beta-(4-dimethylamino-phenyl)-17 beta-hydroxy-17 alpha-(propynylestra-4,9- diene-3-one), an antiglucocorticoid hormone. We conclude that the intracellular half-life of the glucocorticoid receptor in S49 mouse lymphoma cells is not regulated by either glucocorticoid or antiglucocorticoid hormones.