Allelic Lineages Research Articles

An archaic HLA class I receptor allele diversifies natural killer cell-driven immunity in First Nations peoples of Oceania

Genetic variation in host immunity impacts the disproportionate burden of infectious diseases that can be experienced by First Nations peoples. Polymorphic human leukocyte antigen (HLA) class I and killer cell immunoglobulin-like receptors (KIRs) are key regulators of natural killer (NK) cells, which mediate early infection control. How this variation impacts their responses across populations is unclear. We show that HLA-A∗24:02 became the dominant ligand for inhibitory KIR3DL1 in First Nations peoples across Oceania, through positive natural selection. We identify KIR3DL1∗114, widespread across and unique to Oceania, as an allele lineage derived from archaic humans. KIR3DL1∗114+NK cells from First Nations Australian donors are inhibited through binding HLA-A∗24:02. The KIR3DL1∗114 lineage is defined by phenylalanine at residue 166. Structural and binding studies show phenylalanine 166 forms multiple unique contacts with HLA-peptide complexes, increasing both affinity and specificity. Accordingly, assessing immunogenetic variation and the functional implications for immunity are fundamental toward understanding population-based disease associations.

Analysis of the Origin of Emiratis as Inferred from a Family Study Based on HLA-A, -C, -B, -DRB1, and -DQB1 Genes.

Two-hundred unrelated Emirati parents of patients selected for bone marrow transplantation were genotyped for HLA class I (A, B, C) and class II (DRB1, DQB1) genes using reverse sequence specific oligonucleotide bead-based multiplexing. HLA haplotypes were assigned with certainty by segregation (pedigree) analysis, and haplotype frequencies were obtained by direct counting. HLA class I and class II frequencies in Emiratis were compared to data from other populations using standard genetic distances (SGD), Neighbor-Joining (NJ) phylogenetic dendrograms, and correspondence analysis. The studied HLA loci were in Hardy-Weinberg Equilibrium. We identified 17 HLA-A, 28 HLA-B, 14 HLA-C, 13 HLA-DRB1, and 5 HLA-DQB1 alleles, of which HLA-A*02 (22.2%), -B*51 (19.5%), -C*07 (20.0%), -DRB1*03 (22.2%), and -DQB1*02 (32.8%) were the most frequent allele lineages. DRB1*03~DQB1*02 (21.2%), DRB1*16~DQB1*05 (17.3%), B*35~C*04 (11.7%), B*08~DRB1*03 (9.7%), A*02~B*51 (7.5%), and A*26~C*07~B*08~DRB1*03~DQB1*02 (4.2%) were the most frequent two- and five-locus HLA haplotypes. Correspondence analysis and dendrograms showed that Emiratis were clustered with the Arabian Peninsula populations (Saudis, Omanis and Kuwaitis), West Mediterranean populations (North Africans, Iberians) and Pakistanis, but were distant from East Mediterranean (Turks, Albanians, Greek), Levantine (Syrians, Palestinians, Lebanese), Iranian, Iraqi Kurdish, and Sub-Saharan populations. Emiratis were closely related to Arabian Peninsula populations, West Mediterranean populations and Pakistanis. However, the contribution of East Mediterranean, Levantine Arab, Iranian, and Sub-Saharan populations to the Emiratis' gene pool appears to be minor.

Low Diversity of Major Histocompatibility Complex (MHC) Genes in Endangered Malayan Tapir (Tapirus indicus).

