Molecular evolution of neuropeptide Y receptors in vertebrates (Lampetra fluviatilis, Squalus acanthias, Danio rerio)

The three evolutionarily related peptides neuropeptide Y (NPY), peptide YY (PYY) and pancreatic polypeptide (PP) are ligands to at least five G-protein coupled receptors in mammals, which are denoted by numbers. NPY has many physiological effects including stimulation of appetite and regulation of circadian rhythm and blood pressure. This work describes the ancient origin of the NPY receptor genes as deduced from molecular cloning of six receptors in four distantly related vertebrate species. Three of the receptors have been functionally expressed in vitro to determine ligand binding properties. The first Y2 receptor from any non-mammalian species was cloned from the chicken. The receptor was found to exhibit substantial structural and pharmacological differences to mammalian Y2, but showed similar anatomical distribution. A receptor was cloned in a primitive vertebrate, an agnathan fish, the river lamprey Lampetra fluviatilis. Phylogenetic analyses indicated that it represents an orthologue to the ancestor of Y4 and the teleost subtypes Yb and Yc. Three NPY receptors were cloned from a shark, the spiny dogfish Squalus acanthias . These were found to correspond to the three mammalian subtypes Y1, Y4 and y6, and was thereby the first complete Y1 subfamily in any species outside the mammalian lineage. This suggests that all three receptor subtypes arose in the common ancestor of sharks and mammals 420–450 million years ago. The sixth described receptor was cloned from the zebrafish, Danio rerio , and was shown to have equal identity to all three mammalian Y1 subfamily receptors. Phylogenetic analyses including the shark and lamprey sequences suggested that Yb may represent a fourth Y1 subfamily gene. It has previously been found that the genes for Y1, Y4 and y6 are located on separate chromosomes. Taken together, these results show that the NPY receptor family expanded by chromosomal duplications early in vertebrate evolution, prior to the origin of gnathostomes. This work will be important for the determination of the time points for the origin of the many functions of NPY as well as for the understanding of the processes that shaped the vertebrate genome.