Transcriptional activation by thyroid hormone receptor-β involves chromatin remodeling, histone acetylation, and synergistic stimulation by p300 and steroid receptor coactivators

Abstract

Transcriptional regulation by heterodimers of thyroid hormone receptor (TR) and the 9-cis retinoid X receptor (RXR) is a highly complex process involving a large number of accessory factors, as well as chromatin remodeling. We have used a biochemical approach, including an in vitro chromatin assembly and transcription system that accurately recapitulates ligand- and activation function (AF)-2-dependent transcriptional activation by TRβ/RXRα heterodimers, as well as in vitro chromatin immunoprecipitation assays, to study the mechanisms of TRβ-mediated transcription with chromatin templates. Using this approach, we show that chromatin is required for robust ligand-dependent activation by TRβ. We also show that the binding of liganded TRβ to chromatin induces promoter-proximal chromatin remodeling and histone acetylation, and that histone acetylation is correlated with increased TRβ-dependent transcription. Additionally, we find that steroid receptor coactivators (SRCs) and p300 function synergistically to stimulate TRβ-dependent transcription, with multiple functional domains of p300 contributing to its coactivator activity with TRβ. A major conclusion from our experiments is that the primary role of the SRC proteins is to recruit p300/cAMP response element binding protein-binding protein to hormone-regulated promoters. Together, our results suggest a multiple step pathway for transcriptional regulation by liganded TRβ, including chromatin remodeling, recruitment of coactivators, targeted histone acetylation, and recruitment of the RNA polymerase II transcriptional machinery. Our studies highlight the functional importance of chromatin in transcriptional control and further define the molecular mechanisms by which the SRC and p300 coactivators facilitate transcriptional activation by liganded TRβ.