Direct Interaction of the τ₁ Transactivation Domain of the Human Glucocorticoid Receptor with the Basal Transcriptional Machinery

We have used a yeast (Saccharomyces cerevisiae) cell free transcription system to study protein-protein interactions involving the τ₁ transactivation domain of the human glucocorticoid receptor that are important for transcriptional transactivation by the receptor. Purified τ₁ specifically inhibited transcription from a basal promoter derived from the CYC1 gene and from the adenovirus 2 major late core promoter in a concentration-dependent manner. This inhibition or squelching was correlated with the transactivation activity of τ₁. Recombinant yeast TATA-binding protein (yTFIID), although active in vitro, did not specifically reverse the inhibitory effect of τ₁. In addition, no specific interaction between τ₁ and yTFIID could be shown in vitro by affinity chromatography. Taken together, these results indicate that the τ₁ transactivation domain of the human glucocorticoid receptor interacts directly with the general transcriptional apparatus through some target protein(s) that is distinct from the TATA-binding factor. Furthermore, this assay can be used to identify interacting factors, since after phosphocellulose chromatography of a whole-cell yeast extract, a fraction that contained an activity which selectively counteracted the squelching effect of τ₁ was found.