PPARβ/δ controls Mediator recruitment and transcription initiation

Abstract

Repression of transcription by nuclear receptors involves NCOR and SMRT corepressor complexes, which harbour the deacetylase HDAC3 as a subunit. Both deacetylase-dependent and -independent repression mechanisms have been reported for these complexes. In the absence of ligands, the nuclear receptor PPAR{beta}/{delta} recruits NCOR and SMRT and represses expression of its canonical targets including the ANGPTL4 gene. Agonistic ligands cause corepressor dissociation and enable enhanced induction of transcription by coactivators. Vice versa, recently developed synthetic inverse agonists lead to augmented corepressor recruitment and repression that dominates over activating stimuli. Both basal repression of ANGPTL4 and reinforced repression elicited by inverse agonists are partially insensitive to HDAC inhibition. This raises the question of how PPAR{beta}/{delta} represses transcription mechanistically. Here, we show that the PPAR{beta}/{delta} inverse agonist PT-S264 impairs transcription initiation in human cells. Inverse agonist-bound PPAR{beta}/{delta} interferes with recruitment of Mediator, RNA polymerase II, and TFIIB, but not with recruitment of other basal transcription factors, to the ANGPTL4 promoter. We identify NCOR as the main ligand-dependent interactor of PPAR{beta}/{delta} in the presence of PT-S264. In PPAR{beta}/{delta} knockout cells, reconstitution with PPAR{beta}/{delta} mutants deficient in basal repression recruit less NCOR, SMRT, and HDAC3 to chromatin, concomitant with increased binding of RNA polymerase II. PT-S264 restores binding of NCOR, SMRT, and HDAC3, resulting in diminished polymerase II binding and transcriptional repression. In the presence of HDAC inhibitors, ligand-mediated repression of PPAR{beta}/{delta} target genes is only partially relieved. Our findings corroborate deacetylase-dependent and -independent repressive functions of HDAC3-containing complexes. Deacetylase-independent repression mediated by binding of inverse agonists to PPAR{beta}/{delta} involve NCOR/SMRT recruitment and interference with Mediator, TFIIB, and RNA polymerase II binding.