Evolution of MHC class I genes in the European badger (Meles meles)

The major histocompatibility complex (MHC) plays a central role in the adaptive immune system and provides a good model with which to understand the evolutionary processes underlying functional genes. Trans-species polymorphism and orthology are both commonly found in MHC genes; however, mammalian MHC class I genes tend to cluster by species. Concerted evolution has the potential to homogenize different loci, whereas birth-and-death evolution can lead to the loss of orthologs; both processes result in monophyletic groups within species. Studies investigating the evolution of MHC class I genes have been biased toward a few particular taxa and model species. We present the first study of MHC class I genes in a species from the superfamily Musteloidea. The European badger (Meles meles) exhibits moderate variation in MHC class I sequences when compared to other carnivores. We identified seven putatively functional sequences and nine pseudogenes from genomic (gDNA) and complementary (cDNA) DNA, signifying at least two functional class I loci. We found evidence for separate evolutionary histories of the α1 and α2/α3 domains. In the α1 domain, several sequences from different species were more closely related to each other than to sequences from the same species, resembling orthology or trans-species polymorphism. Balancing selection and probable recombination maintain genetic diversity in the α1 domain, evidenced by the detection of positive selection and a recombination event. By comparison, two recombination breakpoints indicate that the α2/α3 domains have most likely undergone concerted evolution, where recombination has homogenized the α2/α3 domains between genes, leading to species-specific clusters of sequences. Our findings highlight the importance of analyzing MHC domains separately.     

MHC class I expression dependent on bacterial infection and parental factors in whitefish embryos (Salmonidae)

Ecological conditions can influence not only the expression of a phenotype, but also the heritability of a trait. As such, heritable variation for a trait needs to be studied across environments. We have investigated how pathogen challenge affects the expression of MHC genes in embryos of the lake whitefish Coregonus palaea. In order to experimentally separate paternal (i.e. genetic) from maternal and environmental effects, and determine whether and how stress affects the heritable variation for MHC expression, embryos were produced in full‐factorial in vitro fertilizations, reared singly, and exposed at 208 degree days (late‐eyed stage) to either one of two strains of Pseudomonas fluorescens that differ in their virulence characteristics (one increased mortality, while both delayed hatching time). Gene expression was assessed 48 h postinoculation, and virulence effects of the bacterial infection were monitored until hatching. We found no evidence of MHC class II expression at this stage of development. MHC class I expression was markedly down‐regulated in reaction to both pseudomonads. While MHC expression could not be linked to embryo survival, the less the gene was expressed, the earlier the embryos hatched within each treatment group, possibly due to trade‐offs between immune function and developmental rate or further factors that affect both hatching timing and MHC expression. We found significant additive genetic variance for MHC class I expression in some treatments. That is, changes in pathogen pressures could induce rapid evolution in MHC class I expression. However, we found no additive genetic variance in reaction norms in our study population.     

Unusually limited nucleotide sequence variation of the expressed major histocompatibility complex class I genes of a New World primate species (Saguinus oedipus)

Although major histocompatibility complex (MHC) class I molecules are, as a rule, highly polymorphic in mammalian species, those of the New World primate Saguinus oedipus (cotton-top tamarin) exhibit limited polymorphism. We have cloned and sequenced twelve MHC class I cDNAs from this species. Since cloned cotton-top tamarin cell lines express three to six MHC class I molecules, this species must have at least three functional MHC class I loci. There was, however, no evidence of locus-specific substitutions in the tamarin cDNAs. Unlike all other species studied, tamarin MHC class I cDNAs displayed limited nucleotide sequence variation. The sequence similarity between the two most divergent tamarin cDNAs was 95%. To ensure that the polymerase chain reaction (PCR) primers employed in these studies had amplified all of the tamarins' expressed MHC class I genes, we used another set of primers to amplify only exons 2 and 3 from RNA and DNA. PCR of genomic DNA resulted in the amplification of six distinct clones, of which only three were well expressed. Two of these nonexpressed genes were pseudogenes and the other was a nonclassical gene. Southern blot analysis demonstrated that the tamarin has 8–11 MHC class I genes, suggesting we had indeed cloned the majority of these genes. Cotton-top tamarins are, therefore, unique among mammalian species studied to date in that they express MHC class I molecules with limited nucleotide sequence variation.The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers M38403-15.     

