HLA-E Alleles Research Articles

The Novel HLA-E Allele, HLA-E*01:151, Identified in a Chinese Individual.

HLA-E*01:151 differs from HLA-E*01:03:01:01 by a single nucleotide substitution at position 890 C>T in exon 5.

Accurate multi-population imputation of MICA, MICB, HLA-E, HLA-F and HLA-G alleles from genome SNP data.

In addition to the classical HLA genes, the major histocompatibility complex (MHC) harbors a high number of other polymorphic genes with less established roles in disease associations and transplantation matching. To facilitate studies of the non-classical and non-HLA genes in large patient and biobank cohorts, we trained imputation models for MICA, MICB, HLA-E, HLA-F and HLA-G alleles on genome SNP array data. We show, using both population-specific and multi-population 1000 Genomes references, that the alleles of these genes can be accurately imputed for screening and research purposes. The best imputation model for MICA, MICB, HLA-E, -F and -G achieved a mean accuracy of 99.3% (min, max: 98.6, 99.9). Furthermore, validation of the 1000 Genomes exome short-read sequencing-based allele calling against a clinical-grade reference data showed an average accuracy of 99.8%, testifying for the quality of the 1000 Genomes data as an imputation reference. We also fitted the models for Infinium Global Screening Array (GSA, Illumina, Inc.) and Axiom Precision Medicine Research Array (PMRA, Thermo Fisher Scientific Inc.) SNP content, with mean accuracies of 99.1% (97.2, 100) and 98.9% (97.4, 100), respectively.

Nomenclature for factors of the HLA system, update January, February, and March 2023.

Nomenclature for factors of the HLA system, update January, February, and March 2023.

86 novel HLA-E alleles discovered through full-gene sequencing of 6227 hematopoietic cell transplant patients and unrelated donors.

Until recently the number of alleles of the nonclassical HLA class I gene HLA-E documented in the IPD-IMGT/HLA Database was small and as a result, the gene was often not considered to be notably polymorphic. Here, we describe our work in identifying and submitting 86 novel HLA-E alleles after full-gene single-molecule real-time (SMRT) DNA sequencing of 6227 DNA samples. These samples were comprised of 2468 patients undergoing hematopoietic cell transplantation and 3759 unrelated potential donors. A total of 111 unique HLA-E alleles were detected in this cohort. The majority of novel alleles (79.1%) contained polymorphisms in intronic regions, highlighting the significant undiscovered variation present in the noncoding regions of the HLA-E gene.

Ramifications of the HLA-I Allelic Reactivity of Anti-HLA-E*01:01 and Anti-HLA-E*01:03 Heavy Chain Monoclonal Antibodies in Comparison with Anti-HLA-I IgG Reactivity in Non-Alloimmunized Males, Melanoma-Vaccine Recipients, and End-Stage Renal Disease Patients.

Serum anti-HLA-I IgG are present in non-alloimmunized males, cancer patients, and transplant recipients. Anti-HLA-I antibodies are also present in intravenous immunoglobulin (IVIg), prepared from the plasma of thousands of healthy donors. However, the HLA-Ia reactivity of IVIg diminishes markedly after passing through HLA-E HC-affinity columns, suggesting that the HLA-I reactivity is due to antibodies formed against HLA-E. Hence, we examined whether anti-HLA-E antibodies can react to HLA-I alleles. Monoclonal IgG antibodies (mAbs) against HCs of two HLA-E alleles were generated in Balb/C mice. The antibodies were analyzed using multiplex bead assays on a Luminex platform for HLA-I reactivity. Beads coated with an array of HLA heterodimers admixed with HCs (LABScreen) were used to examine the binding of IgG to different HLA-Ia (31-HLA-A, 50-HLA-B, and 16-HLA-C) and Ib (2-HLA-E, one each of HLA-F and HLA-G) alleles. A striking diversity in the HLA-Ia and/or HLA-Ib reactivity of mAbs was observed. The number of the mAbs reactive to (1) only HLA-E (n = 25); (2) all HLA-Ib isomers (n = 8); (3) HLA-E and HLA-B (n = 5); (4) HLA-E, HLA-B, and HLA-C (n = 30); (5) HLA-E, HLA-A*1101, HLA-B, and HLA-C (n = 83); (6) HLA-E, HLA-A, HLA-B, and HLA-C (n = 54); and (7) HLA-Ib and HLA-Ia (n = 8), in addition to four other minor groups. Monospecificity and polyreactivity were corroborated by HLA-E monospecific and HLA-I shared sequences. The diverse HLA-I reactivity of the mAbs are compared with the pattern of HLA-I reactivity of serum-IgG in non-alloimmunized males, cancer patients, and ESKD patients. The findings unravel the diagnostic potential of the HLA-E monospecific-mAbs and immunomodulatory potentials of IVIg highly mimicking HLA-I polyreactive-mAbs.

Correlation of Human Leukocyte Antigen-E Genomic Polymorphism with Leukemia and Functional Study of Human Leukocyte Antigen-E Different Type Promoters.