The Malayan tapir (Tapirus indicus) is listed as Endangered on the IUCN Red List due to multiple threats such as habitat loss and human disturbance that have led to its population decline. This decline increases the risk of inbreeding, which could result in the reduction of genome-wide genetic variation and negatively affect the gene responsible for immune response i.e., MHC gene. Class I and II MHC genes are responsible for encoding MHC molecules in the cells that recognise pathogenic peptides and present them to T-Cells on the cell surface for adaptive immune response. However, at present there is no study related to the MHC gene in Malayan tapir yet. This study characterises the MHC class I and II genes from seven individuals, investigates evidence of balancing selection and their relationships with homologous genes of other species. We identified at least one class I gene and four class II genes. Five sequences of alpha1 (α1) and four of alpha2 (α2) domains of class I alleles, two DRA, two DQA, three DRB and three DQB of class II alleles were isolated. α1 and α2 domains of class I and DRB domain of class II displayed evidence of selection with a higher rate of non-synonymous over synonymous substitutions. Within the DRB gene, 24 codons were found to be under selection where 10 are part of the codons forming the Antigen Binding Site. Genes sequences show species-specific monophyletic group formation except for class I and DRB genes with intersperse relationship in their phylogenetic trees which may indicate occurrence of trans-species polymorphism of allelic lineage. More studies using RNA samples are needed to identify the gene's level of expression.

Balanced Polymorphism at the Pgm-1 Locus of the Pompeii Worm Alvinella pompejana and Its Variant Adaptability Is Only Governed by Two QE Mutations at Linked Sites.

The polychaete Alvinella pompejana lives exclusively on the walls of deep-sea hydrothermal chimneys along the East Pacific Rise (EPR), and displays specific adaptations to withstand the high temperatures and hypoxia associated with this highly variable habitat. Previous studies have revealed the existence of a balanced polymorphism on the enzyme phosphoglucomutase associated with thermal variations, where allozymes 90 and 100 exhibit different optimal activities and thermostabilities. Exploration of the mutational landscape of phosphoglucomutase 1 revealed the maintenance of four highly divergent allelic lineages encoding the three most frequent electromorphs over the geographic range of A. pompejana. This polymorphism is only governed by two linked amino acid replacements, located in exon 3 (E155Q and E190Q). A two-niche model of selection, including ‘cold’ and ‘hot’ conditions, represents the most likely scenario for the long-term persistence of these isoforms. Using directed mutagenesis and the expression of the three recombinant variants allowed us to test the additive effect of these two mutations on the biochemical properties of this enzyme. Our results are coherent with those previously obtained from native proteins, and reveal a thermodynamic trade-off between protein thermostability and catalysis, which is likely to have maintained these functional phenotypes prior to the geographic separation of populations across the Equator about 1.2 million years ago.

Fine-Scale Genetic Structure and Natural Selection Signatures of Southwestern Hans Inferred From Patterns of Genome-Wide Allele, Haplotype, and Haplogroup Lineages.

The evolutionary and admixture history of Han Chinese have been widely discussed via traditional autosomal and uniparental genetic markers [e.g., short tandem repeats, low-density single nucleotide polymorphisms). However, their fine-scale genetic landscapes (admixture scenarios and natural selection signatures) based on the high-density allele/haplotype sharing patterns have not been deeply characterized. Here, we collected and generated genome-wide data of 50 Han Chinese individuals from four populations in Guizhou Province, one of the most ethnolinguistically diverse regions, and merged it with over 3,000 publicly available modern and ancient Eurasians to describe the genetic origin and population admixture history of Guizhou Hans and their neighbors. PCA and ADMIXTURE results showed that the studied four populations were homogeneous and grouped closely to central East Asians. Genetic homogeneity within Guizhou populations was further confirmed via the observed strong genetic affinity with inland Hmong-Mien people through the observed genetic clade in Fst and outgroup f3/f4-statistics. qpGraph-based phylogenies and f4-based demographic models illuminated that Guizhou Hans were well fitted via the admixture of ancient Yellow River Millet farmers related to Lajia people and southern Yangtze River farmers related to Hanben people. Further ChromoPainter-based chromosome painting profiles and GLOBETROTTER-based admixture signatures confirmed the two best source matches for southwestern Hans, respectively, from northern Shaanxi Hans and southern indigenes with variable mixture proportions in the historical period. Further three-way admixture models revealed larger genetic contributions from coastal southern East Asians into Guizhou Hans compared with the proposed inland ancient source from mainland Southeast Asia. We also identified candidate loci (e.g., MTUS2, NOTCH4, EDAR, ADH1B, and ABCG2) with strong natural selection signatures in Guizhou Hans via iHS, nSL, and ihh, which were associated with the susceptibility of the multiple complex diseases, morphology formation, alcohol and lipid metabolism. Generally, we provided a case and ideal strategy to reconstruct the detailed demographic evolutionary history of Guizhou Hans, which provided new insights into the fine-scale genomic formation of one ethnolinguistically specific targeted population from the comprehensive perspectives of the shared unlinked alleles, linked haplotypes, and paternal and maternal lineages.

