Comparative genomics of odorant and pheromone receptor genes in rodents

Abstract

We applied a comprehensive data-mining strategy to examine the repertoires of rat and mouse odorant receptors (ORs) and type 1 pheromone receptors (V1Rs) using the mm5 (mouse) and rn3 (rat) genomes. We identified 1576 rat OR genes, including 292 pseudogenes. The rat V1R repertoire is composed of 115 intact genes and 72 pseudogenes. The mouse OR and V1R databases were updated using the new assembly mm5, from which 1375 mouse ORs and 308 V1Rs were identified, with more than 100 putative pseudogenes from mm2 now identified as intact because of the higher sequence quality. With these new data we have conducted a series of genomic analyses of the OR and V1R genes from mouse and rat. Orthologous OR clusters were identified in mouse and rat and comparison analysis was performed at three incremental levels: families, coding sequences, and motifs. At the family level, we found that V1R genes have more species-specific families than OR genes. About 20% of intact V1R genes have no orthologous counterpart in the same family, whereas less than 1% of intact ORs are similarly isolated. At the coding sequence level, OR genes are more conserved between mouse and rat than V1R genes. OR genes share greater similarity with their orthologous counterparts than with their closest neighbor, whereas V1R genes show the opposite tendency. Motifs were identified to obtain biological insights. Motifs specific for species or families were found in OR and V1R genes, which may result in the differential pheromone-dependent behaviors and perception of odors between mouse and rat.

Fig. 1. Chromosomal distribution of rat and mouse OR/V1R genes

Blue, intact OR genes; red, OR pseudogenes; green, intact V1R genes; purple, V1R pseudogenes. The number of OR/V1R genes per 1Mb is shown as bars on each chromosome. The height of each bar is proportional to the number of genes in that locus. (a) The number of OR/V1R genes on each chromosome of rat. “Un” represents the sequences unmapped in current rn3 assembly. There is no OR/V1R gene on either chromosome 18 or Y. (b) Rat chromosomes 1 and 4 are drawn according to the rn3 assembly. The cytogenetic map of each chromosome is shown under the scaffold assembly in scale. The number of OR/V1R genes per 1Mb is shown as bars on each chromosome. The black arrow points to one OR cluster that is very close to V1R genes, but not intermingled with them. Rat Class I OR cluster is indicated with a grey arrowhead. (c) The number of OR/V1R genes on each chromosome of mouse. “Un” represents the sequences from unmapped scaffolds of mm5 assembly. There are no OR genes on chromosome 18. (d) Mouse chromosome 6 and 7 are drawn according to the mm5 assembly. The cytogenetic map of each chromosome is shown under the scaffold assembly in scale. The number of OR/V1R genes per 1Mb is shown as bars on each chromosome. Mouse Class I OR cluster is indicated with a grey arrowhead.

Fig.2. Comparison of rodent ORs and V1Rs through phylogenetic analysis

At the family level, OR genes are more closely related in mouse and rat than V1R genes. All mouse and Rat ORs/V1Rs are pooled and classified into sub-families using the same criterion to define a family as applied to ORs/V1Rs from either species separately. OR/V1R subfamilies are represented in the middle of the figure by a line of small colored bars. ORs/V1Rs belonging to the same subfamily have the same color index. OR/V1R clusters are shown according to their genomic locations on the top and bottom with grey bars proportional to their width on the chromosome. Each OR/V1R gene is represented by one line inside the grey bar with its family color index. Isolated OR/V1R genes are drawn without grey cluster bars. Each cluster is named with the chromosome location and position(Mb) connected by an underscore. Thin grey lines from each cluster or isolated gene are connected to their subfamily in the middle. (a) Families of OR genes. To minimize the number of connection lines, only one line is drawn between the cluster bar and the subfamily bar if there is more than 1 OR from the same cluster belonging to the same subfamily. Orthologous clusters are highlighted with dark lines and Class I ORs are highlighted with blue lines. In total, mouse and rat ORs are divided into 245 families, among which 4 are specific for rat and 12 for mouse. In these rat or mouse specific families there are only 4 intact mouse ORs and 1 intact rat OR, which constitutes less than 1% of all putative functional ORs. Class I ORs, which are from rat cluster 1_160 and mouse cluster 7_90 are the most conserved in terms of genomic location. (b) Families of V1R genes. One grey line is draw from the relative position of each V1R gene in its genomic cluster to the subfamily that it belongs to. Species-specific families are highlighted with red (for rat) and green (for mouse) circles. 14 families are specific for rat, and 10 for mouse, in which there are 35 rat V1Rs and 79 mouse V1Rs, constituting about 20 percent of the functional V1R repertoire.

Fig.3. Similarity analysis of OR/V1R genes

Each gene is represented at the position of the similarity range with its most related sequence. A color index is assigned to each OR/V1R family defined from either species separately. Mouse or rat OR/V1R genes from the same family are in the same color. The height of each bar is proportional to the number of genes in that similarity range. OR genes appear on the left panels, and V1R genes on the right. (a) Comparison between the best orthologs from the other species. In each panel, genes from rat are on the top, and those from mouse are on the bottom. Over 70% of OR genes have orthologs with more than 80 percent identity, whereas less than 20% of V1Rs have orthologs in this range, indicating that OR genes are more conserved between mouse and rat than V1R genes. (b) Comparison between best paralogs from the same species. In the left panel, 52% of rat ORs and 52% of mouse ORs have a paralog with over 80 percent identity. In the right panel, 41% of rat V1Rs and 63% of mouse V1Rs have a paralog at the same level. From the left panel, OR genes are more conserved with their orthologs than with their paralogs. From the right, V1R genes are more similar with their paralogs than with their orthologs.

Fig. 4. Motif analysis of OR and V1R sequences

Recurring motifs were identified from the pool of mouse and rat intact ORs/V1Rs with the MEME program. This was followed by checking the identitiy of the support sequences in each species. Each motif is represented by a bar proportional to its width, positioned using OR I7 and VN3 as the references for the TMs (trans membrane domains). The color index varying from white to black indicates the conservation of each motif, measured as the number of support sequences. The motif numbers are listed on the top of each bar. Further details are shown in supplementary tables 3&4. Red and green circles highlight motifs that are specific for one species. (a) Motifs of OR genes. 100 motifs supported by more than 10 sequences are listed, none of which are specific for either mouse or rat. 10 motifs are specific for Class I ORs, and 59 are specific for Class II. The rest are shared by both classes. (b) Motifs of V1R genes. 83 motifs supported by more than 5 V1R sequences are listed. 12 of them reside only in mouse V1Rs and 3 of them reside only in rat V1Rs. The rest are shared by both species.