Comparative genomic analysis of the major histocompatibility complex class I region in the teleost genus <Emphasis Type="Italic">Oryzias</Emphasis>

The major histocompatibility complex (MHC) class I region of teleosts harbors a tight cluster of the class IA genes and several other genes directly involved in class I antigen presentation. Moreover, the dichotomous haplotypic lineages (termed d- and N- lineages) of the proteasome subunit beta genes, PSMB8 and PSMB10, are present in this region of the medaka, Oryzias latipes. To understand the evolution of the Oryzias MHC class I region at the nucleotide sequence level, we analyzed bacterial artificial chromosome clones covering the MHC class I region containing the d- lineage of Oryzias luzonensis and the d- and N- lineages of Oryzias dancena. Comparison among these three elucidated sequences and the published sequences of the d- and N- lineages of O. latipes indicated that the order and orientation of the encoded genes were completely conserved among these five genomic regions, except for the class IA genes, which showed species-specific variation in copy number. The PSMB8 and PSMB10 genes showed trans-species dimorphism. The remaining regions flanking the PSMB10, PSMB8, and class IA genes showed high degrees of sequence conservation at interspecies as well as intraspecies levels. Thus, the three independent evolutionary patterns under apparently distinctive selective pressures are recognized in the Oryzias MHC class I region. Electronic Supplementary Material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

MHC class I genes in a New World primate, the cotton-top tamarin (Saguinus oedipus), have evolved by an active process of loci turnover

 Lymphocytes of a New World primate, the cotton-top tamarin (Saguinus oedipus), express classical G–related major histocompatibility complex (MHC) class I molecules with unusually limited polymorphism and variability. Three G-related loci, an F locus, an E locus, and two pseudogenes (So-N1 and So-N3) have been identified by cDNA library screening and extensive PCR analysis of both cDNA and genomic DNA from the cotton-top tamarin. Furthermore, each genus of the subfamily Callitrichinae (tamarins and marmosets) appears to express its own unique set of MHC class I genes, likely due to a rapid turnover of loci. The rapid emergence of unique MHC class I genes in the Callitrichinae genera, resulting from an active process of duplication and inactivation of loci, may account for the limited diversity of the MHC class I genes in the cotton-top tamarin. To determine the nature of the entire complement of MHC class I genes in the cotton-top tamarin, we synthesized a genomic DNA library and screened it with MHC class I-specific probes. We isolated nine new MHC class I pseudogenes from this library. These newly isolated tamarin G–related MHC class I pseudogenes are not closely related to any of their functional counterparts in the tamarin, suggesting that they do not share a recent common ancestral gene with the tamarin's currently expressed MHC class I loci. In addition, these tamarin sequences display a high rate of nonsynonymous substitutions in their putative peptide binding region. This indicates that the genes from which they have derived were likely subject to positive selection and, therefore, were once functional. Our data support the notion that an extremely high rate of loci turnover is largely responsible for the limited diversity of the MHC class I genes in the cotton-top tamarin. Received: 15 September 1997 / Revised: 2 July 1998     

