Ancient genomic architecture for mammalian olfactory receptor clusters

Abstract

Background: Mammalian olfactory receptor (OR) genes reside in numerous genomic clusters of up to several dozen genes. Whole-genome sequence alignment nets of five mammals allow their comprehensive comparison, aimed at reconstructing the ancestral olfactory subgenome.

Results: We developed a new and general tool for genome-wide definition of genomic gene clusters conserved in multiple species. Syntenic orthologs, defined as gene pairs showing conservation of both genomic location and coding sequence, were subjected to a graph theory algorithm for discovering CLICs (clusters in conservation). When applied to ORs in five mammals, including the marsupial opossum, more than 90% of the OR genes were found within a framework of 48 multi-species CLICs, invoking a general conservation of gene order and composition. A detailed analysis of individual CLICs revealed multiple differences among species, interpretable through species-specific genomic rearrangements and reflecting complex mammalian evolutionary dynamics. One significant instance involves CLIC #1, which lacks a human member, implying the human-specific deletion of an OR cluster, whose mouse counterpart has been tentatively associated with isovaleric acid odorant detection.

Conclusion: The identified multi-species CLICs demonstrate that most of the mammalian OR clusters have a common ancestry, preceding the split between marsupials and placental mammals. However, only two of these CLICs were capable of incorporating chicken OR genes, parsimoniously implying that all other CLICs emerged subsequent to the avian-mammalian divergence.

Figure 1

Conservation of synteny of OR genes. (a) All ORs from each species are ordered along the axis according to their genomic location from chromosome 1 to X (or by scaffold number in the case of the opossum), and by the internal megabase coordinates in each chromosome. Each point represents an alignment between two ORs from different species in the UCSC alignment net, colored according to the degree of DNA sequence identity (x-axis for the reference species, y-axis for the target species). Diagonals in both directions represent conservation of gene order, whereas reverse diagonals indicate a reverse of gene order relative to the 'plus' DNA strand. Off-diagonal points generally indicate micro-rearrangements, but those that are associated with low percentage identity possibly represent alignment errors. (b) Zoomed human versus mouse comparison, with chain numbers (by UCSC hg17 versus mm6 alignment net) indicated for the 16 alignment chains that contain at least six pairs of syntenic orthologs. Chains #95 and #183 represent disrupted synteny, because the alignment of a succession of ORs from human chromosome 6 is split between mouse chromosomes 13 and 17 (as described by Amadou and coworkers [26]). Chains #375 and #118 capture a genomic inversion. OR, olfactory receptor.

Figure 2

CLIC statistics. (a) Different types of CLICs are characterized by the number of species involved. The fraction of opossum-specific CLICs is indicated by light gray. (b) The total number of genes in CLICs from each type. The opossum-specific fraction is indicated as in panel a. (c) Cumulative plots show the fraction of OR genes that is covered by multi-species CLICs of decreasing size (sorted first according to the number of genes in human, and then by the numbers in mouse, rat, dog, and finally opossum). All multi-species CLICs together cover more than 95% of any eutherian OR repertoire (solid black = human, dashed dark gray = mouse, dashed light gray = rat, solid light gray = dog), but only two-thirds of the opossum repertoire (solid dark gray). The coverage of the combined repertoire of all species is shown by black circles. (d) The total number of clusters included in CLICs from each type and size. CLIC, clusters in conservation.

Figure 3

CLICs of OR genes. (a) CLIC (columns) are shown by human genomic order (see Table 3), with human chromosome numbers indicated (top ticked line). For CLICs that do not contain human clusters, the order was determined by the human location that is syntenic to the region of the mouse OR cluster (Additional data file 1). For each species (h = human, m = mouse, r = rat, d = dog, o = opossum, c = chicken, n = consensus gene count) circle size is proportional to log2(n - 1), where n is the number of genes in the OR clusters within the CLIC. All multi-species CLICs are enumerated (#i at bottom); nonhuman single species CLICs are not shown. (b-d) Detailed depiction of three CLICs indicated by the corresponding capital letter above the CLIC column in panel a. To the left of panels b-d, clusters are represented by circles (colored for species, as in panel a), with gene count indicated. Lines connect every two clusters sharing syntenic orthologs. To the right of panels b-d are schematic genomic representations of the clusters, with OR gene groups in species color and OR family indicated. Grey bars represent flanking non-OR genes (HUGO nomenclature symbols indicated [57]); TRA@ is the T-cell receptor alpha locus. Multiple rows for the same species indicate the inclusion of clusters from multiple chromosomes in the CLIC. A break in local or large-scale synteny is marked by a broken line. For the complete list of the genomic coordinates of all analyzed genes, see Additional data file 2. CLIC, clusters in conservation; OR, olfactory receptor.

Figure 4

The reconstruction of an ancestral cluster. (a) Detailed comparison of gene content and order is shown for the four clusters included in CLIC #39 (see Table 3), containing genes from the OR1 and OR3 families. A schematic ancestral configuration is shown at the bottom row, with inferred minimal number of genes. Genes are presented as triangles colored by subfamily affiliation (bottom row; yellow for subfamily OR1G). Inferred chromosomal rearrangements relative to the ancestor are specified for each species: circle = insertion, X = deletion, arrow = inversion, broken line with number of genes = tandem duplication. A complex duplication in rodent genomes, involving subfamilies OR3A, OR1P, and OR1A, is marked with a rectangle. This duplication was probably formed via several events, some of which occurred after the split between mouse and rat lineages. The same region had experienced another independent event in the dog genome, in which three genes from subfamilies 1A and 1P were inverted as one unit. Tandem duplication in one end of the rodent clusters forms a series of numerous adjacent highly similar genes from the same subfamily (OR1J). The human and mouse orthologous clusters were studied and compared previously [27], and a complex orthology relationship among the genes was described. (b) A phylogenetic tree of CLIC #39 ORs from which the ancestral cluster gene count can be inferred. The phylogenetic tree was generated with Mega version 3.1 [58] using ME algorithm, and Poisson correction for distance calculation. Protein sequences were aligned with Clustalx [59]. The colors of circles next to the phylogenetic branches indicate species (blue = dog, green = rat, orange = mouse, red = human). OR51D1 serves as an out-group. CLIC, clusters in conservation; OR, olfactory receptor.