High Major Histocompatibility Complex Research Articles

Different MHC class I cell surface expression levels in diverse chicken lines, associations with B blood group, and proposed relationship to antigen-binding repertoire

The Major Histocompatibility Complex (MHC) is a cluster of genes with primarily immune-related functions. The MHC class I genes are responsible for self- versus non-self-recognition and viral antigen presentation to T lymphocytes. The chicken MHC class I protein binds its cognate antigen(s) over a repertoire spectrum ranging from promiscuous (generalist) to fastidious (specialist). The MHC class I protein expression level at the cell surface is inversely related to the promiscuity of its peptide-binding repertoire. In our study, erythrocytes from 6 diverse and highly inbred lines of chickens, a closed broiler line, and a highly advanced intercross line were evaluated for MHC class I antigen expression level by flow cytometry using monoclonal antibodies to chicken MHC class I molecules. In chickens, the B blood group antigens include the MHC class I antigen expressed from the MHC. Thus, the B blood group has historically been used as a genetic marker for Marek's Disease virus response. Erythrocytes of the inbred lines were blood typed by serology. The B21 blood type is widely recognized as relatively resistant to Marek's disease and regarded as an MHC class I generalist with low MHC class I expression. The Spanish line, which types serologically as B21.1 (similar to B21), was the lowest MHC class I expressing line. The two sublines (B5.1 and B15.2) of the Fayoumi breed, which significantly differed in their MHC class I expression, also differ in response to multiple pathogens. These defined genetic lines of chickens, with distinct MHC class I expression levels, provide an excellent platform to further interrogate the hypothesis of high or low MHC class I expression (antigenic specialists or generalists, respectively) determining diverse responses to pathogens.

Partial molecular characterization, expression pattern and polymorphism analysis of MHC I genes in Chinese domestic goose (Anser cygnoides).

Major histocompatibility complex (MHC) allelic polymorphism is critically important for mediating antigen presentation in vertebrates. Presently, there are insufficient studies of MHC genetic diversity in domestic Anseriform birds. In this study, we analyzed the expression profile of MHC I genes and screened for MHC I exon 2 polymorphism in one domestic goose population from China using Illumina MiSeq sequencing. The results showed that four MHC I alleles (Ancy-IE2*09/*11/*13/*21) in one goose were identified based on cDNA cloning and sequencing using four primer combinations, and the varying number of cDNA clones implied that these four classical sequences showed differential expression patterns. Through next-generation sequencing, 27 alleles were obtained from 68 geese with 3-10 putative alleles per individual, indicating at least the existence of 5 MHC I loci in the goose. The marked excess of the non-synonymous over the synonymous substitution in the peptide-binding region (PBR) along 27 alleles and five positively selected sites (PSSs) detected around the PBR indicated that balancing selection might be the major force in shaping high MHC variation in the goose. Additionally, IA alleles displaying lower polymorphism were subject to less positive selection pressure than non-IA alleles with a higher level of polymorphism.

Immune genotypes, immune responses, and survival in a wild bird population.

Individuals vary in their immune genotype, inbreeding coefficient f, immune responses, survival to adulthood, and adult longevity. However, whether immune genes predict survival or longevity, whether such relationships are mediated through immune responses, and how f affects immune genotype remain unclear. We use a wild song sparrow (Melospiza melodia) population in which survival to adulthood, adult longevity, and f were measured precisely, and in which immune responses have previously been assessed. We investigate four toll-like receptor (TLR) and the major histocompatibility complex (MHC) class IIB exon 2 genes. We test: a) whether immune genes predict fitness (survival to adulthood or adult longevity); b) whether immune genes predict immune response; c) whether immune response predicts fitness; and d) whether fitness, immune responses, or immune genotypes are correlated with f. We find that survival to adulthood is not associated with immune gene variation, but adult longevity is decreased by high MHC allele diversity (especially in birds that were relatively outbred), and by the presence of a specific MHC supertype. Immune responses were affected by specific immune genotypes. Survival to adulthood and adult longevity were not predicted by immune response, implying caution in the use of immune response as a predictor for fitness. We also found no relationship between f and immune genotype. This finding indicates that immune gene associations with longevity and immune response are not artefacts of f, and suggests that pathogen-mediated selection at functional loci can slow the loss of genetic variation arising from genetic drift and small population size.

