Exonic polymorphism vs intronic simple repeat hypervariability in MHC-DRB genes

Abstract

Gene products encoded by the major histocompatibility complex often exhibit a high degree of polymorphism. In humans the HLA-DR polymorphism is due to more than 50 alleles with varying exon 2 sequences. Each group of DRB alleles contains a certain form of the basic simple repeat motif (gt)n(ga)m in intron 2. Identical alleles can be differentiated on the basis of the hypervariable repeat. In this study focused on cattle (Bos taurus) we identified different Bota-DRB alleles in a limited survey by amplification via polymerase chain reaction and sequencing. In addition DRB exon 2 sequences were also obtained from eight additional hoofed animal species (seven horned artiodactyls and one pig) revealing artiodactyl-specific polymorphic and nonpolymorphic substitutions. In the genus Bos the intronic simple repeat variability was compared with exonic DRB polymorphism. As in humans all Bota-DRB exons were always associated with specifically organized basic simple repeat structures. Yet the extent of simple repeat variability was lower in cattle compared to humans. Selective breeding in the process of domestication might be responsible for the diminished intronic hypervariability. Nevertheless, the hypermutable simple repeat sequences have been preserved in the same position and with the same principal structure for at least 70 x 10(6) years of evolution. Unexpectedly, the rate of intronic simple repeat and exonic changes appear quite similar.

Association of molecular polymorphism in exon 2 of Ovar-DRB1 gene with weight traits in Iranian Makuie sheep breed

“…Quality and quantity of extracted DNA was measured on 0.8% agarose gel prepared in 0.5X TBE buffer (45 mM Tris base, 45 mM boric acid, 1 mM EDTA, pH 8.0), visualized with ethidium bromide (1.0 mg mL -1 ), and photographed under UV light using a Gel-Doc image analysis system (Gel Logic 212 PRO, USA). The amplification of the exon 2 region of the MHC-DRB1 gene was achieved using primers MHC-Forward (5´-TCTCTGCAGCACATTTCCTGG-3´) and MHCReverse (5´-CTCGCCGCTGCACAGTGAAAC-3´) targeting a fragment of 279 bp as described by Ammer et al (20). The PCRs were performed in a final volume of 25 µL containing 100 ng of template DNA, 0.5 µL of each primer, 2.5 µL of 10X PCR buffer, 4 µL of 1.25 mM dNTP (BioFluxbiotech, http://biofluxbiotech.com), 1 µL of 50 mM MgCl 2 (CinnaGen, Tehran, Iran), and 0.5 µL of Tag DNA polymerase (CinnaGen) using a 96-well Eppendorf Mastercycler Gradient (Type 5331, Eppendorf AG, Hamburg, Germany).…”

“…The exon 2 regions of the Ovar-DRB1 gene (a fragment of 279 bp in length) were successfully amplified in our first attempt by the specific primers as described by Ammer et al (20). All extracted DNAs from sheep blood samples yielded a specific single-band PCR product without any nonspecific bands.…”

“…High polymorphism in the exon 2 sequence associates with immune response toward a wide variety of pathogens. Two MHC microsatellite alleles, GT/GA, were identified on the second intron of the Ovar-DRB1 gene, closely linked to the exon 2 sequence and associated with production of the highest levels of antibodies in response to nematode parasites (20). These alleles are also expected to play an important role in the fertility of sheep (21).…”

OLA-DRB1 microsatellite variants are associated with ovine growth and reproduction traits

“…This repeat is supposed to have been conserved between humans and artiodactyls for more than 70 million years [2]. In vitro experiments have revealed protein binding to (GT) n (GA) m sequences, which points towards a biological function of this microsatellite in gene regulation [17].…”

Selection on MHC-linked microsatellite loci in sheep populations

“…Microsatellite loci located within the MHC region have received increasing attention as proxies for inferring the level of polymorphism at MHC loci (Ammer et al, 1992;Ellegren et al, 1993;Schwaiger et al, 1993Schwaiger et al, , 1994. The analysis of variation at these loci has produced contrasting results.…”