GLUCOCORTICOSTEROID RECEPTOR DIMERIZATION INVESTIGATED BY ANALYSIS OFRECEPTOR-BINDING TO GLUCOCORTICOSTEROID RESPONSIVE ELEMENTS USING A MONOMER-DIMER EQUILIBRIUM-MODEL

The aim of this study was to analyze the role of regions of the glucoc orticosteroid 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 t o 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 p art of the ligand binding domain), and two rat GR derivatives limited to the DNA binding domain and proximal sequences. Specific CR monomer and dimer complexes with P-33-labeled palindromic or half-palindromic GREs were identified by gel-shift and methylation interference experim ents. The different complexes were quantified, and the multiple equili brium constants for their formation were determined. The affinity of t he monomer for the GRE was not affected by the deletions of regions ou tside the DBD. However, the affinity of the dimer for the GRE was clea rly increased by the presence of the N-terminal domain and, to a lesse r 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 increa se in dimerization efficiency was observed when both domains were pres ent, suggesting a structural synergy between the N- and C-terminal dom ains in GR homodimerization.