Haplotypic Associations and Differentiation of MHC Class II Polymorphic Alu Insertions at Five Loci With HLA-DRB1 Alleles in 12 Minority Ethnic Populations in China.

The analysis of polymorphic variations in the human major histocompatibility complex (MHC) class II genomic region on the short-arm of chromosome 6 is a scientific enquiry to better understand the diversity in population structure and the effects of evolutionary processes such as recombination, mutation, genetic drift, demographic history, and natural selection. In order to investigate associations between the polymorphisms of HLA-DRB1 gene and recent Alu insertions (POALINs) in the HLA class II region, we genotyped HLA-DRB1 and five Alu loci (AluDPB2, AluDQA2, AluDQA1, AluDRB1, AluORF10), and determined their allele frequencies and haplotypic associations in 12 minority ethnic populations in China. There were 42 different HLA-DRB1 alleles for ethnic Chinese ranging from 12 alleles in the Jinuo to 28 in the Yugur with only DRB1∗08:03, DRB1∗09:01, DRB1∗12:02, DRB1∗14:01, DRB1∗15:01, and DRB1∗15:02 present in all ethnic groups. The POALINs varied in frequency between 0.279 and 0.514 for AluDPB2, 0 and 0.127 for AluDQA2, 0.777 and 0.995 for AluDQA1, 0.1 and 0.455 for AluDRB1 and 0.084 and 0.368 for AluORF10. By comparing the data of the five-loci POALIN in 13 Chinese ethnic populations (including Han-Yunnan published data) against Japanese and Caucasian published data, marked differences were observed between the populations at the allelic or haplotypic levels. Five POALIN loci were in significant linkage disequilibrium with HLA-DRB1 in different populations and AluDQA1 had the highest percentage association with most of the HLA-DRB1 alleles, whereas the nearby AluDRB1 indel was strongly haplotypic for only DRB1∗01, DRB1∗10, DRB1∗15 and DRB1∗16. There were 30 five-locus POALIN haplotypes inferred in all populations with H5 (no Alu insertions except for AluDQA1) and H21 (only AluDPB2 and AluDQA1 insertions) as the two predominant haplotypes. Neighbor joining trees and principal component analyses of the Alu and HLA-DRB1 polymorphisms showed that genetic diversity of these genomic markers is associated strongly with the population characteristics of language family, migration and sociality. This comparative study of HLA-DRB1 alleles and multilocus, lineage POALIN frequencies of Chinese ethnic populations confirmed that POALINs whether investigated alone or together with the HLA class II alleles are informative genetic and evolutionary markers for the identification of allele and haplotype lineages and genetic variations within the same and/or different populations.

Characterization of LILRB3 and LILRA6 allelic variants in the Japanese population.

Leukocyte immunoglobulin (Ig)-like receptors (LILRs) are encoded by members of a human multigene family, comprising 11 protein-coding genes and two pseudogenes. Among the LILRs, LILRB3 and LILRA6 show the highest homology with each other, along with high allelic and copy number variations. Therefore, it has been difficult to discriminate between them, both genetically and functionally, precluding disease association studies of LILRB3 and LILRA6. In this study, we carefully performed variant screening of LILRB3 and LILRA6 by cDNA cloning from Japanese individuals and identified four allelic lineages showing significantly high non-synonymous-to-synonymous ratios in pairwise comparisons. Furthermore, the extracellular domains of the LILRB3*JP6 and LILRA6*JP1 alleles were identical at the DNA level, suggesting that gene conversion-like events diversified LILRB3 and LILRA6. To determine the four allelic lineages from genomic DNA, we established a lineage typing method that accurately estimated the four allelic lineages in addition to specific common alleles from genomic DNA. Analysis of LILRA6 copy number variation revealed one, two, and three copies of LILRA6 in the Japanese-in-Tokyo (JPT) population. Flow cytometric analysis showed that an anti-LILRB3 antibody did not recognize the second most common lineage in the Japanese population, indicating significant amino acid differences across the allelic lineages. Taken together, our findings indicate that our lineage typing is useful for classifying the lineage-specific functions of LILRB3 and LILRA6, serving as the basis for disease association studies.