The structure of songbird MHC class I reveals antigen binding that is flexible at the N-terminus and static at the C-terminus.

Long-distance migratory animals such as birds and bats have evolved to withstand selection imposed by pathogens across the globe, and pathogen richness is known to be particularly high in tropical regions. Immune genes, so-called Major Histocompatibility Complex (MHC) genes, are highly duplicated in songbirds compared to other vertebrates, and this high MHC diversity has been hypothesised to result in a unique adaptive immunity. To understand the rationale behind the evolution of the high MHC genetic diversity in songbirds, we determined the structural properties of an MHC class I protein, Acar3, from a long-distance migratory songbird, the great reed warbler Acrocephalus arundinaceus (in short: Acar). The structure of Acar3 was studied in complex with pathogen-derived antigens and shows an overall antigen presentation similar to human MHC class I. However, the peptides bound to Acar3 display an unusual conformation: Whereas the N-terminal ends of the peptides display enhanced flexibility, the conformation of their C-terminal halves is rather static. This uncommon peptide-binding mode in Acar3 is facilitated by a central Arg residue within the peptide-binding groove that fixes the backbone of the peptide at its central position, and potentially permits successful interactions between MHC class I and innate immune receptors. Our study highlights the importance of investigating the immune system of wild animals, such as birds and bats, to uncover unique immune mechanisms which may neither exist in humans nor in model organisms.

Interspecific introgression of MHC genes in Triturus newts: Evidence from multiple contact zones.

The major histocompatibility complex (MHC) genes are central to the adaptive immune response in vertebrates. Selection generally maintains high MHC variation because the spectrum of recognized pathogens depends on MHC polymorphism. Novel alleles favoured by selection originate by interallelic recombination or de novo mutations but may also be acquired by introgression from related species. However, the extent and prevalence of MHC introgression remain an open question. In this study, we tested for MHC introgression in six hybrid zones formed by six Triturus newt species. We sequenced and genotyped the polymorphic second exons of the MHC class I and II genes and compared their interspecific similarity at various distances from the centre of the hybrid zone. We found evidence for introgression of both MHC classes in the majority of examined hybrid zones, with support for a more substantial class I introgression. Furthermore, the overall MHC allele sharing outside of hybrid zones was elevated between pairs of Triturus species with abutting ranges, regardless of the phylogenetic distance between them. No effect of past hybrid zone movement on MHC allele sharing was found. Finally, using previously published genome-wide data, we demonstrated that MHC introgression was more extensive than genome-wide introgression, supporting its adaptive potential. Our study thus provides evidence for the prevalence of MHC introgression across multiple Triturus hybrid zones, indicating that MHC introgression between divergent hybridizing species may be widespread and adaptive.

Abstract 3514: Bispecific gamma delta T cell engagers containing butyrophilin 2A1/3A1 heterodimeric fusion protein efficiently activate Vg9Vd2 T cells and promote tumor cell killing

