RETINOID X-RECEPTOR-ALPHA BINDS WITH THE HIGHEST AFFINITY TO AN IMPERFECT DIRECT REPEAT RESPONSE ELEMENT

The regulation of gene expression by retinoids is mediated by two clas ses of receptors, retinoic acid receptors and retinoid X receptors (RX R). RXR can bind to specific target genes as homodimers, and these hom odimers can activate gene expression in the presence of the Ligand 9-c is-retinoic acid. A direct repeat of AGGTCA with a 1 base pair spacer (DR1) acts as a RXR homodimer response element in the presence of 9-ci s-retinoic acid. However, it is not known if this represents the highe st affinity binding site for the RXR homodimer. To investigate this qu estion, we used a nonbiased strategy to isolate from a pool of random DNA those sequences that have the highest affinity for RXR alpha homod imers. The imperfect DR1 sequence 5'-GGGGTCAAAGGTCA displayed the high est in vitro binding affinity for RXR alpha homodimers. Transient tran sfection studies confirmed that this sequence is a more potent respons e element than is a perfect DR1 of either AGGTCA or GGGGTCA. The resul ts also indicate that for RXR alpha homodimers, the receptor hound to the 5' half-site dislays different DNA binding specificity than that b ound to the 3' half-site. Thus, DNA binding specificity is determined not only by the amino acid sequence of the protein but also by its pro tein-protein interactions and its position on the response element (5' vs. 3').