Molecular Evolution of Antigen-Processing Genes in Salamanders: Do They Coevolve with MHC Class I Genes?

Proteins encoded by antigen-processing genes (APGs) prepare antigens for presentation by the major histocompatibility complex class I (MHC I) molecules. Coevolution between APGs and MHC I genes has been proposed as the ancestral gnathostome condition. The hypothesis predicts a single highly expressed MHC I gene and tight linkage between APGs and MHC I. In addition, APGs should evolve under positive selection, a consequence of the adaptive evolution in MHC I. The presence of multiple highly expressed MHC I genes in some teleosts, birds, and urodeles appears incompatible with the coevolution hypothesis. Here, we use urodele amphibians to test two key expectations derived from the coevolution hypothesis: 1) the linkage between APGs and MHC I was studied in Lissotriton newts and 2) the evidence for adaptive evolution in APGs was assessed using 42 urodele species comprising 21 genera from seven families. We demonstrated that five APGs (PSMB8, PSMB9, TAP1, TAP2, and TAPBP) are tightly linked (<0.5 cM) to MHC I. Although all APGs showed some codons under episodic positive selection, we did not find a pervasive signal of positive selection expected under the coevolution hypothesis. Gene duplications, putative gene losses, and divergent allelic lineages detected in some APGs demonstrate considerable evolutionary dynamics of APGs in salamanders. Overall, our results indicate that if coevolution between APGs and MHC I occurred in urodeles, it would be more complex than envisaged in the original formulation of the hypothesis.

Cartilaginous fish class II genes reveal unprecedented old allelic lineages and confirm the late evolutionary emergence of DM

Cartilaginous fish (chimaeras, rays and sharks) are the most basal extant jawed vertebrates with an adaptive immune system based on the Major Histocompatibility Complex (MHC). Despite being a key taxon in the evolution of vertebrate adaptive immunity, no comprehensive characterization of MHC class II genes has been undertaken for the group. We performed extensive bioinformatic searches on a taxonomically diverse dataset of transcriptomes and genomes of cartilaginous fish targeting MHC class II sequences. Class IIα and IIβ sequences were retrieved from all taxa analyzed and showed typical features of classical class II genes. Phylogenetic trees of the immunoglobulin superfamily domain showed two divergent and remarkably ancient lineages of class II genes in Selachians (sharks), originating >350 million years ago. Close linkage of lineage-specific pairs of IIα and IIβ genes was found, confirming previous results, with genes from distinct lineages segregating as alleles. Nonclassical class II DM sequences were not retrieved from these data and classical class II sequences lacked the conserved residues shown to interact with DM molecules, supporting claims that the DM system arose only in the lobe-finned fish lineage leading to tetrapods. Based on our search methods, other divergent class II genes are unlikely in cartilaginous fish.

Balancing selection versus allele and supertype turnover in MHC class II genes in guppies.

