NATURAL VITAMIN-D-3 RESPONSE ELEMENTS FORMED BY INVERTED PALINDROMES - POLARITY-DIRECTED LIGAND SENSITIVITY OF VITAMIN-D-3 RECEPTOR RETINOID-X RECEPTOR HETERODIMER-MEDIATED TRANSACTIVATION

VDR, the nuclear receptor for 1,25-dihydroxyvitamin D-3 (VD), is a mem ber of the superfamily of nuclear hormone receptors and controls multi ple aspects of homeostasis, cell growth, and differentiation, VDR can function as a homodimer, but heterodimerization with the retinoid X re ceptor (RXR), retinoic acid receptor, or thyroid hormone receptor incr eases its affinity for response elements in the promoter of target gen es, All natural VD response elements identified so far consist of dire ct repeats of a variety of hexameric core binding motifs with a prefer ential spacing of three nucleotides (DR3s), However, all four VD signa lling pathways function also on response elements formed by inverted p alindromes, although these sequences were not of natural origin, Here, we report the identification of two VD response elements consisting o f inverted palindromes spaced by nine nucleotides (IP9s) in the promot ers of the human calbindin D,, gene and the rat osteocalcin gene, Like most DR3-type VD response elements, both IP9s are preferentially boun d by VDR-RXR heterodimers with a 5'-RXR-VDR-3' polarity, whose transcr iptional activity can be enhanced by costimulation with 9-cis retinoic acid, We demonstrate that changing the response element orientation r elatively to the basal promoter decreases the sensitivity of transcrip tional activation by VD by about 10-fold. Our findings indicate that i nverted palindromes are as functional as direct repeats, Furthermore, we suggest that the orientation of a nuclear receptor complex in relat ion to the basic transcriptional machinery, which is directed by heter odimer polarity and response element orientation, influences the ligan d sensitivity of the respective target gene expression.