Nonactivated and activated glucocorticoid-receptor complexes in WEHI-7 and rat thymus cells

Abstract

Our own results and those of others have indicated that nonactivated glucocorticoid-receptor complexes are oligomeric proteins with Stokes radius R s = 8–9 nm, and that activation is accompanied by a reduction in size to R s = 5–6 nm. The most convincing evidence for the large size of the nonactivated compared to the activated complex has been obtained with cytosols stabilized with molybdate. It has been suggested, however, that molybdate causes aggregation of complexes. Here we show that nonactivated rat thymus complexes in cytosols with molybdate and 400 M KCl have R s = 8 nm. Furthermore, cytosols from WEHI-cells, which are exceptionally stable, show clear indications of 8 nM nonactivated complexes even without molybdate. The principal complexes in thymus cells under physiological conditions are the nonactivated, activated and nuclear-bound forms. We have studied the rapid kinetics of formation and interconversion of these complexes in intact cells at 37°C, using our newly-developed mini-column procedure to assay nonactivated and activated complexes. These kinetic results, along with many earlier results, can be accounted for quantitatively with a simple cyclic (irreversible) model in which the dissociation rate constant of the steroid plays a key role. The model predicts correctly the different degrees of activation in the cell with glucocorticoids such as triamcinolone acetonide and dexamethasone on the one hand, and cortisol and corticosterone on the other, without assuming steroid-specific allosteric influences of each of these steroids on the receptor.