Thyroid hormone receptor functions as monomeric ligand-induced transcription factor on octameric half-sites....

The thyroid hormone (3,5,3'-triiodothyronine) receptor (T3R) belongs to the nuclear receptor superfamily of ligand-inducible transcription factors. T3Rs are known to bind as homodimers and heterodimers with retinoid X receptors (RXRs) to two hexameric half-sites in directly repeated, palindromic, and inverted palindromic orientations. The binding of T3R monomers to individual half-sites was often reported, but no clear ligand-induced transactivational activity has been shown. Here, we analyzed interactions of T3R monomers with individual half-sites of the sequence NNAGGTCA. We found that the two nucleotides 5' of the AGGTCA core half-site strongly influence T3R binding and transcriptional activity: octameric half-sites of the consensus sequence (T/C)(A/G)AGGTCA were bound by T3Rs with the highest affinity. This suggests T3R functioning also as a monomeric transcription factor like the orphan nuclear receptors NGFI-B and FTZ-F1. Moreover, we observed that the function of T3R-RXR heterodimers on response elements composed of two half-sites in a directly repeated orientation spaced by 4 nucleotides is determined in major parts by the 5'-flanking sequence of the upstream half-site. Consequently, we noted that the affinity of T3R homodimers is influenced by both 5'-flanking sequences. Our findings suggest that the binding of dimerizing receptors like T3R and other nuclear receptors to their cognate response elements is determined not only by the half-site core sequence, orientation, and number of spacing nucleotides, but also by the nucleotide sequence preceding the half-sites.