GCN5 and ADA adaptor proteins regulate triiodothyronine/GRIP1 and SRC-1 coactivator-dependent gene activation by the human thyroid hormone receptor

We have used yeast genetics and in vitro protein-protein interaction experiments to explore the possibility that GCN5 (general control nonrepressed protein 5) and several other ADA (alteration/deficiency in activation) adaptor proteins of the multimeric SAGA complex can regulate T-3/GRIP1 (glucocorticoid receptor interacting protein 1) and SRC-1 (steroid receptor coactivator-1) coactivator-dependent activation of transcription by the human T-3 receptor beta 1 (hTR beta 1). Here, we show that in vivo activation of a T-3/GRIP1 or SRC-1 coactivator-dependent T-3 hormone response element by hTR beta 1 is dependent upon the presence of yeast GCN5, ADA2, ADA1, or ADA3 adaptor proteins and that the histone acetyltransferase (HAT) domains and bromodomain (BrD) of yGCN5 must be intact for maximal activation of transcription. We also observed that hTR beta 1 can bind directly to yeast or human GCN5 as well as hADA2, and that the hGCN5(387-837) sequence could bind directly to either GRIP1 or SRC-1 coactivator. Importantly, the T-3-dependent binding of hTR beta 1to hGCN5(387-837) could be markedly increased by the presence of GRIP1 or SRC1. Mutagenesis of GRIP1 nuclear receptor (NR) Box II and III LXXLL motifs also substantially decreased both in vivo activation of transcription and in vitro T-3-dependent binding of hTR beta 1 to hGCN5. Taken together, these experiments support a multistep model of transcriptional initiation wherein the binding of T-3 to hTR beta 1 initiates the recruitment of p160 coactivators and GCN5 to form a trimeric transcriptional complex that activates target genes through interactions with ADA/SAGA adaptor proteins and nucleosomal histones.