Glucocorticosteroid receptor dimerization investigated by analysis of receptor binding to glucocorticosteroid...

The aim of this study was to analyze the role of regions of the glucocorticosteroid receptor (GR) outside the DNA binding domain (DBD) in GR binding and homodimerization efficiencies by using a model according to which GR monomers and dimers are in equilibrium and able to bind to each half-palindromic motif of a GRE. We studied wild-type human GR (hGR), an N-terminal domain deleted mutant (lacking amino acids 1-417), a C-terminal deleted mutant (lacking amino acids 550-777, the main part of the ligand binding domain), and two rat GR derivatives limited to the DNA binding domain and proximal sequences. Specific GR monomer and dimer complexes with 33P-labeled palindromic or half-palindromic GREs were identified by gel-shift and methylation interference experiments. The different complexes were quantified, and the multiple equilibrium constants for their formation were determined. The affinity of the monomer for the GRE was not affected by the deletions of regions outside the DBD. However, the affinity of the dimer for the GRE was clearly increased by the presence of the N-terminal domain and, to a lesser extent, by that of the main part of the C-terminal domain. By using this model, we also obtained a GR dimerization constant in the absence of specific binding to GRE. Dimerization of the DBD was not increased by the presence of only one of the GR terminal domains, but an increase in dimerization efficiency was observed when both domains were present, suggesting a structural synergy between the N- and C-terminal domains in GR homodimerization.