Both coactivator LXXLL motif-dependent and -independent interactions are required for peroxisome proliferator-activated receptor gamma (PPAR gamma) function

Nuclear receptor activation is dependent on recruitment of coactivators, in cluding CREB-binding protein (GBP/p300) and steroid receptor coactivator-1 (SRC-1), A three-dimensional NMR approach was used to probe the coactivator binding interface in the peroxisome proliferator-activated receptor gamma (PPAR gamma) ligand binding domain (LBD), In the presence of a CBP peptide, peaks corresponding to 20 residues in helices 3, 4, 5, and 12 of the LED w ere attenuated. Alanine mutants revealed that M301A, V315A, Y320A, L468A, a nd E471A were required for binding of both CBP and SRC-1 and for cell-based transcription. Several additional amino acids in helix 4 of the PPAR gamma LBD were defective with respect to CBP recruitment, but retained relativel y normal SRC-1 recruitment. Thus these amino acid residues may be important determinants of specificity for nuclear receptor LED interactions with dis crete coactivator molecules.