A candidate subspecies discrimination system involving a vomeronasal receptor gene with different alleles fixed in M. m. domesticus and M. m. musculus

Abstract

Assortative mating, a potentially efficient prezygotic reproductive barrier, may prevent loss of genetic potential by avoiding the production of unfit hybrids (i.e., because of hybrid infertility or hybrid breakdown) that occur at regions of secondary contact between incipient species. In the case of the mouse hybrid zone, where two subspecies of Mus musculus (M. m. domesticus and M. m. musculus) meet and exchange genes to a limited extent, assortative mating requires a means of subspecies recognition. We based the work reported here on the hypothesis that, if there is a pheromone sufficiently diverged between M. m. domesticus and M. m. musculus to mediate subspecies recognition, then that process must also require a specific receptor(s), also sufficiently diverged between the subspecies, to receive the signal and elicit an assortative mating response. We studied the mouse V1R genes, which encode a large family of receptors in the vomeronasal organ (VNO), by screening Perlegen SNP data and identified one, Vmn1r67, with 24 fixed SNP differences most of which (15/24) are nonsynonymous nucleotide substitutions between M. m. domesticus and M. m. musculus. We observed substantial linkage disequilibrium (LD) between Vmn1r67 and Abpa27, a mouse salivary androgen-binding protein gene that encodes a proteinaceous pheromone (ABP) capable of mediating assortative mating, perhaps in conjunction with its bound small lipophilic ligand. The LD we observed is likely a case of association rather than residual physical linkage from a very recent selective sweep, because an intervening gene, Vmn1r71, shows significant intra(sub)specific polymorphism but no inter(sub)specific divergence in its nucleotide sequence. We discuss alternative explanations of these observations, for example that Abpa27 and Vmn1r67 are coevolving as signal and receptor to reinforce subspecies hybridization barriers or that the unusually divergent Vmn1r67 allele was not a product of fast positive selection, but was derived from an introgressed allele, possibly from Mus spretus.

Figure 1. A comparison of polymorphism and divergence in two V1R genes.

Reduced data sets summarizing SNP differences for Vmn1r67 (Panel A) and the closely linked gene Vmn1r71 (Panel B) in five wild-derived inbred strains each for M. m. domesticus and M. m. musculus. Strain abbreviations are: LEW = LEWES; PER = PERA; TIR = TIRANO; ZAL = ZALENDE; CZ1 = CZECHI; CZ2 = CZECHII; and SKI = SKIVE. Fixed differences (divergence) between M. m. domesticus and M. m. musculus are highlighted in green (there are 24 in Vmn1r67 and none in Vmn1r71) and polymorphisms in both genes in the two subspecies are highlighted in gray to facilitate comparison. In addition, nonsynonymous sites in Vmn1r67 are indicated by yellow highlighting of the site numbers. Nucleotide diversity (π) and nucleotide polymorphism (θ) were calculated for both subspecies using DNAsp and are shown for each dataset. Full sequence data appear in Files S2 and S3.

Figure 2. A comparison of a murid rodent phylogeny and a V1R gene phylogeny.

Panel A: A canonical phylogeny of murid rodents (adapted from [57]); Panel B: A Vmn1r67 gene phylogeny for murid rodents using F84 nucleotide distances and neighbor-joining, with bootstrap values expressed as percentages (see Methods). Full sequence data appear in File S4.

Figure 3. Amino acid differences in a V1R gene between M. m. domesticus and M. m. musculus.

Diagrammatic representation of the amino acid sequence of Vmn1r67 (Panel A) and the three-dimensional structure on which it was threaded (Panel B); amino acids in Panel A that were not threaded on the d1ln6a model in Panel B are shaded gray. The amino acid positions affected by fifteen nonsynonymous differences fixed between M. m. domesticus and M. m. musculus (File S6) are shown as diamonds. Fourteen nonsynonymous sites detected by a test for homoplasy (File S5) are colored red. For orientation to the three-dimensional models in Panel B, the first helix is colored light orange. Panel B views are 1) front, 2) rear, 3) left side, 4) right side, 5) top and 6) bottom.

Figure 4. Linkage disequilibrium (LD) between Vmn1r67 and Abpa27, with the gene Vmn1r71 mapping between them.

The proximal end of mouse chromosome 7 shown at the top of the figure represents the relative positions of the three genes which appear diagrammed as gray-shaded blocks below the chromosome and over their portion of the LD diagram. A comparison of the degree of LD, represented as r² values, appears in the triangle at the bottom of the figure and a shading scale for LD minimum to maximum appears under the LD diagram. The wild-derived inbred mouse strains used in this study are numbered on the left side of the LD diagram with a strain legend immediately below. The genes Vmn1r67 and Abpa27 show strong LD, while Vmn1r71 shows no significant LD with either of them.