Abstract A primary mechanism of cancer immunotherapy resistance involves downregulation of specific antigens or major histocompatibility complex (MHC) based antigen presentation, which renders tumor cells invisible to αβ T cells, but not γδ T cells. Recently, a two-step model of γδ T cell activation has emerged, wherein one butyrophilin (BTN, i.e., BTN2A1) directly binds the γδ TCR but is only activated if certain molecular patterns (e.g., phosphoantigens) facilitate recruitment of a second BTN (i.e., BTN3A1) into a complex to form a BTN2A1/3A1 heterodimer. The BTN2A1/3A1 complex specifically activates the Vγ9Vδ2 T cells found in the peripheral blood in response to phosphoantigen build up in tumor cells. The unique mechanism of action and specificity of γδ TCR/BTN interactions suggests that therapeutic proteins comprising specific BTN heterodimers could be used to target specific γδ T cell populations to tumor cells which lack expression of high affinity neoantigens or MHC class I molecules. In this study, we generated bispecific γδ T cell engagers (GADLEN) containing heterodimeric BTN2A1 and BTN3A1 extracellular domains (ECD) fused via inert Fc linkers to scFv domains targeting a tumor-antigen (CD19 or CD33) to test their ability to modulate Vγ9Vδ2 T cells and to promote tumor cell killing in co-culture assays. GADLEN induced proliferation, degranulation and cytokine production including IFNγ and TNFα in Vγ9Vδ2 T cells but requires co-stimulation of a natural cytotoxicity receptor (via anti-NKG2D) or T-cell co-stimulatory receptor (via anti-CD28), highlighting the parallels of signal 1 and signal 2 requirements of TCR activation in αβ- and γδ- T cells. In Vγ9Vδ2 T cells and tumor co-culture assays, the addition of GADLEN alone enhanced killing of CD19+ lymphoma cells such as Daudi and Raji, expressing known ligands for CD28 (e.g., CD80 and CD86). Similarly, GADLEN enhanced Vγ9Vδ2 T cell-mediated killing of K562 cells engineered to express CD19, which express MICA/B (ligands for NKG2D) on the cell surface. In addition, the CD33-targeted GADLEN alone was able to specifically kill acute myeloid leukemia cells expressing CD33 and costimulatory ligands (e.g., CD80 and CD86) in a mechanism similar to the CD19-targeted GADLEN construct. These results highlight the ability of GADLENs in promoting targeted killing of tumor cells by providing the “active” heterodimeric BTN2A1 and BTN3A1 that is critical to enhance cytotoxic killing by Vγ9Vδ2 T cells. Furthermore, this study provides mechanistic insights into BTN-mediated activation of Vγ9Vδ2 T cells and offers proof of concept for using antigen-targeted butyrophilin heterodimeric fusion proteins for the treatment of cancer. Citation Format: Anne Lai, Arpita Patel, Faraha Brewer, Kinsley Evans, Kellsey Johannes, Louis Gonzalez, Kyung Jin Yoo, George Fromm, Keith Wilson, Taylor H. Schreiber, Suresh de Silva. Bispecific gamma delta T cell engagers containing butyrophilin 2A1/3A1 heterodimeric fusion protein efficiently activate Vg9Vd2 T cells and promote tumor cell killing [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3514.

Unravelling the sex-specific diversity and functions of adrenal gland macrophages.

SummaryDespite the ubiquitous function of macrophages across the body, the diversity, origin, and function of adrenal gland macrophages remain largely unknown. We define the heterogeneity of adrenal gland immune cells using single-cell RNA sequencing and use genetic models to explore the developmental mechanisms yielding macrophage diversity. We define populations of monocyte-derived and embryonically seeded adrenal gland macrophages and identify a female-specific subset with low major histocompatibility complex (MHC) class II expression. In adulthood, monocyte recruitment dominates adrenal gland macrophage maintenance in female mice. Adrenal gland macrophage sub-tissular distribution follows a sex-dimorphic pattern, with MHC class IIlow macrophages located at the cortico-medullary junction. Macrophage sex dimorphism depends on the presence of the cortical X-zone. Adrenal gland macrophage depletion results in altered tissue homeostasis, modulated lipid metabolism, and decreased local aldosterone production during stress exposure. Overall, these data reveal the heterogeneity of adrenal gland macrophages and point toward sex-restricted distribution and functions of these cells.

High MHC diversity confers no advantage for phenotypic quality and reproductive performance in a wild bird.

