THE AH RECEPTOR CAN BIND LIGAND IN THE ABSENCE OF RECEPTOR-ASSOCIATEDHEAT-SHOCK-PROTEIN-90

The Ah receptor (AhR) is a soluble ligand-dependent DNA regulatory pro tein that mediates many of the biological responses to 2,3,7,8-tetrach lorodibenzo-p-dioxin (TCDD, dioxin) and related chemicals. In the abse nce of ligand, the cytosolic form of the AhR is found complexed with a t least two molecules of hsp90, a heat shock protein of 90 kDa. In add ition to its role in AhR protein folding and ability to repress the in herent nuclear localization, dimerization, and DNA binding activity of the AhR, it has been reported that hsp90 is absolutely required for m aintaining the AhR in its high-affinity ligand binding conformation. T he ability of high salt conditions (0.4 M KCl) to dissociate the multi meric AhR complex into its monomeric form provides us with an avenue t o examine the role of hsp90 in AhR ligand binding activity. In contras t to previous reports, we demonstrate that salt-dissociated ''hsp90-fr ee'' AhR from several species still retains the ability to specificall y bind ligand ([H-3]TCDD). Although partial inactivation of ligand bin ding of salt-dissociated rat hepatic AhR was observed (to a maximum of 50% of total AhR binding), the presence of bound ligand protected aga inst this inactivation. Little or no inactivation of the ligand bindin g ability of salt-dissociated guinea pig or rabbit AhR occurred. Our r esults not only indicate a significant species-difference in AhR ligan d binding stability and/or activity, but also demonstrate that AhR lig and binding activity does not absolutely require the presence of recep tor-bound hsp90. a 1998 Academic Press.