MODULATION OF NUCLEAR RECEPTOR INTERACTIONS BY LIGANDS - KINETIC-ANALYSIS USING SURFACE-PLASMON RESONANCE

Many nuclear hormone receptors, including the human 1,25-dihydroxyvita min D-3 receptor (VDR), bind cooperatively to DNA as either homodimers or heterodimers with the 9-cis-retinoic acid receptor (RXR). Protein- protein interactions mediated by residues within both the DNA- and lig and-binding domains contribute to this binding. We have previously rep orted that the ligands for VDR and RXR can modulate the affinity of th e receptors' interaction with DNA [Cheskis, B., & Freedman, L. P. (199 4) Mol. Cell. Biol. 14, 3329-3338]. To examine this in more detail, we report here the use of surface plasmon resonance (SPR) to characteriz e the kinetics of both protein-protein and protein-DNA interactions by VDR and RXR in the presence and absence of their cognate ligands. We find that 1,25 dihydroxyvitamin D-3 binding favors both VDR-RXR hetero dimerization and, as a result. DNA binding by the complex. Conversely, the ligand reduces VDR homodimerization in solution and the affinity of VDR-DNA interaction. 9-cis-Retinoic acid attenuates the stimulating effect of 1,25-dihydroxyvitamin D-3 by decreasing the rate of VDR-RXR heterodimer formation and simultaneously by increasing the affinity o f RXR homodimerization. Thus, using SPR, we have shown that a major ro le for such ligands is to regulate nuclear receptor dimerization both in solution and on DNA. The ligands appear to do so dynamically, modul ating the overall affinity of these complexes. This mechanism therefor e creates a fast and sensitive way to regulate DNA binding in response to changes in ligand concentration.