Genes of the major histocompatibility complex (MHC) encode antigen‐binding molecules and are an integral part of the acquired immune response of vertebrates. In general, high individual MHC diversity is expected to increase fitness by broadening the spectrum of pathogens recognized by the immune system, in accordance with the heterozygote advantage mechanism. On the other hand, the optimality hypothesis assumes that individuals with optimal (intermediate), rather than maximum, diversity of the MHC will achieve the highest fitness because of inherent costs associated with expressing diverse MHC alleles.Here, we tested for associations between individual diversity of the MHC class I and class II genes (binding antigens of intra‐ and extracellular pathogens respectively) and a range of fitness‐related traits (condition, ornament expression and reproduction) in an urban population of the Eurasian coot Fulica atra.Contrary to our expectation, we found that high within‐individual allelic diversity of MHC genes (both class I and II) was associated with poorer condition (lower blood haemoglobin concentrations), weaker expression of the putative ornament (smaller frontal shield), later onset of breeding and smaller clutches. An analysis of functional MHC allele clusters (supertypes) provided further support for negative associations of MHC diversity with phenotypic quality and reproductive performance, but most of these relationships could not be explained by the presence of specific maladaptive supertypes. Finally, we found little empirical support for the optimality hypothesis in the Eurasian coot.Our results suggest that the costs of high MHC diversity outweighed any benefits associated with broad MHC repertoire, which could be driven by depauperate pathogen diversity in an urban landscape. To the best of our knowledge, this is one of the first studies providing consistent evidence for negative associations of MHC diversity with a range of fitness‐related traits in a natural avian population.

Using de novo genome assembly and high-throughput sequencing to characterize the MHC region in a non-model bird, the Eurasian coot

Genes of the Major Histocompatibility Complex (MHC) form a key component of vertebrate adaptive immunity, as they code for molecules which bind antigens of intra- and extracellular pathogens (MHC class I and II, respectively) and present them to T cell receptors. In general, MHC genes are hyper-polymorphic and high MHC diversity is often maintained within natural populations (via balancing selection) and within individuals (via gene duplications). Because of its complex architecture with tandems of duplicated genes, characterization of MHC region in non-model vertebrate species still poses a major challenge. Here, we combined de novo genome assembly and high-throughput sequencing to characterize MHC polymorphism in a rallid bird species, the Eurasian coot Fulica atra. An analysis of genome assembly indicated high duplication rate at MHC-I, which was also supported by targeted sequencing of peptide-binding exons (at least five MHC-I loci genotyped). We found high allelic richness at both MHC-I and MHC-II, although signature of diversifying selection and recombination (gene conversion) was much stronger at MHC-II. Our results indicate that Eurasian coot retains extraordinary polymorphism at both MHC classes (when compared to other non-passerine bird species), although they may be subject to different evolutionary mechanism.

Influence of T Cell-Mediated Immune Surveillance on Somatic Mutation Occurrences in Melanoma.

BackgroundNeoantigens are presented on the cancer cell surface by peptide-restricted human leukocyte antigen (HLA) proteins and can subsequently activate cognate T cells. It has been hypothesized that the observed somatic mutations in tumors are shaped by immunosurveillance.MethodsWe investigated all somatic mutations identified in The Cancer Genome Atlas (TCGA) Skin Cutaneous Melanoma (SKCM) samples. By applying a computational algorithm, we calculated the binding affinity of the resulting neo-peptides and their corresponding wild-type peptides with the major histocompatibility complex (MHC) Class I complex. We then examined the relationship between binding affinity alterations and mutation frequency.ResultsOur results show that neoantigens derived from recurrent mutations tend to have lower binding affinities with the MHC Class I complex compared to peptides from non-recurrent mutations. Tumor samples harboring recurrent SKCM mutations exhibited lower immune infiltration levels, indicating a relatively colder immune microenvironment.ConclusionsThese results suggested that the occurrences of somatic mutations in melanoma have been shaped by immunosurveillance. Mutations that lead to neoantigens with high MHC class I binding affinity are more likely to be eliminated and thus are less likely to be present in tumors.

