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

The peroxisome-proliferator-activated receptor gamma (PPAR gamma) is a memb er of the steroid/thyroid nuclear receptor superfamily of ligand-activated transcription factors. PPAR gamma forms a heterodimer with the retinoid X r eceptor alpha (RXR alpha) and binds to a common consensus response element consisting of a direct repeat of two hexanucleotides spaced by one nucleoti de (DR1 motif). However, other hexamer configurations for binding of PPAR g amma have not been considered. By using PCR-mediated random site selection, the DNA sequence preferences for PPAR gamma binding were examined. In this study, we have demonstrated that PPAR gamma has dual DNA-binding specifici ty; binding to both the DR1 motif and a palindromic sequence with three bas es as spacers (Pal3 motif). The consensus sequence selected by equimolar am ounts of PPAR gamma and RXR alpha was a perfect DR1 motif, whereas a relati vely large population of Pal3 was observed when a 30-fold molar excess of P PAR gamma over RXR alpha. was used. Gel-shift analysis revealed that the PP AR gamma homodimer could bind to Pal3 and that the affinity constant of the PPAR gamma homodimer for Pal3 was nearly the same as that of the PPAR gamm a /RXR alpha heterodimer for DR1, The addition of RXR alpha decreased the b inding affinity of PPAR gamma for Pal3, indicating that the DNA-binding spe cificity of PPAR gamma could be altered by heterodimer formation with RXR a lpha.