Allosteric effects of dexamethasone and RU486 on glucocorticoid receptor-DNA interactions

The glucocorticoid receptor (GR) is a DNA-binding protein that can regulate the transcription of a large number of genes in a ligand-dependent fashion. Although much progress has been made on the mechanism of transcriptional regulation by GR, a potential allosteric effect of GR-binding ligands on specific GR-DNA interactions is controversial. In this study, gel-shift methods are used to measure the effects of a classical agonist dexamethasone and a prototypical antagonist RU486 on the in vitro interactions of GR with DNA substrates, which contain glucocorticoid response elements (GREs) from promoters of GR-regulated genes. These studies show that cell extracts containing human GR bind specifically and with high affinity to GREs in the absence of ligand. An agonist dexamethasone and antagonist RU486 do not affect the affinity of GR for DNA but subtly alter the electrophoretic mobility of the GR-DNA complex. Importantly, the dissociation rate of GR from DNA increases as a function of the concentration of GRE-containing DNA. At a fixed DNA concentration, dexamethasone-bound GR dissociates from DNA significantly faster than does ligand-free GR or RU486-bound GR. These results are consistent with a model for transcriptional activation in which a dynamic complex is formed between agonist-bound GR and DNA.