The DY sub-region of the sheep MHC contains an A/B gene pair
The major histocompatibility complex (MHC) class II region of ruminants appears to have a structure broadly similar to that of the human class II or HLA-D region. Restriction fragment length polymorphism (RFLP) studies of class II genes in cattle (Andersson et al. 1988; Anderson and Rask 1988; Sigurdardottir et al. 1988, 1991 b), and in sheep (Scott et al. 1987), have provided an estimate of the number and type of class II genes in these species. The subsequent cloning and sequencing of sheep and cattle class II genes (Muggli-Cockett and Stone 1989; Groenen et al. 1990; van der Poel et al. 1990; Andersson et al. 1991; Scott et al. 1991 a, b; Ballingall et al. 1992; Sigurdardottir et al. 1991 a, 1992), have demonstrated that they are highly homologous to their human counterparts. Of more interest, therefore, are loci within the ruminant MHC which differ from the HLA class II region.
Three distinguishing features of the ruminant class II region described to date are, firstly, the apparent absence of a DP-like isotype, secondly, the variability in the number of DQ genes between haplotypes (Andersson and Rask 1988), and thirdly, the presence of class II genes presumed to be unique to the ruminant (Andersson et al. 1988). The presence of two such genes, designated DYA and DYB, was deduced from RFLP studies of cattle DNA. These genes were shown to segregate together with the DOB gene in one region separated by a recombination distance of 17 cM from the region which contains the DQA, DQB, DRB, DRA, and C4 loci (Andersson et al. 1988). Subsequently, Bota-DYA was cloned from a phage library and sequenced (van der Poel et al. 1990; Acc. Nos. m30119 and m30118). The sequence of part of a similar gene in the goat, obtained by PCR by using primers derived from the cattle sequence, has recently been reported (Mann et al. 1993; Acc. No. m94325). However, there has been no report of the cloning of a B gene partner for the DYA gene. A novel cattle class II B gene designated Bota-DIB was cloned from a phage library and sequenced by Stone and Muggli-Cockett (1990). This was shown to be a single copy gene of limited polymorphism, which on the basis of RFLP analysis was probably not Bota-DYB but did appear to be distinct from other known cattle class II genes. The species distribution of this B gene was shown to be restricted to Cervidae, Giraffidae, and Bovidae (Stone and Muggli-Cockett 1993). However, it is not known whether any of these novel genes are functional.
Expressed human class II genes usually occur as A/B gene pairs situated close to each other on the chromosome. This is also the case with Bota-DQ genes (Groenen et al. 1990) and Ovar-DQ genes (Deverson et al. 1991; Wright and Ballingall 1994). We used the techniques of cosmid cloning and DNA-mediated gene transfection to determine whether there is a sheep equivalent of the Bota-DYA gene, whether there is a DYB gene partner, and whether there is a protein product.
A cosmid library was constructed from DNA prepared from a Finnish Landrace ram. The library was screened with Ovar-DQA, Ovar-DQB, HLA-DQA, and HLA-DQB gene probes at low stringency. A cosmid clone, 365, was obtained which hybridized weakly to both the Ovar gene probes. Restriction maps of the clone were produced for the enzymes Eco R1, Bam HI, Hin dIII, Sac I and Sma I. When the maps were compared to those published for the phage clones containing the Bota-DYA (van der Poel et al. 1990) and the Bota-DIB gene (Stone and Muggli-Cockett 1990), there was an imperfect match (Figure 1 shows the Eco RI maps). However, the sequence data for the A and B genes in cosmid 365 are more convincing. The sequences of exons 2 and 3 of the A gene in cosmid 365 and the Bota-DYA gene, together with the partial sequence from the third exon of the Cahi-DYA gene are shown in Figure 2 A. The predicted amino acid translations of these genes together with those of other published sheep MHC class II A genes are shown in Figure 2 B. The A gene in cosmid 365 had all the salient features of an MHC class II A gene. It showed a high sequence similarity to the cattle and caprine DYA genes and much less so to the Ovar-DRA gene (Ballingall et al. 1992; Acc. No z11600) and the Ovar-DQA1 and DQA2 (Scott et al. 1991 a; Acc. Nos. m33304 and m33305), as detailed in Table 1. The cosmid A gene showed low sequence similarity to the sheep DNA (formerly DZA) gene (unpublished observations). The A gene described here is clearly the sheep homologue of the Bota-DYA gene.
The sequences of the second, third, and fourth exons of the B gene in cosmid 365 are shown in Figure 3 A together with those of the Bota-DIB gene (Stone and Muggli-Cockett 1990). Unfortunately, the presence of a Bam HI site in exon 2 of the sheep gene caused a truncation at this point, during the cloning procedure and so a part of exon 2, the whole of exon 1, and all the upstream regulatory elements were missing. The predicted amino acid translations of exons 2, 3, and 4 are shown together with those of an Ovar-DQB (Scott et al. 1991 a; Acc. No. m33323) and an expressed Ovar-DRB gene (Ballingall et al. 1992; Acc. No. z11522) in Figure 3 B.
