A REGULATORY ROLE FOR RIP140 IN NUCLEAR RECEPTOR ACTIVATION

Transcriptional regulation of gene expression by nuclear receptors req uires negatively and positively acting cofactors. Recent models for re ceptor activation propose that certain receptors in the absence of lig ands can recruit corepressors while ligand binding results in conforma tional changes leading to the recruitment of coactivators. Previous wo rk has established a coactivator role for the SRC-1 Family members as well as an involvement of the coactivators CBP/p300 in nuclear recepto r signaling. However, in addition to coactivators, ligand-activated nu clear receptors bind a number of different proteins that possibly serv e other functions. Using peroxisome proliferator-activated receptor-al pha (PPAR alpha) as bait in a yeast two-hybrid screening, we have isol ated nuclear factor RIP140 whose function in receptor activation is un clear. We now report a detailed characterization of RIP140 action with a focus on the retinoid X receptor (RXR) heterodimeric receptors PPAR and thyroid hormone receptor (TR). We show that putative PPAR ligands enhance the interaction of RIP140 with the rat PPAR subtypes alpha an d gamma in solution but not with PPAR/RXR heterodimers on DNA. However , RIP140 forms ternary complexes in the presence of RXR ligands. Simil ar experiments with Tn support the high affinity of RIP140 to the RXR subunit and also suggest that either partner in the TR/RXR heterodimer can independently respond to ligand. Coactivation experiments in yeas t and mammalian cells confirm the coactivator role for SRC-1, but not for RIP140. We provide important evidence that the in vitro binding of RIP140 and SRC-1 to nuclear receptors is competitive. Since RIP140 ge nerally down-regulates receptor activity in mammalian cells and specif ically down-regulates coactivation mediated by SRC-1, we propose a mod el in which RIP140 indirectly regulates nuclear receptor AF-2 activity by competition for coactivators such as SRC-1.