Selection pressure from parasites is thought to be a major force shaping the extreme polymorphism of the major histocompatibility complex (MHC) genes, but the modes and consequences of selection remain unclear. Here, we analyse MHC class II and microsatellite diversity in 16 guppy (Poecilia reticulata) populations from two islands (Trinidad and Tobago) that have been separated for at least 10 ky. Within-population MHC diversity was high, but allele sharing was limited within islands and even lower between islands, suggesting relatively fast turnover of alleles. Allelic lineages strongly supported in phylogenetic analyses tended to be island-specific, suggesting rapid lineage sorting, and an expansion of an allelic lineage private to Tobago was observed. New alleles appear to be generated locally at a detectably high frequency. We did not detect a consistent signature of local adaptation, but FST outlier analysis suggested that balancing selection may be the more general process behind spatial variation in MHC allele frequencies in this system, particularly within Trinidad. We found no evidence for divergent allele advantage within populations, or for decreased genetic structuring of MHC supertypes compared to MHC alleles. The dynamic and complex nature of MHC evolution we observed in guppies, coupled with some evidence for balancing selection shaping MHC allele frequencies, are consistent with Red Queen processes of host-parasite coevolution.

Global allele polymorphism indicates a high rate of allele genesis at a locus under balancing selection.

When selection favours rare alleles over common ones (balancing selection in the form of negative frequency-dependent selection), a locus may maintain a large number of alleles, each at similar frequency. To better understand how allelic richness is generated and maintained at such loci, we assessed 201 sequences of the complementary sex determiner (csd) of the Asian honeybee (Apis cerana), sampled from across its range. Honeybees are haplodiploid; hemizygotes at csd develop as males and heterozygotes as females, while homozygosity is lethal. Thus, csd is under strong negative frequency-dependent selection because rare alleles are less likely to end up in the lethal homozygous form. We find that in A. cerana, as in other Apis, just a few amino acid differences between csd alleles in the hypervariable region are sufficient to trigger female development. We then show that while allelic lineages are spread across geographical regions, allelic differentiation is high between populations, with most csd alleles (86.3%) detected in only one sample location. Furthermore, nucleotide diversity in the hypervariable region indicates an excess of recently arisen alleles, possibly associated with population expansion across Asia since the last glacial maximum. Only the newly invasive populations of the Austral-Pacific share most of their csd alleles. In all, the geographic patterns of csd diversity in A. cerana indicate that high mutation rates and balancing selection act together to produce high rates of allele genesis and turnover at the honeybee sex locus, which in turn leads to its exceptionally high local and global polymorphism.

Two Lineages of KLRA with Contrasting Transcription Patterns Have Been Conserved at a Single Locus during Ruminant Speciation.

Cattle possess the most diverse repertoire of NK cell receptor genes among all mammals studied to date. Killer cell receptor genes encoded within the NK complex and killer cell Ig-like receptor genes encoded within the leukocyte receptor complex have both been expanded and diversified. Our previous studies identified two divergent and polymorphic KLRA alleles within the NK complex in the Holstein-Friesian breed of dairy cattle. By examining a much larger cohort and other ruminant species, we demonstrate the emergence and fixation of two KLRA allele lineages (KLRA*01 and -*02) at a single locus during ruminant speciation. Subsequent recombination events between these allele lineages have increased the frequency of KLRA*02 extracellular domains. KLRA*01 and KLRA*02 transcription levels contrasted in response to cytokine stimulation, whereas homozygous animals consistently transcribed higher levels of KLRA, regardless of the allele lineage. KLRA*02 mRNA levels were also generally higher than KLRA*01 Collectively, these data point toward alternative functional roles governed by KLRA genotype and allele lineage. On a background of high genetic diversity of NK cell receptor genes, this KLRA allele fixation points to fundamental and potentially differential function roles.

Two-dimensional site frequency spectrum for detecting, classifying and dating incomplete selective sweeps