A novel form of immunotherapy using antigen peptides conjugated on PD-L1 antibody

Immune checkpoint inhibitors (ICIs), including programmed cell death protein 1 (PD-1)/programmed death-ligand 1 (PD-L1) and cytotoxic T-lymphocyte-associated protein 4 have shown promising cancer clinical outcomes. However, IC therapy has low patient response rates (10%–15%). Thus, ICIs and sufficient antigen combinations into the tumor microenvironment (TME) is important to produce strong tumor-specific adaptive immune responses. Mice were treated with cisplatin, and human cancer cells were exposed to inflammatory cytokines, to confirm increased PD-L1 and major histocompatibility complex (MHC) I expression by tumor cells or dendritic cells. TC-1, CT26, B16-F1, or B16-F10 tumor cells, and bone marrow-derived dendritic cells, were treated with interferon (IFN)-β, IFN-γ, or tumor necrosis factor-α to identify the molecular mechanisms underlying tumor PD-L1 and MHC I upregulation, and to examine MHC I, CD40, CD80, CD86, or PD-L1 levels, respectively. For synergistic combination therapy, αPD-L1 monoclonal antibody (mAb) covalently linked to the long E7 peptide was generated. Chemotherapy shifted the TME to express high PD-L1 and MHC I, resulting in targeted ICI cargo delivery and enhanced generation and activation of tumor antigen-specific T cells. Synergistic effects of vaccination and IC blockade in the TME were demonstrated using an anti-PD-L1 mAb covalently conjugated to the E7 long peptide.

Improved high-throughput MHC typing for non-model species using long-read sequencing.

The major histocompatibility complex (MHC) plays a critical role in the vertebrate immune system. Accurate MHC typing is critical to understanding not only host fitness and disease susceptibility, but also the mechanisms underlying host‐pathogen co‐evolution. However, due to the high degree of gene duplication and diversification of MHC genes, it is often technically challenging to accurately characterise MHC genetic diversity in non‐model species. Here we conducted a systematic review to identify common issues associated with current widely used MHC typing approaches. Then to overcome these challenges, we developed a long‐read based MHC typing method along with a new analysis pipeline. Our approach enables the sequencing of fully phased MHC alleles spanning all key functional domains and the separation of highly similar alleles as well as the removal of technical artefacts such as PCR heteroduplexes and chimeras. Using this approach, we performed population‐scale MHC typing in the Tasmanian devil (Sarcophilus harrisii), revealing previously undiscovered MHC functional diversity in this endangered species. Our new method provides a better solution for addressing research questions that require high MHC typing accuracy. Since the method is not limited by species or the number of genes analysed, it will be applicable for studying not only the MHC but also other complex gene families.

Updated Overall Survival Data and Predictive Biomarkers of Sintilimab Plus Pemetrexed and Platinum as First-Line Treatment for Locally Advanced or Metastatic Nonsquamous NSCLC in the Phase 3 ORIENT-11 Study

