Abstract

The pancreatic polypeptide-fold family of peptides consists of three 36-amino acid peptides, namely neuropeptide Y (NPY), peptide YY, and pancreatic polypeptide (PP). These peptides regulate important functions, including food intake, circadian rhythms, mood, blood pressure, intestinal secretion, and gut motility, through four receptors: Y1, Y2, Y4, and Y5. Additional receptor subtypes have been proposed based on pharmacology observed in native tissues. Recent studies with other G-protein-coupled receptors have shown that homo- and heterodimerization may be important in determining receptor function and pharmacology. In the present study, the recently cloned rhesus (rh) Y4 receptor was evaluated using radioligand binding, and the pharmacological profile was found to be very similar to the human Y4 receptor. To study homo- and heterodimerization involving the Y4 receptor using bioluminescence resonance energy transfer 2 (BRET²), the carboxy termini of the rhesus Y1, Y2, Y4, and Y5 receptors were fused to Renilla luciferase, and rhY4 was also fused to green fluorescent protein. Dimerization was also studied using Western blot analysis. Using both BRET² and Western analysis, we found that the rhY4 receptor is present at the cell surface as a homodimer. Furthermore, agonist stimulation using the Y4-selective agonists PP and 1229U91 can dissociate these dimers in a concentration-dependent manner. In contrast, rhY4 did not heterodimerize with other members of the NPY receptor family or with human opioid δ and μ receptors. Therefore, homodimerization is an important component in the regulation of the Y4 receptor.