The two-dimensional site frequency spectrum (2D SFS) was investigated to describe the intra-allelic variability (IAV) maintained within a derived allele (D) group that has undergone an incomplete selective sweep against an ancestral allele group. We observed that recombination certainly muddles the ancestral relationships of allelic lineages between the two allele groups; however, the 2D SFS reveals intriguing signatures of recombination as well as the genealogical structure of the D group, particularly the size of a mutation and the time to the most recent common ancestor (TMRCA). Coalescent simulations were performed to achieve powerful and robust 2D SFS-based statistics with special reference to accurate evaluation of IAV, significance of recombination effects, and distinction between hard and soft selective sweeps. These studies were extended to a case wherein an incomplete selective sweep is no longer in progress and ceased in the recent past. The 2D SFS-based method was applied to 100 intronic linkage disequilibrium regions randomly chosen from the East Asian population of modern humans to examine the P value distributions of the summary statistics under the null hypothesis of neutrality in a nonequilibrium demographic model. We argue that about 96% of intronic variants are non-adaptive with a 10% false discovery rate. Furthermore, this method was applied to six genomic regions in Eurasian populations that were claimed to have experienced recent selective sweeps. We found that two of these genomic regions did not have significant signals of selective sweeps, but the remaining four had undergone hard and soft sweeps and were dated, in terms of TMRCA, after the major out-of-Africa dispersal of modern humans.

Molecular Basis of ABO Variants Including Identification of 16 Novel ABO Subgroup Alleles in Chinese Han Population

Introduction: The characteristic of ABO blood subgroup is crucial for elucidating the mechanisms of such variant phenotypes and offering useful information in blood transfusion. Methods: In total, 211 ABO variants including part of available family members were investigated in this study. The phenotypes of these individuals were typed with serologic methods. The full coding regions of ABO gene and the erythroid cell-specific regulatory elements in intron 1 of them were amplified with polymerase chain reaction and then directly sequenced. The novel alleles were confirmed by cloning and sequencing. Phylogenetic tree was made using CLUSTAL W software. 3D structural analyses of the glycosyltransferases (GTs) with some typical mutations were performed by PyMOL software. Results: Forty-eight distinctly rare ABO alleles were identified in 211 Chinese variant individuals, including 16 novel ABO alleles. All of the alleles were categorized as 5 groups: 16 ABO*A alleles, 23 ABO*B alleles, 4 ABO*BA alleles, 4 ABO*cisAB alleles, and 1 ABO*O alleles. ABO*A2.08 and ABO*BA.02 were the relatively predominant A and B subgroup alleles, respectively. According to the phylogenetic tree, 28 alleles (5 common alleles and 23 alleles identified in our laboratory) were classified into 3 major allelic lineages. The structural analysis of 3D homology modeling predicted reduced protein stability of the mutant GTs and may explain the reduced ABO antigen expression. Conclusions: The molecular basis of ABO variants was analyzed, and 16 novel ABO alleles were identified. The results extended the information of ABO variants and provided a basis for better transfusion strategies and helped to improve blood transfusion safety.

MHC class I polymorphic Alu insertion (POALIN) allele and haplotype frequencies in the Arabs of the United Arab Emirates and other world populations

Polymorphic Alu insertions (POALINs) are found throughout the human genome and have been used in various studies to infer geographic origin of human populations. The main aim of this study was to determine the allele and haplotype frequencies of five POALINs, AluHF, AluHG, AluHJ, AluTF and AluMICB, within the major histocompatibility complex (MHC) class I region of 95 UAE Arabs, and correlate their frequencies to those of the HLA-A, HLA-C and HLA-B class I allele lineages. Evolutionary relationships between the POALINs of the Arabs and those previously studied in populations of African, Asian and European descent were compared. At each of the five Alu loci (AluHF, AluHG, AluHJ, AluTF and AluMICB), Alu insertion was designated as Alu(locus)*02 and absence was Alu(locus)*01. The AluHG insertion (AluHG*02) had the highest frequency (0.332), followed by AluHF*02 (0.300), AluHJ*02 (0.263), AluMICB*02 (0.111) and AluTF*02 (0.058). Of the 270 Alu-HLA haplotypes pairs in the UAE Arabs, 110 had no Alu insertion, and 54 had an Alu insertion at >50% per haplotype. An Alu insertion >75% per haplotype was found between AluMICB*02 and HLA-B*14, HLA-B*22, HLA-B*44, HLA-B*55, HLA-B*57 and HLA-B*73, and with HLA-C*01 and HLA-C*18; AluHJ*02 with HLA-A*01, HLA-A*19, HLA-A*24 and HLA-A*32; AluHG*02 with HLA-A*02 and HLA-B*18; and AluHF*02 with HLA-A*10. The genotyped allele and haplotype frequencies of the MHC POALINs in UAE Arabs were compared with the results of 30 previously published Asian, European, American and African populations. Phylogenetic and multidimensional scaling (MDS) analysis of the relative MHC POALINs allele and haplotype frequencies revealed that the UAE Arabs have a similar lineage to Caucasians and the most distant genetic relationship to the Waorani native American population of Ecuador. The structure of both the phylogenetic tree and the MDS analysis supports the Out of Africa theory of human evolution. The nature of the clusters suggests the Arabian Middle East represents a crossroads from which human populations migrated towards Asia in the east and Europe to the north-west.