Modo-UG, a marsupial nonclassical MHC class I locus

Modo-UG is a class I gene located in the MHC of the marsupial Monodelphis domestica, the gray, short-tailed opossum. Modo-UG is expressed as three alternatively spliced mRNA forms, all of which encode a transmembrane form with a short cytoplasmic tail that lacks phosphorylation sites typically found in classical class I molecules. The three alternative mRNAs would encode a full-length form, an isoform lacking the α2 domain, and one lacking both α2 and α3 domains. Genotyping both captive-bred and wild M. domestica from different geographic regions revealed no variation in the residues that make up Modo-UG’s peptide-binding groove. Modo-UG’s low polymorphism is contrasting to that of a nearby class I locus, Modo-UA1, which has a highly polymorphic peptide-binding region. Absence of functional polymorphism in Modo-UG is therefore not a general feature of opossum class I genes but the result of negative selection. Modo-UG is the first MHC linked marsupial class I to be described that appears to clearly have nonclassical features.Electronic Supplementary Material Supplementary material is available for this article at     

A prominent role for invariant T cells in the amphibian Xenopus laevis tadpoles

Invariant T (iT) cells expressing an invariant or semi-invariant T cell receptor (TCR) repertoire have gained attention in recent years because of their potential as specialized regulators of immune function. These iT cells are typically restricted by nonclassical MHC class I molecules (e.g., CD1d and MR1) and undergo differentiation pathways distinct from conventional T cells. While the benefit of a limited TCR repertoire may appear counterintuitive in regard to the advantage of the diversified repertoire of conventional T cells allowing for exquisite specificity to antigens, the full biological importance and evolutionary conservation of iT cells are just starting to emerge. It is generally considered that iT cells are specialized to recognize conserved antigens equivalent to pathogen-associated molecular pattern. Until recently, little was known about the evolution of iT cells. The identification of class Ib and class I-like genes in nonmammalian vertebrates, despite the heterogeneity and variable numbers of these genes among species, suggests that iT cells are also present in ectothermic vertebrates. Indeed, recent studies in the amphibian Xenopus have revealed a drastic overrepresentation of several invariant TCRs in tadpoles and identified a prominent nonclassical MHC class I-restricted iT cell subset critical for tadpole antiviral immunity. This suggests an important and perhaps even dominant role of multiple nonclassical MHC class I-restricted iT cell populations in tadpoles and, by extension, other aquatic vertebrates with rapid external development that are under pressure to produce a functional lymphocyte repertoire with small numbers of cells.     

Spatially variable coevolution between a haemosporidian parasite and the MHC of a widely distributed passerine

The environment shapes host–parasite interactions, but how environmental variation affects the diversity and composition of parasite‐defense genes of hosts is unresolved. In vertebrates, the highly variable major histocompatibility complex (MHC) gene family plays an essential role in the adaptive immune system by recognizing pathogen infection and initiating the cellular immune response. Investigating MHC‐parasite associations across heterogeneous landscapes may elucidate the role of spatially fluctuating selection in the maintenance of high levels of genetic variation at the MHC. We studied patterns of association between an avian haemosporidian blood parasite and the MHC of rufous‐collared sparrows (Zonotrichia capensis) that inhabit environments with widely varying haemosporidian infection prevalence in the Peruvian Andes. MHC diversity peaked in populations with high infection prevalence, although intra‐individual MHC diversity was not associated with infection status. MHC nucleotide and protein sequences associated with infection absence tended to be rare, consistent with negative frequency‐dependent selection. We found an MHC variant associated with a ~26% decrease in infection probability at middle elevations (1501–3100 m) where prevalence was highest. Several other variants were associated with a significant increase in infection probability in low haemosporidian prevalence environments, which can be interpreted as susceptibility or quantitative resistance. Our study highlights important challenges in understanding MHC evolution in natural systems, but may point to a role of negative frequency‐dependent selection and fluctuating spatial selection in the evolution of Z. capensis MHC.     

