Cloning and characterization of PRAX-1 - A new protein that specifically interacts with the peripheral benzodiazepine receptor

Using a cytoplasmic domain of the peripheral benzodiazepine receptor (PBR) as a bait in the yeast two-hybrid system, we have isolated a cDNA encoding a new protein that specifically interacts with PER. We named it PRAX-1, for peripheral benzodiazepine receptor-associated protein 1. PRAX-1 is a 1857- amino acid protein, the sequence of which was structurally unrelated to any known proteins. The gene encoding PRAX-1 is located in the q22-q23 region of the long arm of the human chromosome 17. The PRAX-1 mRNA is 7.5 kilobase pairs, predominantly expressed in the central nervous system, pituitary gl and, and thymus, At the protein level, we found the PRAX-1 as a single 220- 250-kDa protein in the brain and in many different human cell lines tested using specific antibody raised against PRAX-1, Parallel analysis of the PRA X-1 mRNA and protein expression performed in mouse and rat gave similar res ults. Immunocytochemistry analysis carried out to define the distribution o f the PRAX-1 protein in the rat brain showed that PRAX-1 was prevalent in t he mesolimbic system, specially abundant in the CA1 subfield of the hippoca mpus. Exhibiting several domains involved in protein-protein interaction (t hree proline-rich domains, three leucine-zipper motifs, and an Src homology region 3-like domain), the PRAX-1 may be looked upon as a new adaptator pr otein. We show that both the Src homology region 3-like domain and a prolin e-rich domain in PRAX-1 are required for the interaction with PER, PRAX-1 i s a cytoplasmic protein that also partially colocalizes with PER in the mit ochondria, as determined by confocal microscopy and Western blotting. Altog ether our observations support a model of interaction implicating PER and t his newly described protein, PRAX-1, As being the first cytoplasmic protein associated with PER, PRAX-1 is a new tool that opens new fields for explor ing PER biological roles.