COOPERATIVE FORMATION OF HIGHER-ORDER PEROXISOME PROLIFERATOR-ACTIVATED RECEPTOR AND RETINOID-X RECEPTOR COMPLEXES ON THE PEROXISOME PROLIFERATOR RESPONSIVE ELEMENT OF THE RAT HYDRATASE-DEHYDROGENASE GENE

Peroxisome proliferator-activated receptor (PPAR) forms a heterodimer with retinoid X receptor (RXR) that binds to the peroxisome proliferat or responsive element (PPRE) to regulate the expression of target gene s, PPRE of the rat enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase (HD) gene has previously been shown to consist of three imperfect TGA CCT half-sites and form two distinct complexes (C1 and C2) with the nu clear extracts from H4IIEC3 cells. The present study identifies anothe r imperfect TGACCT motif involved in the PPAR/RXR-mediated trans-activ ation process and demonstrates that these four imperfect TGACCT motifs constitute an unique binding site consisting of two DR1 elements over lapping a DR2 element. PPAR and RXR cooperatively bind the two DR1 ele ments to form C1 complex or bind DR2 element to form C2 complex with a 1:1 ratio. Saturation of the HD PPRE probes with receptor proteins ca nnot convert the heterodimeric C2 complex to the higher order C1 compl ex, suggesting that they are formed independently. Transfection analys es indicate that mutation of any one of these TGACCT motifs or truncat ion of the entire HD PPRE into a separate DR1 and DR2 element signific antly reduced the transcriptional response of HD PPRE to peroxisome pr oliferators. The rat HD PPRE differentially binds with one or two PPAR /RXR heterodimers providing the peroxisome proliferator signaling path way with two levels of response.