Genetic diversity of MHC class II <Emphasis Type="Italic">DRB</Emphasis> alleles in the continental and Japanese populations of the sable <Emphasis Type="Italic">Martes zibellina</Emphasis> (Mustelidae,Carnivora, Mammalia)

The sable (Martes zibellina) is a medium-sized mustelid inhabiting forest environments in Siberia, northern China, the Korean Peninsula, and Hokkaido Island, Japan. To further understand the molecular evolution of the major histocompatibility complex (MHC), we sequenced part of exon 2 in MHC class II DRB genes, including codons encoding the antigen binding site, from 33 individuals from continental Eurasia and Japan. We identified 16 MHC class II DRB alleles (Mazi-DRBs), some of which were geographically restricted and others broadly distributed, and eight putative pseudogenes. A single-breakpoint recombination analysis detected a recombination site in the middle of exon 2. A mixed effects model of evolution analysis identified five amino acid sites presumably under positive selection. These sites were all located in the region 3′ to the recombination site, suggesting that positive selection and recombination could be committed to the diversity of the M. zibellina DRB gene. In a Bayesian phylogenetic tree, all Mazi-DRBs and the presumed pseudogenes grouped within a Mustelidae clade. The Mazi-DRBs showed trans-species polymorphism, with some alleles most closely related to alleles from other mustelid species. This result suggests that the sable DRBs have evolved under long-lasting balancing selection.     

Marsupials and monotremes possess a novel family of MHC class I genes that is lost from the eutherian lineage

Background

Major histocompatibility complex (MHC) class I genes are found in the genomes of all jawed vertebrates. The evolution of this gene family is closely tied to the evolution of the vertebrate genome. Family members are frequently found in four paralogous regions, which were formed in two rounds of genome duplication in the early vertebrates, but in some species class Is have been subject to additional duplication or translocation, creating additional clusters. The gene family is traditionally grouped into two subtypes: classical MHC class I genes that are usually MHC-linked, highly polymorphic, expressed in a broad range of tissues and present endogenously-derived peptides to cytotoxic T-cells; and non-classical MHC class I genes generally have lower polymorphism, may have tissue-specific expression and have evolved to perform immune-related or non-immune functions. As immune genes can evolve rapidly and are subject to different selection pressure, we hypothesised that there may be divergent, as yet unannotated or uncharacterised class I genes.

Results

Application of a novel method of sensitive genome searching of available vertebrate genome sequences revealed a new, extensive sub-family of divergent MHC class I genes, denoted as UT, which has not previously been characterized. These class I genes are found in both American and Australian marsupials, and in monotremes, at an evolutionary chromosomal breakpoint, but are not present in non-mammalian genomes and have been lost from the eutherian lineage. We show that UT family members are expressed in the thymus of the gray short-tailed opossum and in other immune tissues of several Australian marsupials. Structural homology modelling shows that the proteins encoded by this family are predicted to have an open, though short, antigen-binding groove.

Conclusions

We have identified a novel sub-family of putatively non-classical MHC class I genes that are specific to marsupials and monotremes. This family was present in the ancestral mammal and is found in extant marsupials and monotremes, but has been lost from the eutherian lineage. The function of this family is as yet unknown, however, their predicted structure may be consistent with presentation of antigens to T-cells.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1745-4) contains supplementary material, which is available to authorized users.     

Extraordinary MHC class II B diversity in a non‐passerine,wild bird: the Eurasian Coot Fulica atra (Aves: Rallidae)

The major histocompatibility complex (MHC) hosts the most polymorphic genes ever described in vertebrates. The MHC triggers the adaptive branch of the immune response, and its extraordinary variability is considered an evolutionary consequence of pathogen pressure. The last few years have witnessed the characterization of the MHC multigene family in a large diversity of bird species, unraveling important differences in its polymorphism, complexity, and evolution. Here, we characterize the first MHC class II B sequences isolated from a Rallidae species, the Eurasian Coot Fulica atra. A next‐generation sequencing approach revealed up to 265 alleles that translated into 251 different amino acid sequences (β chain, exon 2) in 902 individuals. Bayesian inference identified up to 19 codons within the presumptive peptide‐binding region showing pervasive evidence of positive, diversifying selection. Our analyses also detected a significant excess of high‐frequency segregating sites (average Tajima's D = 2.36, P < 0.05), indicative of balancing selection. We found one to six different alleles per individual, consistent with the occurrence of at least three MHC class II B gene duplicates. However, the genotypes comprised of three alleles were by far the most abundant in the population investigated (49.4%), followed by those with two (29.6%) and four (17.5%) alleles. We suggest that these proportions are in agreement with the segregation of MHC haplotypes differing in gene copy number. The most widespread segregating haplotypes, according to our findings, would contain one single gene or two genes. The MHC class II of the Eurasian Coot is a valuable system to investigate the evolutionary implications of gene copy variation and extensive variability, the greatest ever found, to the best of our knowledge, in a wild population of a non‐passerine bird.     