IntroductionSintilimab plus chemotherapy significantly prolonged progression-free survival (PFS) compared with chemotherapy alone in nonsquamous NSCLC in the ORIENT-11 study. Updated overall survival (OS) and PFS data and corresponding biomarker analyses are reported here. MethodsIn this study, a total of 397 patients with previously untreated, locally advanced or metastatic nonsquamous NSCLC were assigned to sintilimab plus chemotherapy combination treatment (combo) group or placebo plus chemotherapy treatment group. The patients were stratified by programmed death-ligand 1 (PD-L1) expression levels. Immune signature profiles from tumor RNA sequencing and PD-L1 immunohistochemistry were correlated with clinical outcome to identify predictive biomarkers. ResultsAs of January 2021, with median follow-up of 22.9 months, median OS was significantly improved in the combo group compared with the placebo plus chemotherapy treatment group (not reached versus 16.8 mo; hazard ratio [HR] = 0.60, 95% confidence interval [CI]: 0.45–0.79, p = 0.0003). High or medium immune cell infiltration was strongly associated with improved PFS in the combo group, in contrast to absent or low immune cell infiltration, which suggests that chemotherapy could not prime “immune deserts” to obtain benefit from programmed cell death protein-1 inhibition. In particular, high major histocompatibility complex (MHC) class II presentation pathway expression was significantly correlated with prolonged PFS (HR = 0.32, 95% CI: 0.19–0.54, p < 0.0001) and OS (HR = 0.36, 95% CI: 0.20–0.64, p = 0.0005) in the combo group. Importantly, patients with low or absent PD-L1 but high MHC class II expression could still benefit from the combo treatment. In contrast, MHC class I antigen presentation pathway was less relevant in this combination setting. ConclusionsThe addition of sintilimab to chemotherapy resulted to significantly longer OS in nonsquamous NSCLC. Expression of MHC class II antigen presentation pathway could identify patients benefiting most from this combination.

Major histocompatibility complex class II genetic diversity and the genetic influence on gut microbiota in Guizhou minipigs.

Major histocompatibility complex (MHC) is an important complex that presents antigen to T cells. The second exon of swine MHC (SLA) class II genes has antigen binding sites that bind with extracellular antigen. Populations with high MHC gene diversity result in low gut microbiota, and individuals with MHC gene heterozygote have lower gut microbiota diversity than that of homozygote. The pig is an animal with organs physiologically and anatomically similar to humans than any other mammal, and the pig is also suitably developed as a laboratory animal to establish the animal models of human disease. However, the relationship between SLA genetic diversity and the gut microbes of the pig is ambiguous. We studied the characterization of SLA class II genes and calculated the genetic diversity, and then we selected experimental animal groups divided by different SLA genotypes to investigate the gut microbiota composition by sequencing V3 to V4 hypervariable regions of bacterial 16s rRNA from fecal samples. Our results showed that Guizhou minipigs had a low SLA genetic diversity, which may be due to the small founder population. The Guizhou minipig population deviated from neutral selection and balancing selection, which shows that Guizhou minipigs experience a strong artificial selection in recent years. We observed that the sex differences influenced gut microbiota much more deeply than that of genetics. Our results also showed that the individual with heterozygote of genes at the SLA class II region had much higher abundant gut microbiota than that of the homozygote.

High Levels of Genetic Variation in MHC-Linked Microsatellite Markers from Native Chicken Breeds.

The major histocompatibility complex (MHC) is a highly polymorphic gene region that regulates cellular communication in all specific immune responses. In this study, we investigated 11 microsatellite (MS) markers in the MHC-B region of chicken populations from four countries: Sri Lanka, Bangladesh, South Korea, and Nigeria. The MS markers were divided into two sets. Set 1 included five novel MS markers, which we assessed using 192 samples from 21 populations. Set 2 included six previously reported markers, which we assessed using 881 samples from 29 populations. The Set 1 MS markers had lower polymorphism (polymorphic information content (PIC) < 0.5) than the Set 2 markers (PIC = 0.4–0.9). In all populations, the LEI0258 marker was the most polymorphic, with a total of 38 alleles (PIC = 0.912, expected heterozygosity (He) = 0.918). Local populations from Sri Lanka, Bangladesh, and Nigeria had higher allele diversity and more haplotypes for Set 2 MS markers than Korean and commercial populations. The Sri Lankan Karuwalagaswewa village population had the highest MHC diversity (mean allele number = 8.17, He = 0.657), whereas the white leghorn population had the lowest (mean allele number = 2.33, He = 0.342). A total of 409 haplotypes (89 shared and 320 unique), with a range of 4 (Rhode Island red) to 46 (Karuwalagaswewa village (TA)), were identified. Among the shared haplotypes, the B21-like haplotype was identified in 15 populations. The genetic relationship observed in a neighbour-joining tree based on the DA distance agreed with the breeding histories and geographic separations. The results indicated high MHC diversity in the local chicken populations. The difference in the allelic pattern among populations presumably reflects the effects of different genotypes, environments, geographic variation, and breeding policies in each country. The selection of MHC allele in domestic poultry can vary due to intensification of poultry production. Preserved MHC diversity in local chicken provides a great opportunity for future studies that address the relationships between MHC polymorphisms and differential immune responses.

