IDENTIFICATION OF FUNCTIONAL DOMAINS OF THE ARYL-HYDROCARBON RECEPTOR

Functional domains of the mouse aryl hydrocarbon receptor (Ahr) were i nvestigated by deletion analysis. Ligand binding was localized to a re gion encompassing the PAS B repeat. The ligand-mediated dissociation o f Ahr from the 90-kDa heat shock protein (HSP90) does not require the aryl hydrocarbon receptor nuclear translocator (Amt), but it is slight ly enhanced by this protein. One HSP90 molecule appears to bind within the PBS region. The other molecule of HSP90 appears to require intera ction at two sites: one over the basic helix-loop-helix region, and th e other located within the PAS region. Each mutant was analyzed for di merization with full-length mouse Amt and subsequent binding of the di mer to the xenobiotic responsive element (XRE). In order to minimize a ny artificial steric hindrances to dimerization and XRE binding, each Ahr mutant was also tested with an equivalently deleted Arnt mutant. T he basic region of Ahr is required for XRE binding but not for dimeriz ation. Both the first and second helices of the basic helix-loop-helix motif and the PAS region are required for dimerization. These last re sults are analogous to those previously obtained for Arnt (Reisz-Porsz asz, S., Probst, M.R., Fukunaga, B. N., and Hankinson, O. (1994) Mol. Cell. Biol. 14, 6075-6086) compatible with the notion that equivalent regions of Ahr and Amt associate with each other. Deletion of the carb oxyl-terminal half of Ahr does not affect dimerization or XRE binding but, in contrast to an equivalent deletion of Arnt, eliminates biologi cal activity as assessed by an in vitro transcriptional activation ass ay, suggesting that this region of Ahr plays a more prominent role in transcriptional activation of the cyp1a1 gene than the corresponding r egion of Amt.