Unable to display preview. Download preview PDF.
- Andersson, L., Lunden, A., Sigurdardottir, S., Davies, C. J., and Rask, L. Linkage relationships in the bovine MHC region: High recombination frequency between class II subregions. Immunogenetics 27: 273–280, 1988Google Scholar
- Andersson, L. and Rask, L. Characterisation of the MHC class II region in cattle. The number of DQ genes varies between haplotypes. Immunogenetics 27: 110–120, 1988Google Scholar
- Andersson, L., Sigurdardottir, S., Borsch, C., and Gustafsson, K. Evolution of MHC polymorphism: extensive sharing of polymorphic sequence motifs between human and bovine DRB alleles. Immunogenetics 33: 188–193, 1991Google Scholar
- Ballingall, K. T., Wright, H., Redmond, J., Dutia, B. M., Hopkins, J., Lang, J., Deverson, E. V., Howard, J. C., Puri, N., and Haig, D. M. Expression and characterisation of ovine major histocompatibility complex class II (OLA-DR) genes. Anim Genet 23: 347–359, 1992Google Scholar
- Campbell, R. D. and Trowsdale, J. A map of the human MHC. Immunol Today 14: 347–352, 1993Google Scholar
- Deverson, E. V., Wright, H., Watson, S., Ballingall, K. T., Huskisson, N., Diamond, A. G., and Howard, J. C. Class II major histocompatibility complex genes of the sheep. Anim Genet 22: 211–225, 1991Google Scholar
- Groenen, M. A. M., Van der Poel, J. J., Dijkhof, R. J. M., and Giphart, M. J. The nucleotide sequence of the bovine MHC class II DQB and DRB genes. Immunogenetics 31: 37–44, 1990Google Scholar
- Mann, A. J., Abraham, L. J., Cameron, P. U., Robinson, W., Giphart, M. J., and Dawkins, R. L. The caprine MHC contains DYA genes. Immunogenetics 37: 292–295, 1993Google Scholar
- Muggli-Cockett, N. E. and Stone, R. T. Partial nucleotide sequence of a bovine major histocompatibility class II DRB-like gene. Anim Genet 20: 361–369, 1989Google Scholar
- Scott, P. C., Choi, C.-L., and Brandon, M. R. Genetic organisation of the ovine MHC class II region. Immunogenetics 25: 116–122, 1987Google Scholar
- Scott, P. C., Gogolin-Ewens, K. J., Adams, T. E., and Brandon, M. R. Nucleotide sequence, polymorphism and evolution of ovine MHC class II DQA genes. Immunogenetics 34: 69–79, 1991 aGoogle Scholar
- Scott, P. C., Maddox, J. F., Gogolin-Ewens, K. J., and Brandon, M. R. The nucleotide sequence and evolution of ovine MHC class II B genes: DQB and DRB. Immunogenetics 34: 80–87, 1991 bGoogle Scholar
- Sigurdardottir, S., Lunden, A., and Andersson, L. Restriction fragment length polymorphism of DQ and DR class II genes of the bovine major histocompatibility complex. Anim Genet 19: 133–150, 1988Google Scholar
- Sigurdardottir, S., Borsch, C., Gustafsson, K., and Andersson, L. Cloning and sequencing of 14 DRB3 alleles of the bovine MHC by using the polymerase chain reaction. Anim Genet 22: 199–210, 1991 aGoogle Scholar
- Sigurdardottir, S., Mariani, P., Groenen, M. A. M., Van der Poel, J. J., and Andersson, L. Organisation and polymorphism of bovine major histocompatibility complex class II genes as revealed by genomic hybridisations with bovine probes. Anim Genet 22: 465–476, 1991 bGoogle Scholar
- Sigurdardottir, S., Borsch, C., Gustafsson, K., and Andersson, L. Gene duplications and sequence polymorphism of bovine class II DQB genes. Immunogenetics 35: 205–213, 1992Google Scholar
- Stone, R. T. and Muggli-Cockett, N. E. Partial nucleotide sequence of a novel bovine major histocompatibility complex class II beta chain gene, BoLA DIB. Anim Genet 21: 352–360, 1990Google Scholar
- Stone, R. T. and Muggli-Cockett, N. E. BoLA-DIB: species distribution, linkage with DOB and Northern analysis. Anim Genet 24: 41–45, 1993Google Scholar
- Van der Poel, J. J., Groenen, M. A. M., Dijkhof, R. J. M., Ruyter, D., and Giphart, M. J. The nucleotide sequence of the bovine MHC class II alpha genes: DRA, DQA and DYA. Immunogenetics 31: 29–36, 1990Google Scholar
- Van Eijk, M. J. T., Russ, I., and Lewin, H. A. Order of bovine DRB3, DYA and PRL determined by sperm typing. Mammalian Genome 4: 113–118, 1993Google Scholar
- Wright, H. and Ballingall, K. T. Mapping and characterisation of the DQ subregion of the ovine MHC. Anim Genet, in pressGoogle Scholar