The odorant receptor repertoire of teleost fish

Abstract

Background: Vertebrate odorant receptors comprise three types of G protein-coupled receptors: the OR, V1R and V2R receptors. The OR superfamily contains over 1,000 genes in some mammalian species, representing the largest gene superfamily in the mammalian genome.

Results: To facilitate an informed analysis of OR gene phylogeny, we identified the complete set of 143 OR genes in the zebrafish genome, as well as the OR repertoires in two pufferfish species, fugu (44 genes) and tetraodon (42 genes). Although the genomes analyzed here contain fewer genes than in mammalian species, the teleost OR genes can be grouped into a larger number of major clades, representing greater overall OR diversity in the fish.

Conclusion: Based on the phylogeny of fish and mammalian repertoires, we propose a model for OR gene evolution in which different ancestral OR genes or gene families were selectively lost or expanded in different vertebrate lineages. In addition, our calculations of the ratios of non-synonymous to synonymous codon substitutions among more recently expanding OR subgroups in zebrafish implicate residues that may be involved in odorant binding.

Figure 1

Chromosomal distribution of zebrafish OR genes. The majority of OR genes are organized in large clusters at only a few loci in the zebrafish genome. OR genes are depicted as boxes above (plus strand) or below (minus strand) a line representing each chromosome that encodes ORs. Genes are color-coded according to subfamily.

Figure 2

Phylogeny of zebrafish and other vertebrate OR families. (a) Phylogeny of zebrafish receptors. A neighbor joining tree was constructed based on an alignment of the predicted amino acid sequences of 143 intact genes and 4 full-length pseudogenes identified from the zebrafish genome [see Additional file 2]. OR genes are named by subfamily and colored by family. The eight gene families are labeled A-H. The zebrafish melanocortin receptor branch (dotted line labeled "mcr") indicates the root of the tree. Bootstrap scores for each family are indicated in parentheses. (b) Phylogenetic relationship among zebrafish and mouse odorant receptors. The following sets of genes were aligned and used to construct a tree by neighbor joining: the mouse odorant receptors, mORs [8,9]; the subset of 136 intact zebrafish ORs with no disruptions identified in this study (highlighted in red); and mouse melanocortin receptors (mcr). Note the presence of OR gene subfamilies OR112, OR113, and OR114 (Family A) within the Class I clade and zebrafish OR101-1 (Family B) within the Class II clade. Bootstrap scores corresponding to selected nodes are indicated.(c) Phylogeny of the complete OR repertoires of zebrafish, fugu and tetraodon identified in this study. One-hundred-thirty-six non-disrupted genes from zebrafish, 42 non-disrupted genes from fugu, and 42 non-disrupted genes from tetraodon were used in this neighbor joining analysis. Families are labeled A-H and correspond to the zebrafish families shown in panel A. Bootstrap scores corresponding to selected nodes are indicated.

Figure 3

Sequence logos of zebrafish and mouse OR families. Conservation of predicted amino acid sequence for the zebrafish (a) and mouse (b) OR repertoires is shown graphically (see the text). Y axis, information content. X axis, residue position. For this analysis, positions with gaps in more than 95% of sequences, as well as poorly aligned N- and C-terminal sequences, were removed. Positions in the species-specific logos are identical according to this alignment. The logo was generated from this alignment using the program WebLogo (G.E. Crooks, G. Hon, J.-M. Chandonia and S.E. Brenner, personal communication), available at .

Figure 4

Sites under positive and negative selection in OR coding sequences. Nucleotide alignments were generated from the corresponding amino acid alignment [see Additional file 2] after removal of N- and C-terminal sequences and gap removal with respect to OR124-3, and subjected to two analyses of selective pressure. (a) Analysis of dN/dS ratios by sub-regions of OR coding sequences indicates that OR genes in general are under negative or purifying selection (dN/dS < 1). However, analysis of informative pairwise comparisons reveals that transmembrane domains (TMs) 3–6, and to a lesser degree TM1, have significantly higher average pairwise dN/dS ratios when compared with the average for non-transmembrane coding sequence (asterisks; p < 1 × 10^-3). Error bars show standard errors of the means. (b) A schematic representation of OR124-3 (an example OR) with transmembrane domains one through seven shown from left to right. SLAC analysis reveals sites under positive selection (dN/dS > 1) with p < 0.1 (red), p < 0.2 (orange), p < 0.5 (yellow), and sites under negative selection (dN/dS < 1) with p < 0.05 (dark blue), p < 0.1 (light blue), p < 0.2 (turquoise), p < 0.5 (green). The null hypothesis is that a site is neutrally evolving with dN/dS = 1. Yellow and green residues may be considered neutrally evolving. Sites on the representative OR124-3 sequence with dN/dS > 1 and p < 0.1 are: V108 (dN/dS = 1.39), F145 (dN/dS = 1.52), E261 (dN/dS = 1.48) and T262 (dN/dS = 1.42). Snake plot generated using the RbDe web service [48].