Dual DNA-binding specificity of peroxisome-proliferator-activated receptor gamma controlled by heterodimer formation with retinoid X receptor alpha.

Abstract

The peroxisome-proliferator-activated receptor gamma (PPARgamma) is a member of the steroid/thyroid nuclear receptor superfamily of ligand-activated transcription factors. PPARgamma forms a heterodimer with the retinoid X receptor alpha (RXRalpha) and binds to a common consensus response element consisting of a direct repeat of two hexanucleotides spaced by one nucleotide (DR1 motif). However, other hexamer configurations for binding of PPARgamma have not been considered. By using PCR-mediated random site selection, the DNA sequence preferences for PPARgamma binding were examined. In this study, we have demonstrated that PPARgamma has dual DNA-binding specificity; binding to both the DR1 motif and a palindromic sequence with three bases as spacers (Pal3 motif). The consensus sequence selected by equimolar amounts of PPARgamma and RXRalpha was a perfect DR1 motif, whereas a relatively large population of Pal3 was observed when a 30-fold molar excess of PPARgamma over RXRalpha was used. Gel-shift analysis revealed that the PPARgamma homodimer could bind to Pal3 and that the affinity constant of the PPARgamma homodimer for Pal3 was nearly the same as that of the PPARgamma/RXRalpha heterodimer for DR1. The addition of RXRalpha decreased the binding affinity of PPARgamma for Pal3, indicating that the DNA-binding specificity of PPARgamma could be altered by heterodimer formation with RXRalpha.