Expression estimation and eQTL mapping for HLA genes with a personalized pipeline.

The HLA (Human Leukocyte Antigens) genes are well-documented targets of balancing selection, and variation at these loci is associated with many disease phenotypes. Variation in expression levels also influences disease susceptibility and resistance, but little information exists about the regulation and population-level patterns of expression. This results from the difficulty in mapping short reads originated from these highly polymorphic loci, and in accounting for the existence of several paralogues. We developed a computational pipeline to accurately estimate expression for HLA genes based on RNA-seq, improving both locus-level and allele-level estimates. First, reads are aligned to all known HLA sequences in order to infer HLA genotypes, then quantification of expression is carried out using a personalized index. We use simulations to show that expression estimates obtained in this way are not biased due to divergence from the reference genome. We applied our pipeline to the GEUVADIS dataset, and compared the quantifications to those obtained with reference transcriptome. Although the personalized pipeline recovers more reads, we found that using the reference transcriptome produces estimates similar to the personalized pipeline (r ≥ 0.87) with the exception of HLA-DQA1. We describe the impact of the HLA-personalized approach on downstream analyses for nine classical HLA loci (HLA-A, HLA-C, HLA-B, HLA-DRA, HLA-DRB1, HLA-DQA1, HLA-DQB1, HLA-DPA1, HLA-DPB1). Although the influence of the HLA-personalized approach is modest for eQTL mapping, the p-values and the causality of the eQTLs obtained are better than when the reference transcriptome is used. We investigate how the eQTLs we identified explain variation in expression among lineages of HLA alleles. Finally, we discuss possible causes underlying differences between expression estimates obtained using RNA-seq, antibody-based approaches and qPCR.

Distribution of ancient α1 and α2 domain lineages between two classical MHC class I genes and their alleles in grass carp.

Major histocompatibility complex (MHC) class I molecules play a crucial role in the immune response by binding and presenting pathogen-derived peptides to specific CD8+ T cells. From cDNA of 20 individuals of wild grass carp (Ctenopharyngodon idellus), we could amplify one or two alleles each of classical MHC class I genes Ctid-UAA and Ctid-UBA. In total, 27 and 22 unique alleles of Ctid-UAA and Ctid-UBA were found. The leader, α1, transmembrane and cytoplasmic regions distinguish between Ctid-UAA and Ctid-UBA, and their encoded α1 domain sequences belong to the ancient lineages α1-V and α1-II, respectively, which separated several hundred million years ago. However, Ctid-UAA and Ctid-UBA share allelic lineage variation in their α2 and α3 sequences, in a pattern suggestive of past interlocus recombination events that transferred α2+α3 fragments. The allelic Ctid-UAA and Ctid-UBA variation involves ancient variation between domain lineages α2-I and α2-II, which in the present study was dated back to before the ancestral separation of teleost fish and spotted gar (> 300 million years ago). This is the first report with compelling evidence that recombination events combining different ancient α1 and α2 domain lineages had a major impact on the allelic variation of two different classical MHC class I genes within the same species.

HLA class I allele lineages and haplotype frequencies in Arabs of the United Arab Emirates.

