TRANSACTIVATION BY THE HUMAN ARYL-HYDROCARBON RECEPTOR AND ARYL-HYDROCARBON RECEPTOR NUCLEAR TRANSLOCATOR PROTEINS - DIRECT INTERACTIONS WITH BASAL TRANSCRIPTION FACTORS

The aryl hydrocarbon (or dioxin) receptor (AhR) is a ligand-activated basic helix-loop-helix (bHLH) protein that heterodimerizes with the bH LH protein AhR nuclear translocator (ARNT) to form a complex that bind s to xenobiotic regulatory elements in the enhancers of target genes. We used a series of fusion proteins, with a heterologous DNA-binding d omain, to study independently the irans-activating function of the hum an AhR and ARNT proteins in yeast. The results confirm that both the h uman AhR and ARNT contain carboxyl-terminal trans-activation domains. The AhR has a complex trans-activation domain that is composed of mult iple segments that function independently and exhibit varying levels o f activation. Furthermore, these regions within the AhR cooperate when linked together, resulting in a synergistic activation of transcripti on. Fusion proteins of the AhR and ARNT trans-activation domains with the LexA DNA-binding domain, expressed in bacteria and purified to nea r-homogeneity, stimulated transcription of a minimal promoter in vitro in yeast nuclear extracts. Using this in vitro transcription assay, i t was also possible to demonstrate that the AhR and ARNT trans-activat ion domains, in the absence of a DNA-binding domain, inhibited activat ed and basal transcription. Furthermore, in vitro the receptor bound s electively to the basal transcription factors, the TATA-binding protei n and TFIIF, whereas ARNT bound preferentially to TFIIF. Taken togethe r, these results suggest that AhR and ARNT activate target gene expres sion, at least in part, through direct interactions with basal transcr iption factors.