Analysis of cattle olfactory subgenome: the first detail study on the characteristics of the complete olfactory receptor repertoire of a ruminant

BACKGROUND: Mammalian olfactory receptors (ORs) are encoded by the largest mammalian multigene family. Understanding the OR gene repertoire in the cattle genome could lead to link the effects of genetic differences in these genes to variations in olfaction in cattle. RESULTS: We report here a whole genome analysis of the olfactory receptor genes of Bos taurus using conserved OR gene-specific motifs and known OR protein sequences from diverse species. Our analysis, using the current cattle genome assembly UMD 3.1 covering 99.9% of the cattle genome, shows that the cattle genome contains 1,071 OR-related sequences including 881 functional, 190 pseudo, and 352 partial OR sequences. The OR genes are located in 49 clusters on 26 cattle chromosomes. We classified them into 18 families consisting of 4 Class I and 14 Class II families and these were further grouped into 272 subfamilies. Comparative analyses of the OR genes of cattle, pigs, humans, mice, and dogs showed that 6.0% (n = 53) of functional OR cattle genes were species-specific. We also showed that significant copy number variations are present in the OR repertoire of the cattle from the analysis of 10 selected OR genes. CONCLUSION: Our analysis revealed the almost complete OR gene repertoire from an individual cattle genome. Though the number of OR genes were lower than in pigs, the analysis of the genetic system of cattle ORs showed close similarities to that of the pig

Genetic Diversity and mRNA Expression of Porcine <i>MHC</i> Class I Chain-Related 2 (<i>SLA-MIC2</i>) Gene and Development of a High-Resolution Typing Method

<div><p>The genetic structure and function of MHC class I chain-related (<i>MIC</i>) genes in the pig genome have not been well characterized, and show discordance in available data. Therefore, we have experimentally characterized the exon-intron structure and functional copy expression pattern of the pig <i>MIC</i> gene, <i>SLA-MIC2</i>. We have also studied the genetic diversity of <i>SLA-MIC2</i> from seven different breeds using a high-resolution genomic sequence-based typing (GSBT) method. Our results showed that the <i>SLA-MIC2</i> gene has a similar molecular organization as the human and cattle orthologs, and is expressed in only a few tissues including the small intestine, lung, and heart. A total of fifteen <i>SLA-MIC2</i> alleles were identified from typing 145 animals, ten of which were previously unreported. Our analysis showed that the previously reported and tentatively named <i>SLA-MIC2*05</i>, <i>07</i>, and <i>01</i> alleles occurred most frequently. The observed heterozygosity varied from 0.26 to 0.73 among breeds. The number of alleles of the <i>SLA-MIC2</i> gene in pigs is somewhat lower compared to the number of alleles of the porcine <i>MHC</i> class I and II genes; however, the level of heterozygosity was similar. Our results indicate the comprehensiveness of using genomic DNA-based typing for the systemic study of the <i>SLA-MIC2</i> gene. The method developed for this study, as well as the detailed information that was obtained, could serve as fundamental tools for understanding the influence of the <i>SLA-MIC2</i> gene on porcine immune responses.</p></div

Comparison of amino acid sequences of MIC genes among pigs, humans, and cattle.

<p>A representative sequence of each functional <i>MIC</i> gene from each species was selected, and amino acid sequences were compared throughout the entire coding region to evaluate sequence conservation. The accession numbers for the sequences are <i>BoLA-MIC1</i> (BK006541), <i>BoLA-MIC2</i> (BK006542), and <i>BoLA-MIC3</i> (BK006543) for cattle, and <i>MICA</i> (NM_000247) and <i>MICB</i>- (NM_005931) for humans. Potential sites for N-linked glycosylation are underlined, and cysteine residues are indicated in squares for <i>SLA-MIC2</i>. Gaps are indicated by dashes and identical residues are indicated by dots. Stars above the sequences indicate conserved N-linked glycosylation sites, and plus signs above the sequences indicate a cysteine residue that is conserved across species. The starting points of protein domains are indicated above the annotated sequence, and the numbers above the sequence indicate the number of amino acids starting from the α1 domain excluding the leader peptide.</p

Comparison of the mRNA expression levels of <i>SLA-MIC2</i> in various pig tissues.

<p><b>a)</b> The RT-PCR products (1080 bp) of <i>SLA-MIC2</i> exons 2 to 6 in different tissues, amplified from RNA isolated from nine-week-old male pigs. <b>b)</b> Standard <i>GAPDH</i> gene expression levels in different tissues, as visualized by the intensity of RT-PCR product staining on an agarose gel. <b>c)</b> The photodensity ratios between the amplified <i>SLA-MIC2</i> and <i>GAPDH</i> products.</p

Amino acid sequence similarities between swine leukocyte antigen <i>SLA</i>-<i>MIC2</i> and <i>MIC</i> orthologs of humans and cattle.

<p>The different protein domains are indicated below the x-axis. <i>MICA</i> and <i>MICB</i> are from humans; <i>BoLA-MIC1</i>, <i>2</i>, and <i>3</i> are from cattle.</p

Differences in porcine <i>MIC2</i> heterozygosity among seven breeds of pigs.

<p>Note: Het-O: observed heterozygosity; Het-E: Nei’s expected heterozygosity; ne: effective number of alleles; HWE shows P-value for heterozygous protein</p><p>S deficiency from the Hardy–Weinberg equilibrium likelihood ratio test</p><p>**P < 0.00</p><p>^aaverage observed heterozygosity</p><p>^baverage expected heterozygosity</p><p>Differences in porcine <i>MIC2</i> heterozygosity among seven breeds of pigs.</p

General strategy of genomic sequence-based genotyping for pig <i>SLA-MIC2</i>.

<p>The diagram shows the location of each primer for PCR and sequencing. The sizes (bp) of introns and exons are indicated.</p

A phylogenetic tree showing the relationships of <i>MIC</i> orthologous genes in different mammals including pigs (<i>MIC2</i>), cattle (<i>BoLA-MIC1</i>, <i>2</i>, and <i>3</i>), humans (<i>MICA</i> and <i>B</i>), chimpanzees (<i>Patr-MICA/B</i>), rhesus macaques (<i>Mamu-MIC1</i> and <i>2</i>), and mice and rats (<i>Mr1</i>).

<p>A phylogenetic tree was constructed using the sequences corresponding to <i>MIC</i> exons 2, 3, and 4 using the neighbor joining method. The numbers on the nodes indicate the bootstrap values above 50% (n = 1000). The accession numbers of sequences are indicated in parentheses. <i>SLA-1</i>*<i>0401</i>(AF464016), one of the most common <i>SLA</i> molecules of swine, was used as an out-group. Bar below the tree indicates distance scale.</p