Human leukocyte antigen (HLA)-E is one of the least polymorphic nonclassical major histocompatibility complex (MHC) I genes; its nucleotide variability can affect immune response. In this study, we assess the correlation between HLA-E polymorphism and leukemia and further study the transcriptional activity of promoter variation at nucleotide position-26. A total of 142 healthy blood donors and 111 leukemia patients were collected. The genomic sequence of HLA-E was amplified by high-fidelity reaction system and identified by Sanger and cloning sequencing. The dual luciferase reporter gene assay was used to detect the transcription activity of promoter variation at nucleotide position-26. In the HLA-E genomic sequence results, a total of 16 alleles and 32 genotypes were detected; the HLA-E*01:01:01:06 allele had a significantly lower frequency in leukemia patients than in healthy participants (p = 0.026 < 0.05). And the HLA-E*01:03:02:01, *01:03:02:01 genotype showed the greatest difference in frequency between the two groups of participants (p = 0.028 < 0.05). Eight HLA-E alleles were first reported worldwide in Chinese individuals. The results of the dual luciferase reporter gene experiment showed that the transcription activity of the mutant-type promoter (HLA-E*01:01:01:06 with "T" allele at nucleotide position-26) was significantly lower compared with the wild-type promoter (HLA-E*01:01:01:01 with "G" allele at nucleotide position-26) (p = 0.0242 < 0.05). HLA-E*01:01:01:06 allele has a protective effect against leukemia through decreasing transcription activity by "T" variation at nucleotide position-26.

Dissection of the NKG2C NK cell response against Puumala Orthohantavirus.

BackgroundInfections with the Puumala orthohantavirus (PUUV) in humans may cause hemorrhagic fever with renal syndrome (HFRS), known as nephropathia epidemica (NE), which is associated with acute renal failure in severe cases. In response to PUUV-infections, a subset of potent antiviral NKG2C+ NK cells expand, whose role in virus defence and pathogenesis of NE is unclear. NKG2C+ NK cell proliferation is mediated by binding of NKG2C/CD94 to HLA-E on infected cells. The proliferation and activation of NKG2C+ NK cells via the NKG2C/HLA-E axis is affected by different NKG2C (NKG2Cwt/del) and HLA-E (HLA-E*0101/0103) alleles, which naturally occur in the human host. Homozygous (NKG2Cdel/del) and heterozygous (NKG2Cwt/del) deletions of the NKG2C receptor results in an impaired NKG2C/CD94 mediated proliferation and activation of NKG2C+ cells. We therefore analyzed the PUUV-mediated NKG2C+ NK cell responses and the impact of different NKG2C and HLA-E alleles in NE patients.Methodology/Principal findingsNKG2C+ NK cell expansion and effector functions in PUUV-infected cells were investigated using flow cytometry and it was shown that PUUV-infected endothelial cells led to a NKG2C/CD94 mediated NKG2C+ NK cell activation and expansion, dependent on the HLA-G-mediated upregulation of HLA-E. Furthermore, the NKG2Cdel and HLA-E*0101/0103 alleles were determined in 130 NE patients and 130 matched controls, and it was shown that in NE patients the NKG2Cwt/del allele was significantly overrepresented, compared to the NKG2Cwt/wt variant (p = 0.01). In addition, in vitro analysis revealed that NKG2Cwt/del NK cells exhibited on overall a lower proliferation (p = 0.002) and lower IFNγ expression (p = 0.004) than NKG2Cwt/wt NK cells.Conclusions/SignificanceOur results corroborate the substantial impact of the NKG2C/HLA-E axis on PUUV-specific NK cell responses. A weak NKG2C+ NK cell response, as reflected by NKG2Cwt/del variant, may be associated with a higher risk for a severe hantavirus infections.

A novel and efficient approach to high-throughput production of HLA-E/peptide monomer for T-cell epitope screening

Over the past two decades, there has been a great interest in the study of HLA-E-restricted αβ T cells during bacterial and viral infections, including recently SARS-CoV-2 infection. Phenotyping of these specific HLA-E-restricted T cells requires new tools such as tetramers for rapid cell staining or sorting, as well as for the identification of new peptides capable to bind to the HLA-E pocket. To this aim, we have developed an optimal photosensitive peptide to generate stable HLA-E/pUV complexes allowing high-throughput production of new HLA-E/peptide complexes by peptide exchange. We characterized the UV exchange by ELISA and improved the peptide exchange readout using size exclusion chromatography. This novel approach for complex quantification is indeed very important to perform tetramerization of MHC/peptide complexes with the high quality required for detection of specific T cells. Our approach allows the rapid screening of peptides capable of binding to the non-classical human HLA-E allele, paving the way for the development of new therapeutic approaches based on the detection of HLA-E-restricted T cells.

HLA-E diversity unfolded: Identification and characterization of 170 novel HLA-E alleles.