The high degree of polymorphism of the HLA system provides suitable genetic markers to study the diversity and migration of different world populations and is beneficial for forensic identification, anthropology, transplantation and disease associations. Although the United Arab Emirates (UAE) population of about nine million people is heterogeneous, information is limited for the HLA class I allele and haplotype frequencies of the Bedouin ethnic group. We performed low-resolution PCR-SSP genotyping of three HLA class I loci at HLA-A, -B and -C for 95 unrelated healthy Bedouins from the cities of Al Ain and Abu Dhabi in the UAE. A total of 54 HLA allele lineages were detected; the most frequent low-resolution allele lineages at each HLA locus were A*02 (0.268), B*51 (0.163) and C*07 (0.216). The inferred estimates for the two most frequent HLA-A and HLA-B haplotypes were HLA-A*02~HLA-B*50 (0.070) and HLA-A*02~HLA-B*51 (0.051), and the most frequent 3-locus haplotype was HLA-A*02 ~ HLA-B*50~HLA-C*06 (0.068). The HLA allele lineage frequencies of the UAE Arabs were compared to those previously reported for 70 other world populations, and a strong genetic similarity was detected between the UAE Arabs and the Saudi Arabians from the west with evidence of a limited gene flow between the UAE Arabs and Pakistani across the Gulf from the east, and the UAE Arabs and Omani from the south of the Gulf Peninsula.

Plasmodium vivax Pv12 B-cell epitopes and HLA-DRβ1*-dependent T-cell epitopes in vitro antigenicity

Malaria is an infectious disease caused by parasites from the genus Plasmodium (P. falciparum and P. vivax are responsible for 90% of all clinical cases); it is widely distributed throughout the world’s tropical and subtropical regions. The P. vivax Pv12 protein is involved in invasion, is expressed on merozoite surface and has been recognised by antibodies from individuals exposed to the disease. In this study, B- and T-cell epitopes from Pv12 were predicted and characterised to advance in the design of a peptide-based vaccine against malaria.For evaluating the humoral response of individuals exposed to natural P. vivax infection from two endemic areas in Colombia, BepiPred-1.0 software was used for selecting B-cell epitopes. B-cell epitope 39038 displayed the greatest recognition by naturally-acquired antibodies and induced an IgG2/IgG4 response. NetMHCIIpan-3.1 prediction software was used for selecting peptides having high affinity binding for HLA-DRβ1* allele lineages and this was confirmed by in-vitro binding assays. T-epitopes 39113 and 39117 triggered a memory T-cell response (Stimulation Index≥2) and significant cytokine production.Combining in-silico, in-vitro and functional assays, two Pv12 protein regions (containing peptides 39038, 39040, 39113 and 39117) have thus been characterised as promising vaccine candidates against P. vivax malaria.

Genetic variation and selection of MHC class I loci differ in two congeneric frogs.

Major histocompatibility complex (MHC) genes encode proteins in the acquired immune response pathway that often show distinctive selection-driven patterns in wild vertebrate populations. We examined genetic variation and signatures of selection in the MHC class I alpha 1 (A1)- and alpha 2 (A2)-domain encoding exons of two frog congeners [Agalychnis callidryas (n = 20) and A. lemur (n = 20)] from a single locality in Panama. We also investigated how historical demographic processes may have impacted MHC genetic diversity by analyzing a neutral mitochondrial marker. We found that both MHC domains were highly variable in both species, with both species likely expressing three loci. Our analyses revealed different signatures of selection between the two species, most notably that the A. callidryas A2 domain had experienced positive selection while the A2 domain of A. lemur had not. Diversifying selection acted on the same number of A1 and A2 allelic lineages, but on a higher percentage of A1 sites compared to A2 sites. Neutrality tests of mitochondrial haplotypes predominately indicated that the two species were at genetic equilibrium when the samples were collected. In addition, two historical tests of demography indicated both species have had relatively stable population sizes over the past 100,000 years; thus large population size changes are unlikely to have greatly influenced MHC diversity in either species during this time period. In conclusion, our results suggest that the impact of selection on MHC diversity varied between these two closely related species, likely due to a combination of distinct ecological conditions and past pathogenic pressures.