High adaptive variability and virus-driven selection on major histocompatibility complex (MHC) genes in invasive wild rabbits in Australia

The rabbit haemorrhagic disease virus (RHDV) was imported into Australia in 1995 as a biocontrol agent to manage one of the most successful and devastating invasive species, the European rabbit (Oryctolagus cuniculus cuniculus). During the first disease outbreaks, RHDV caused mortality rates of up to 97% and reduced Australian rabbit numbers to very low levels. However, recently increased genetic resistance to RHDV and strong population growth has been reported. Major histocompatibility complex (MHC) class I immune genes are important for immune responses against viruses, and a high MHC variability is thought to be crucial in adaptive processes under pathogen-driven selection. We asked whether strong population bottlenecks and presumed genetic drift would have led to low MHC variability in wild Australian rabbits, and if the retained MHC variability was enough to explain the increased resistance against RHD. Despite the past bottlenecks we found a relatively high number of MHC class I sequences distributed over 2–4 loci. We identified positive selection on putative antigen-binding sites of the MHC. We detected evidence for RHDV-driven selection as one MHC supertype was negatively associated with RHD survival, fitting expectations of frequency-dependent selection. Gene duplication and pathogen-driven selection are possible (and likely) mechanisms that maintained the adaptive potential of MHC genes in Australian rabbits. Our findings not only contribute to a better understanding of the evolution of invasive species, they are also important in the light of planned future rabbit biocontrol in Australia.     

Blood parasites shape extreme major histocompatibility complex diversity in a migratory passerine

Pathogens are one of the main forces driving the evolution and maintenance of the highly polymorphic genes of the vertebrate major histocompatibility complex (MHC). Although MHC proteins are crucial in pathogen recognition, it is still poorly understood how pathogen‐mediated selection promotes and maintains MHC diversity, and especially so in host species with highly duplicated MHC genes. Sedge warblers (Acrocephalus schoenobaenus) have highly duplicated MHC genes, and using data from high‐throughput MHC genotyping, we were able to investigate to what extent avian malaria parasites explain temporal MHC class I supertype fluctuations in a long‐term study population. We investigated infection status and infection intensities of two different strains of Haemoproteus, that is avian malaria parasites that are known to have significant fitness consequences in sedge warblers. We found that prevalence of avian malaria in carriers of specific MHC class I supertypes was a significant predictor of their frequency changes between years. This finding suggests that avian malaria infections partly drive the temporal fluctuations of the MHC class I supertypes. Furthermore, we found that individuals with a large number of different supertypes had higher resistance to avian malaria, but there was no evidence for an optimal MHC class I diversity. Thus, the two studied malaria parasite strains appear to select for a high MHC class I supertype diversity. Such selection may explain the maintenance of the extremely high number of MHC class I gene copies in sedge warblers and possibly also in other passerines where avian malaria is a common disease.     

