LIGAND-DEPENDENT INTERACTION OF THE ARYL-HYDROCARBON RECEPTOR WITH A NOVEL IMMUNOPHILIN HOMOLOG IN-VIVO

In an effort to identify regulators of aryl hydrocarbon receptor (AHR) signaling, we have employed the yeast two-hybrid system to screen for human proteins that interact in a ligand-dependent manner with the AH R. After screening 1.4 x 10(6) clones from a human B cell library, two distinct clones were identified that associated specifically with the liganded receptor. No clones were identified that interacted preferen tially with the unliganded AHR. One of the ligand dependent clones, AR A9, encodes a novel 330-amino acid protein with regions of amino acid sequence similarity to the 52-kDa FK506-binding protein known to be as sociated with the glucocorticoid receptor. Yeast two-hybrid experiment s with ARA9 demonstrated a strong interaction with the AHR that is enh anced 11-fold in the presence of the ligand beta-naphthoflavone. In vi tro experiments using proteins generated in reticulocyte lysates confi rmed this interaction and indicated that ARA9 can be co-immunoprecipit ated with the AHR using antisera raised specifically for either the AH R or the 90-kDa heat shock protein. The observation that ARA9 has a hi gh affinity for both the 90-KDa heat shock protein-associated and liga nd-activated forms of the AHR suggests that ARA9 is a component of the AHR-signaling pathway in vivo.