GPR50 is the mammalian ortholog of Mel1c: evidence of rapid evolution in mammals

Abstract

Background: The melatonin receptor subfamily contains three members Mel1a, Mel1b and Mel1c, found in all vertebrates except for Mel1c which is found only in fish, Xenopus species and the chicken. Another receptor, the melatonin related receptor known as GPR50, found exclusively in mammals and later identified as a member of the melatonin receptor subfamily because of its identity to the three melatonin receptors despite its absence of affinity for melatonin. The aim of this study was to describe the evolutionary relationships between GPR50 and the three other members of the melatonin receptor subfamily.

Results: Using an in silico approach, we demonstrated that GPR50 is the ortholog of the high affinity Mel1c receptor. It was necessary to also study the synteny of this gene to reach this conclusion because classical mathematical models that estimate orthology and build phylogenetic trees were not sufficient. The receptor has been deeply remodelled through evolution by the mutation of numerous amino acids and by the addition of a long C-terminal tail. These alterations have modified its affinity for melatonin and probably affected its interactions with the other two known melatonin receptors MT1 and MT2 that are encoded by Mel1a and Mel1b genes respectively. Evolutionary studies provided evidence that the GPR50 group evolved under different selective pressure as compared to the orthologous groups Me11 a, b, and c.

Conclusion: This study demonstrated that there are only three members in the melatonin receptor subfamily with one of them (Me11c) undergoing rapid evolution from fishes and birds to mammals. Further studies are necessary to investigate the physiological roles of this receptor.

Figure 1

Phylogenetic analysis of the GPR50/MT1/MT2/Mel1c genes. (A) Overall phylogenetic tree showing 3 groups of genes: GPR50, MT1/MT2, and Mel1c genes and the animal orders where each branch is expressed. The trees (npl) are the fusion of three phylogenetic trees built based on Neighbour joining, maximum Parsimony and maximum Likelihood (see "Materials and Methods" section for further details). The italic letters correspond to the name given to the branches for the likelihood ratio tests (B) Phylogenetic tree of GPR50 genes. Please note that only mammalian species appear in the tree. (C) Phylogenetic tree of MT1 genes (D) Phylogenetic tree of MT2 genes. (E) Phylogenetic tree of Mel1c genes that do seem to appear only in non mammalian species. Bootstrap values are reported for each npl method.

Figure 2

Synteny of Mel1c/GPR50 genes and neighbours in vertebrate genomes. Note that genes are found on chromosome 5 in zebra fish and on chromosome 4 in chicken while they are found on chromosome X in other depicted species. Please note that synteny is mostly conserved for bHLHPAS, 2610030H06 RIK, Mel1c, HMG2A, CD99, and myotubularin related protein in opossum and mammalian species despite the integration of new genes coding for hypothetical proteins (opossum, chimpanzee, cow), ribosomal proteins (dog, chimpanzee, man), NGFI-A binding protein (chimpanzee, man), Utbf (mouse) and MAGE (cattle) proteins. It is also of note that several genes surrounding Mel1c in zebra fish (pdcd8, nono, and the two hypothetical proteins) present high identities with genes found on chromosome X in mouse but not in the GPR50 locus (unpublished data). p.d.: predicted gene. Chrm: chromosome.

Figure 3

Sequence alignment of human MT1, MT2 and GPR50 with bovine rhodopsin (pdb 1F88). Sequence identities are reported white on a black background, whereas sequence similarities are boxed (A). The positions of the transmembrane helices, as observed in the bovine rhodopsin structure, are reported above its sequence. Arrows indicate the positions of the amino acids that, in GPR50, evolved under positive selection. Stars indicate amino acids which have been shown to play a key role for melatonin binding in MT1 (dark blue), MT2 (light blue) or both (red). A ribbon representation of the GPR50 3D structure model is represented (B), with transmembrane helices colored according to the sequence alignment. Amino acids evolving under positive selection and amino acids important for melatonin binding in MT1/MT2 are shown according to the colors reported in the sequence alignment.

Figure 4

Sequence alignment of chicken Mel1c, zebra fish Mel1c and human GPR50. Sequence identities are reported white on a black background, whereas sequence similarities are boxed. The positions of the transmembrane helices are reported above its sequence. Arrows indicate the positions of the amino acids that, in GPR50, evolved under positive selection.

Figure 5

Schematic diagram of the Mel1c/GPR50 gene organization in zebrafish, chicken, opossum, mouse, man, and horse. The stop codon following the second exon in zebrafish and chicken is replaced by the insertion of a protein fragment reminiscent of a DNA directed RNA polymerase II in mammals (light color).

Figure 6

Alignment of the repeated heptad found in the C-terminal extension of human GPR50 and comparison with the repeated heptad observed in the C-terminal domain (CTD) of RNA polymerase II (RNAPII). The three positions (2, 5 and 7) occupied by phosphorylable serine residues in RNAPII CTD are boxed.