Ligand-triggered stabilization of vitamin D Receptor/Retinoid X receptor heterodimer conformations on DR4-type response elements

Abstract

Nuclear receptors integrate an incoming signal in the form of a nuclear hormone by undergoing a conformational change that results via co-activator proteins in an activation of the basal transcriptional machinery. The vitamin D 3 receptor is the nuclear receptor for 1α,25-dihydroxy vitamin D 3 (1α,25(OH) 2D 3) and is known to function as a heterodimer with the retinoid X receptor on DR3-type 1α,25(OH) 2D 3 response elements. Here, it could be demonstrated that DR4-type response elements are at least as effective as DR3-type 1α,25(OH) 2D 3 response elements. Gel shift clipping analysis showed that vitamin D 3 receptor-retinoid X receptor heterodimers form in response to 1α,25(OH) 2D 3 and retinoid X receptor ligands, the pan-agonist 9- cis retinoic acid (9cRA) and the retinoid X receptor-selective retinoid CD2425, different conformations on the DR4-type element of the rat Pit-1 gene. Interestingly, on this response element the heterodimeric complexes of retinoid X receptor with the thyroid hormone receptor, the retinoic acid receptor and the benzoate ester receptor also displayed characteristic individual ligand-dependent complex formation. On the level of complex formation, utilizing DNA affinity and functional assays, only vitamin D 3 receptor-retinoid X receptor heterodimers showed a synergistic interaction of both ligands. However, the sensitivity of vitamin D 3 receptor-retinoid X receptor heterodimers to 1α,25(OH) 2D 3 was found to be much higher than to retinoid X receptor ligands. Taken together, this study demonstrates a unique interaction potential of vitamin D 3 receptor and retinoid X receptor but also establishes DR4-type response elements as multi-functional DNA binding sites with a potential to integrate various hormone signalling pathways.