S. Sigurdardóttir

Linkage relationships in the bovine MHC region. High recombination frequency between class II subregions.

Class II genes of the bovine major histocompatibility complex (MHC) have been investigated by Southern blot analysis using human DNA probes. Previous studies revealed the presence of bovine DOβ, DQα, DQβ, DRαand DRβgenes, and restriction fragment length polymorphisms for each of these genes were documented. In the present study, the presence of three additional class II genes, designated DZα, DYα, and DYβ, are reported. DZαwas assumed to correspond to the human DZαgene while the other two were designated DY because their relationship to human class II genes could not be firmly established. The linkage relationships among bovine class II genes and two additional loci, TCP1B and C4, were investigated by family segregation analysis and analysis of linkage disequilibrium. The results clearly indicated that all these loci belong to the same linkage group. This linkage group is divided into two subregions separated by a fairly high recombination frequency. One region includes the C4, DQα, DQβ, DRαand DRβloci and the other one is composed of the DOβDYα, DYβ, and TCPIB loci. No recombinant was observed within any of these subregions and there was a strong or fairly strong linkage disequilibrium between loci within groups. In contrast, as many as five recombinants among three different families were detected in the interval between these subregions giving a recombination frequency estimate of 0.17 ± 0.07. The fairly high recombination frequency observed between class 11 genes in cattle is strikingly different from the corresponding recombination estimates in man and mouse. The finding implies either a much larger molecular distance between some of the bovine class II genes or alternatively the presence of a recombinational “hot spot” in the bovine class II region.

Evolution of MHC polymorphism: extensive sharing of polymorphic sequence motifs between human and bovine DRB alleles.

The evolution ofMHC polymorphism has been studied by comparing the amino acid and nucleotide sequences of 14 bovine and 32 humanDRB alleles. The comparison revealed an extensive sharing of polymorphic sequence motifs in the two species. Almost identical sets of residues were found at several highly polymorphic amino acid positions in the putative antigen recognition site. Consequently, certain bovine alleles were found to be more similar to certain human alleles than to other bovine alleles. In contrast, the frequencies of silent nucleotide substitutions were found to be much higher in comparisons between species than within species implying that none of the human or bovine DRB alleles originated before the divergence of these distantly related species. The results suggest that the observed similarity inDRB polymorphism is due to convergent evolution and possibly the sharing of short ancestral sequence motifs. However, the relative role of the latter mechanism is difficult to assess due to the biased base composition in the first domain exon of polymorphic class 11 β genes. The frequency of silent substitutions betweenDRB alleles was markedly lower in cattle than in man suggesting that theDRB diversity has evolved more rapidly in the former species.

Gene duplications and sequence polymorphism of bovine class II DQB genes

The genetic diversity of bovine class II DQB genes was investigated by polymerase chain reaction amplification and DNA sequencing. The first domain exon was amplified from genomic DNA samples representing 14 class II haplotypes, defined by restriction fragment length polymorphism (RFLP) analysis. The presence of a polymorphism in the copy number of DQB genes was confirmed since two DQB sequences were isolated from certain haplotypes. Four subtypes of bovine DQB genes were found. DQB1 is the major type and was found in almost all haplotypes. DQB2 is very similar DQB1 but was found only in the duplicated haplotypes DQ9 to 12. DQB3 and DQB4 are two quite divergent genes only present in certain duplicated haplotypes. The bovine DQB complexity thus resembles that in the human DRB region. Bovine DQB genes were found to be highly polymorphic as ten DQB1 alleles and four DQB2 alleles were identified. The observed sequence polymorphism correlated well with previously defined DQB RFLPs. Bovine and human DQB alleles show striking similarities at the amino acid level. In contrast, the frequency of silent substitutions is much higher in comparisons of DQB alleles between species than within species ruling out the possibility that any of the contemporary DQB alleles have been maintained since the divergence of humans and cattle. The frequency of silent substitutions between DQB alleles was markedly lower in cattle than in humans, in agreement with a previous comparison of human and bovine DRB alleles.

Simplifying genetic locus assignment of HLA-DRB genes

The DR haplotypes of the human major histocompatibility complex have been arranged in five haplotypic groups based on genomic cloning and sequence analyses. To date, the expressed DRB sequences have been assigned to four different loci: DRB1, 3, 4 and 5. DRB1 alleles are present in all haplotypes, whereas DRB3, 4 and 5 are present only in some haplotypes. Here, Göran Andersson and colleagues suggest that DRB3, 4 and 5 sequences may be treated as a single allelic series. They argue that such a model is appropriate, since DRB3, 4 and 5 sequences are inherited in an allelic fashion, have similar genomic localization, exhibit similar levels of gene expression and are, with a few rare exceptions, not present in the same haplotype.

Primate DRB genes from the DR3 and DR8 haplotypes contain ERV9 LTR elements at identical positions

The HLA-DRB genes of the human major histocompatibility complex constitute a multigene family with a varying number of DRB genes in different haplotypes. To gain further knowledge concerning the evolutionary relationship, the complete nucleotide sequence was determined for a region spanning introns 4 and 5 of the three DRB genes (DRB1*0301, DRB2 and DRB3*0101) from a DR52 haplotype and the single DRB gene (DRB1*08021) in the DR8 haplotype. These analyses identified an endogenous retroviral long terminal repeat element (ERV9 LTR3), inserted at identical positions in intron 5 of the functional DRB genes in these two haplotypes. Comparison of the nucleotide sequence from introns 4 and 5 including the ERV9 LTR elements revealed a strong similarity between the three expressed DRB genes. The DRB3*0101 and DRB1*08021 genes were most similar in this comparison. These findings provide further evidence for a separate duplication in a primordial DR52 haplotype followed by a gene contraction event in the DR8 haplotype. A homologous element was found in a chimpanzee DRB gene from a DR52 haplotype. This represents the first characterized ERV9 LTR element in a nonhuman species. The corresponding introns of the DRB genes in the DR4 haplotype contain no ERV9 LTRs. In contrast, these genes have insertions of distinct Alu repeats, implying distinct evolutionary histories of DR52 and DR53 haplotypes, respectively. Phylogenetic analyses of DRB introns from DR52, DR53, and DR8 haplotypes showed a close relationship between the DRB2 and DRB4 genes. Thus, the ancestral DR haplotype that evolved to generate the DR52 and DR53 haplotypes most likely shared a primordial common DRB gene.

A Phylogenetic Investigation of MHC Class II DRB Genes Reveals Convergent Evolution in the Antigen Binding Site

The evolution of Mhc class II polymorphism has been studied by comparing the nucleotide and amino sequences of a number of pig, cattle, horse, dog, rodent, and human DRB alleles. Very similar sets of residues were found at several highly polymorphic amino acid positions in the putative antigen recognition site (ARS). As a consequence of this, in the ARS, certain alleles in several species have counterparts in other species that are more similar to each other than to any other intra-species allele. In contrast, the frequencies of silent nucleotide substitutions were found to be much higher in comparison between species than within species, implying that none of these alleles originated before the divergence of these distantly related species. An analysis of codon usage, as well as the use of silent nucleotides in individual codons, suggests that many of these similarities are due to convergent evolution.