Origin and molecular evolution of LPXRFamide peptides and PQRFamide peptides in the brain of vertebrates

Abstract

Among the RFamide peptide groups, PQRFamide peptides, such as neuropeptide FF (NPFF) and NPAF, share a common C-terminal Pro-Gln-Arg-Phe-NH2 motif. LPXRFamide (X = L or Q) peptides, such as gonadotropin-inhibitory hormone (GnIH), frog growth hormone-releasing peptide (fGRP), goldfish LPXRFamide peptide and mammalian RFamide-related peptides (RFRPs), share a C-terminal Leu-Pro-Leu/GlnArg-Phe-NH2 motif. Such a similar C-terminal structure suggests that these two groups may have diverged from a common ancestral gene. In this study, I sought to clarify the evolutionary origin and divergence of these two groups, by identifying novel RFamide peptides from the brain of lower vertebrates. At the start, I used the brain of lamprey (Petromyzon marinus), one of the only two extant representative species of the oldest lineage of vertebrates, the Agnatha, which diverged from the main line of vertebrate evolution approximately 450 million years ago. A novel RFamide peptide was identified by immunoaffinity purification using the antiserum against LPXRFamide peptide. The identified RFamide peptide did not contain a C-terminal LPXRFamide motif, but had the sequence SWGAPAEKFWMRAMPQRFamide (lamprey PQRFa). A cDNA of the precursor encoded one lamprey PQRFa and two related peptides. These related peptides, which also had the Cterminal PQRFamide motif, were further identified as mature endogenous ligands. Phylogenetic analysis revealed that lamprey PQRFamide peptide precursor belongs to the PQRFamide peptide group. In situ hybridization demonstrated that lamprey PQRFamide peptide mRNA is expressed in the regions predicted to be involved in neuroendocrine and behavioral functions. This is the first demonstration of the presence of RFamide peptides in the agnathan brain. Lamprey PQRFamide peptides are considered to have retained the most ancestral features of PQRFamide peptides. On the other 55