The glucocorticoid receptor in GH1 cells. Evidence from dense amino acid labeling and whole cell studies for an equilibrium model explaining the influence of hormone on the intracellular distribution of receptor.

The glucocorticoid receptor in GH1 cells cultured without glucocorticoids and harvested in hypotonic buffer is primarily a cytosolic protein with a sedimentation coefficient of 10 S in low salt gradients. In gradients containing 0.4 M KCl, the receptor sediments at 3.5 S. Following incubation of cells for various times in culture medium containing dense amino acids, discrete peaks of newly synthesized dense receptor and normal density 3.5 S receptor are resolved in salt-containing gradients. The half-life of the 3.5 S receptor is 9.5 h. In contrast, the 10 S receptor gradually becomes more dense as the time of incubation of cells with dense medium increases and discrete normal and dense receptor peaks are not resolved in sucrose gradients. These results suggest that the 10 S receptor is an oligomer and is in a rapidly exchanging equilibrium in intact cells at 37 degrees C with its 3.5 S hormone-binding subunit. When GH1 cells are incubated for 1 h at 37 degrees C with various concentrations of [3H]9 alpha-fluoro-11 beta, 16 alpha, 17 alpha, 21-tetrahydroxypregna-1,4-diene-3,20-dione, 16,17-acetonide ([3H]triamcinolone acetonide), receptor is found both in the cytosol and the nucleus. Sedimentation of the cytosol in low salt gradients reveals that a 4 S receptor form is present along with 10 S receptor. As the concentration of [3H]triamcinolone acetonide is increased, the amounts of both 4 S cytosolic receptor and nuclear receptor increase and the amount of 10 S cytosolic receptor decreases. Following removal of [3H]triamcinolone acetonide from cells, the amounts of both 4 S cytosolic receptor and nuclear receptor decrease and the amount of 10 S cytosolic receptor increases. Density labeling indicates that this increase in 10 S receptor is not due to new receptor synthesis. The 4 S receptor form, but not the 10 S form, binds to purified DNA. Resedimentation studies suggest that the 4 S receptor found in low salt gradients is identical with or derived from the 3.5 S form found in salt-containing gradients. A model for hormone-mediated receptor translocation to the nucleus is proposed in which binding of hormone shifts the equilibrium between the oligomeric 10 S receptor and its 4 S subunit toward the 4 S form. The 4 S form is in equilibrium with 4 S receptor bound to chromatin. The model provides a molecular explanation for many experimental findings concerning glucocorticoid and other steroid receptors.