Population demographic history and adaptability of the vulnerable Lolokou Sucker Frog.

Amphibians are experiencing worldwide declines due to increasing anthropogenetic disturbances. However, the genetic variability and hence adaptability are still unknown for most frogs. We integrated the mitochondrial (ND2 gene), nuclear (TYR gene) and major histocompatibility complex (MHC) loci, to clarify the demographic patterns and immune-gene diversity of the Lolokou Sucker Frog (Amolops loloensis). Demographic analysis of the ND2 and TYR genes suggested that the Lolokou Sucker Frog experienced a population expansion within the last 10,000years. High MHC diversity was detected, which has likely resulted from positive selection, indicating the current diversity bodes well for the species' adaptive potential to pathogenic challenges. These findings broaden our knowledge on the population history and evolution adaptation of the reclusive torrent frog, and conservation implications are provided.

Many loci make light work: High individual diversity despite low population diversity and random mating at class I MHC in a Critically Endangered island songbird.

Multi-cellular organisms are under constant attack from parasites, making immune defence a critical aspect of fitness. In vertebrate animals, genes of the major histocompatibility complex (MHC) determine the breadth of pathogens to which individuals can respond. Having many MHC alleles can confer better protection against infectious disease, and balancing selection at MHC is widespread. Indeed, MHC loci are famously variable, with some populations harbouring thousands of alleles (Biedrzycka et al., 2018; Robinson, Soormally, Hayhurst, & Marsh, 2016). MHC has also long fascinated behavioural ecologists because mate choice-for example, preferring MHC-dissimilar partners-may amplify the effects of natural selection (Penn & Potts, 1999). But despite keen interest in the evolutionary ecology of MHC, extensive duplication (Minias, Pikus, Whittingham, & Dunn, 2019) has made these genes challenging to study. In a From the Cover article in this issue of Molecular Ecology, Stervander, Dierickx, Thorley, Brooke, and Westerdahl (2020) characterizes class I MHC in a Critically Endangered songbird, relating genotype to mate choice and survivorship. By inferring copy number and patterns of allelic co-segregation, the authors pave the way to elucidating the genomic architecture of MHC in this bottlenecked population. These insights help reconcile apparently counterintuitive findings: no effect of MHC genotype on mate choice or survival, and high MHC diversity within individuals despite low diversity at the population level. The latter finding is cause for optimism regarding conservation prospects. Moreover, these results suggest that ancient duplication events can have longstanding effects on the adaptive landscapes of natural and sexual selection.

High MHC gene copy number maintains diversity despite homozygosity in a Critically Endangered single-island endemic bird, but no evidence of MHC-based mate choice.

Small population sizes can, over time, put species at risk due to the loss of genetic variation and the deleterious effects of inbreeding. Losing diversity in the major histocompatibility complex (MHC) could be particularly harmful, given its key role in the immune system. Here, we assess MHC class I (MHC-I) diversity and its effects on mate choice and survival in the Critically Endangered Raso lark Alauda razae, a species restricted to the 7km2 islet of Raso, Cape Verde, since ~1460, whose population size has dropped as low as 20 pairs. Exhaustively genotyping 122 individuals, we find no effect of MHC-I genotype/diversity on mate choice or survival. However, we demonstrate that MHC-I diversity has been maintained through extreme bottlenecks by retention of a high number of gene copies (at least 14), aided by cosegregation of multiple haplotypes comprising 2-8 linked MHC-I loci. Within-locus homozygosity is high, contributing to low population-wide diversity. Conversely, each individual had comparably many alleles, 6-16 (average 11), and the large and divergent haplotypes occur at high frequency in the population, resulting in high within-individual MHC-I diversity. This functional immune gene diversity will be of critical importance for this highly threatened species' adaptive potential.

