A single nucleotide in an estrogen-related receptor alpha site can dictate mode of binding and peroxisome proliferator-activated receptor gamma coactivator 1alpha activation of target promoters.

The orphan nuclear receptor estrogen-related receptor alpha (ERRalpha, NR3B1) is a constitutively active transcription factor that controls multiple processes, most notably mitochondrial function. ERRalpha preferentially binds to a nine-nucleotide extended half-site sequence TNAAGGTCA, referred to as the ERRE, as either a monomer or a dimer, although how the mode of DNA binding is dictated remains to be determined. Here, we used variants of the extended half-site sequence and selective DNA binding domain mutants of ERRalpha to investigate the effects of ERRE sequence specificity on ERRalpha DNA binding mode, transactivation and interaction with the coactivator protein peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC-1alpha). We found that the base at the N position of the TNAAGGTCA sequence dictated ERRalpha binding preference as a monomer or dimer. In addition, we demonstrated that the threonine residue at position 124 (Thr(124)) was a determinant of ERRalpha DNA-dependent dimerization. Transfection experiments also indicated that substituting a thymidine for a cytosine at the N position in the ERRE of the native ERRalpha target promoter trefoil factor 1 (TFF1) considerably diminished the transcriptional response of the ERRalpha/PGC-1alpha complex. These results suggest that a single nucleotide in an ERRalpha binding site can determine specific configuration to the receptor and productive interaction with the coactivator PGC-1alpha.