MHC evolution in three salmonid species: a comparison between class II alpha and beta genes

The genes of the major histocompatibility complex (MHC) are amongst the most variable in vertebrates and represent some of the best candidates to study processes of adaptive evolution. However, despite the number of studies available, most of the information on the structure and function of these genes come from studies in mammals and birds in which the MHC class I and II genes are tightly linked and class II alpha exhibits low variability in many cases. Teleost fishes are among the most primitive vertebrates with MHC and represent good organisms for the study of MHC evolution because their class I and class II loci are not physically linked, allowing for independent evolution of both classes of genes. We have compared the diversity and molecular mechanisms of evolution of classical MH class II α and class II β loci in farm populations of three salmonid species: Oncorhynchus kisutch, Oncorhynchus mykiss and Salmo salar. We found single classical class II loci and high polymorphism at both class II α and β genes in the three species. Mechanisms of evolution were common for both class II genes, with recombination and point mutation involved in generating diversity and positive selection acting on the peptide-binding residues. These results suggest that the maintenance of variability at the class IIα gene could be a mechanism to increase diversity in the MHC class II in salmonids in order to compensate for the expression of one single classical locus and to respond to a wider array of parasites.     

Variety matters: adaptive genetic diversity and parasite load in two mouse opossums from the Brazilian Atlantic forest

The adaptive potential of a species to a changing environment and in disease defence is primarily based on genetic variation. Immune genes, such as genes of the major histocompatibility complex (MHC), may thereby be of particular importance. In marsupials, however, there is very little knowledge about natural levels and functional importance of MHC polymorphism, despite their key role in the mammalian evolution. In a previous study, we discovered remarkable differences in the MHC class II diversity between two species of mouse opossums (Gracilinanus microtarsus, Marmosops incanus) from the Brazilian Atlantic forest, which is one of the most endangered hotspots for biodiversity conservation. Since the main forces in generating MHC diversity are assumed to be pathogens, we investigated in this study gastrointestinal parasite burden and functional associations between the individual MHC constitution and parasite load. We tested two contrasting scenarios, which might explain differences in MHC diversity between species. We predicted that a species with low MHC diversity would either be under relaxed selection pressure by low parasite diversity (‘Evolutionary equilibrium’ scenario), or there was a recent loss in MHC diversity leading to a lack of resistance alleles and increased parasite burden (‘Unbalanced situation’ scenario). In both species it became apparent that the MHC class II is functionally important in defence against gastrointestinal helminths, which was shown here for the first time in marsupials. On the population level, parasite diversity did not markedly differ between the two host species. However, we did observe considerable differences in the individual parasite load (parasite prevalence and infection intensity): while M. incanus revealed low MHC DAB diversity and high parasite load, G. microtarsus showed a tenfold higher population wide MHC DAB diversity and lower parasite burden. These results support the second scenario of an unbalanced situation.     

Genetic structure in insular and mainland populations of house sparrows (Passer domesticus) and their hemosporidian parasites

Small and isolated populations usually exhibit low levels of genetic variability, and thus, they are expected to have a lower capacity to adapt to changes in environmental conditions, such as exposure to pathogens and parasites. Comparing the genetic variability of selectively neutral versus functional loci allows one to assess the evolutionary history of populations and their future evolutionary potential. The genes of the major histocompatibility complex (MHC) control immune recognition of parasites, and their unusually high diversity is genes which is likely driven by parasite‐mediated balancing selection. Here, we examined diversity and differentiation of neutral microsatellite loci and functional MHC class I genes in house sparrows (Passer domesticus), living in six insular and six mainland populations, and we aimed to determine whether their diversity or differentiation correlates with the diversity and the prevalence of infection of hemosporidian parasites. We found that island bird populations tended to have lower neutral genetic variability, whereas MHC variability gene was similar between island and mainland populations. Similarly, island populations tended to show greater genetic differentiation than mainland populations, especially at microsatellite markers. The maintenance of MHC genetic diversity and its less marked structure in the island populations could be attributed to balancing‐selection. The greater MHC differentiation among populations was negatively correlated with similarity in blood parasites (prevalence and diversity of parasite strains) between populations. Even at low prevalence and small geographical scale, haemosporidian parasites might contribute to structure the variability of immune genes among populations of hosts.