Can extreme MHC class I diversity be a feature of a wide geographic range? The example of Seba's short-tailed bat (Carollia perspicillata).

The major histocompatibility complex (MHC) is one of the most diverse genetic regions under pathogen-driven selection because of its central role in antigen binding and immunity. The highest MHC variability, both in terms of the number of individual alleles and gene copies, has so far been found in passerine birds; this is probably attributable to passerine adaptation to both a wide geographic range and a diverse array of habitats. If extraordinary high MHC variation and duplication rates are adaptive features under selection during the evolution of ecologically and taxonomically diverse species, then similarly diverse MHC architectures should be found in bats. Bats are an extremely species-rich mammalian group that is globally widely distributed. Many bat species roost in multitudinous groups and have high contact rates with pathogens, conspecifics, and allospecifics. We have characterized the MHC class I diversity in 116 Panamanian Seba’s short-tailed bats (Carollia perspicillata), a widely distributed, generalist, neotropical species. We have detected a remarkable individual and population-level diversity of MHC class I genes, with between seven and 22 alleles and a unique genotype in each individual. This diversity is comparable with that reported in passerine birds and, in both taxonomic groups, further variability has evolved through length polymorphisms. Our findings support the hypothesis that, for species with a geographically broader range, high MHC class I variability is particularly adaptive. Investigation of the details of the underlying adaptive processes and the role of the high MHC diversity in pathogen resistance are important next steps for a better understanding of the role of bats in viral evolution and as carriers of several deadly zoonotic viruses.Electronic supplementary materialThe online version of this article (10.1007/s00251-019-01128-7) contains supplementary material, which is available to authorized users.

Transcriptome analysis revealed potential mechanisms of differences in physiological stress responses between caged male and female magpies

BackgroundUnder caged conditions, birds are affected more severely by environmental stressors such as dietary structure, activity space, human disturbances, and pathogens, which may be reflected in the gene expression in peripheral blood or other tissues. Elucidating the molecular mechanism of these stress responses will help improve animal welfare.ResultsIn the present study, the blood transcriptomes of six male and five female caged magpies (Pica pica) were sequenced, and a total of ~ 100 Gb in clean reads were generated using the Illumina HiSeq 2000 sequencer. A total of 420,291 unigenes were identified after assembly, of which 179,316 were annotated in five databases, 7471 were assigned to 269 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and 566 were assigned to the Clusters of Orthologous Groups (COG) functional classification “defense mechanisms”. Analysis of differentially expressed genes (DEGs) showed that 2657 unigenes were differentially expressed between males and females (q < 0.1), and these DEGs were assigned to 45 KEGG pathways involving stress resistance, immunity, energy metabolism, reproduction, lifespan regulation, and diseases. Further analysis revealed that females might be more sensitive to stress through upregulation of c-Jun N-terminal kinases (JNKs) and 5’AMP-activated protein kinase (AMPK), and were also possibly more sensitive to dynamic changes in energy. Females expressed higher major histocompatibility complex (MHC) class II levels than males, enhancing resistance to pathogens, and the DEGs related to reproduction included MAPK, CaMK, CPEB, and Cdc25. The genes related to stress, energy, and immunity were also likely related to the regulation of longevity. The upregulated JNKs in females might prolong lifespan and relieve antioxidant stress. Females may also activate the AMPK pathway and implement dietary restrictions to prolong lifespan, whereas males may upregulate SIRT1 and CRAB to increase lifespan.ConclusionsFemale magpies might be more sensitive to stress and dynamic changes in energy thus enhanced resistance to pathogens, and the genes related to stress, energy, and immunity were also possibly related to the regulation of longevity. Further confirmations with techniques such as RT-qPCR and western blot are necessary to validate the above arguments.