Recruitment of nuclear receptor corepressor and coactivator to the retinoic acid receptor by retinoid ligands - Influence of DNA-heterodimer interactions

Ligand activation of retinoic acid receptors (RARs) involves coordinated ch anges in their interaction with coregulatory molecules. Binding of the agon ist all-trans-retinoic acid to the RAR results in increased interaction wit h coactivator molecules as well as a decreased interaction with corepressor molecules. Thus, an all-trans-retinoic acid antagonist might function eith er by preventing agonist induction of such events or, additionally, by acti vely increasing repression via corepressor recruitment. We demonstrate that the repression of the transcriptional activity of a constitutively active RAR gamma-VP-16 chimeric receptor by the inverse agonist AGN193109 requires a functional Co-R box and that binding of this ligand to RAR gamma leads t o an increased interaction with the corepressor N-CoR both in glutathione S -transferase pull-down and yeast two-hybrid analyses. Detection of nuclear receptor corepressor (N-CoR) association with RAR gamma was greatly facilit ated by inclusion of a RARE oligonucleotide in coimmunoprecipitation analys es, a result of an increase in association of the ternary complex consistin g of RAR, RXR, and DNA. Similarly, this DNA-dependent increase in heterodim er formation likewise resulted in an increase in agonist-mediated recruitme nt efficiency of the coactivator SRC-1, Under conditions which favor ternar y complex formation, a RAR neutral antagonist is distinguished from an inve rse agonist with respect to corepressor recruitment as is a RAR partial ago nist distinguished from an agonist with respect to coactivator recruitment. These results indicate that it is possible to design RAR ligands with dist inct recruitment capabilities for coregulators, both coactivators as well a s corepressors, In addition, using this recruitment assay, we show that SRC -1 and the related coactivator molecule ACTR associate with the ternary com plex via utilization of different helical motifs within their conserved rec eptor interaction domains.