HLA‐E is a member of the nonclassical HLA class Ib genes. Even though it is structurally highly similar to the classical HLA class Ia genes, it is less diverse and only 45 alleles and 12 proteins were known in December 2019 (IPD‐IMGT/HLA, release 3.38.0). Since 2017, we have genotyped over 3 million voluntary stem cell donors for HLA‐E by sequencing the most relevant allele‐determining bases of exons 2 and 3. As expected, most donors harbor the two predominant alleles HLA‐E*01:01 and/or HLA‐E*01:03. However, in 1666 (0.05%) of our samples we detected 345 distinct novel HLA‐E sequences. The most frequent one was identified in 162 samples and has by now been named HLA‐E*01:114. To characterize these novel alleles in full‐length, we used both short‐read Illumina and long‐read PacBio sequencing to obtain fully phased and highly accurate sequences. This resulted in 234 submissions to IPD‐IMGT/HLA comprising 170 novel HLA‐E alleles, which encode for 93 novel HLA‐E proteins, as well as 64 confirmations or sequence extensions. Consequently, the number of HLA‐E alleles in the database (release 3.42.0) has now increased to 256 HLA‐E alleles and 110 HLA‐E proteins.

HLA-E typing of more than 2.5 million potential hematopoietic stem cell donors: Methods and population-specific allele frequencies

We developed a cost-efficient workflow for genotyping HLA-E by NGS and applied it for genotyping more than 2.5 million potential stem cell donors. The data obtained were used to determine HLA-E allele frequency distributions for 104 populations. Our results confirm the known dominance of the alleles E*01:01 and E*01:03, which have a combined frequency of more than 0.99 in 97 of the 104 populations. E*01:01 is more frequent in Africa and the western part of South America, E*01:03 in Southeast and East Asia. E*01:03 shows a pronounced regional substructure at the high-resolution level with E*01:03:01G being particularly common in a large connected region extending from Turkey to China, E*01:03:02G in Northwestern Europe and E*01:03:03 in Central and Eastern Europe as well as Central Asia. The presented results are relevant both as a basis for further population genetics studies and for optimizing stem cell donor searches.

Single molecule real-time DNA sequencing of the full HLA-E gene for 212 reference cell lines.

We have developed a genotyping assay that produces fully phased, unambiguous HLA-E genotyping using Pacific Biosciences' single molecule real-time DNA sequencing. In total 212 cell lines were genotyped, including the panel of 107 established at the 10th International Histocompatibility Workshop. Our results matched the previously known HLA-E genotype in 94 (44.3%) cell lines, in all cases either improving or equalling previous genotyping resolution. Three (1.4%) cells had discrepant HLA-E genotyping data and 115 (54.2%) had no previous HLA-E data. The HLA-E genotypes for four (1.9%) cell lines resulted in a change of zygosity by identifying two distinct haplotypes. We discovered eight novel HLA-E alleles, extended the known reference sequence of seven and confirmed the existence of a further 10.

Protective Effect of HLA-E∗0101∕∗0103 Genotype in Survival of Patients After Allogeneic Hematopoietic Stem Cell Transplant.

HLA-E is located on the nonclassical major histocompatibility complex class I and acts as the ligand for natural killer cells. Consequently, it has a main role in the regulation of innate immune responses by involving cell identification by natural killer cells. Differences in expression levels among HLA-E alleles have been suggested to affect transplant outcomes. In this study, we evaluated the effects of different HLA-E genotypes on allogeneic hematopoietic stem cell transplant in southern Iran. We investigated 200 patients (donors and recipients) who underwent allogeneic hematopoietic stem-cell transplant and 100 normal participants (control group) in a case-control study. Detection of HLA-E polymorphisms was performed using a sequence-specific primer polymerase chain reaction method. Statistical analyses indicated that genotypes in the transplant group were not distributed in accordance with Hardy-Weinberg equilibrium (χ²=76.56; P < .001), whereas genotypes in the control group were distributed in accordance with Hardy-Weinberg equilibrium (χ²=0.39; P = .53). No significant differences were observed in cumulative incidence of acute (P = .76; hazard ratio = 0.80; 95% confidence interval, 0.19-3.31) and chronic (P = .75, hazard ratio = 0.048; 95% confidence interval, 0.00) graft-versus-host disease in recipients harboring HLA-E∗0103 allele compared with those homozygous for the HLA-E∗0101 allele. The HLA-E∗0103 allele showed a trend toward lower cumulative incidence of relapse compared with the homozygous HLA-E∗0101 genotype (8% vs 21.5%; P = .37; hazard ratio = 2.50; 95% confidence interval, 0.32-19.20). Genotypes of the HLA-E molecule did not correlate with acute and chronic graft-versus-host disease in hematopoietic stem cell transplant recipients except for the HLA-E∗0101∗∕∗0103 genotype, which was protective in survival of our study patients.

A novel HLA-E allele, HLA-E*01:03:01:05, identified in a healthy Chinese blood donor.

HLA-E*01:03:01:05 differs from E*01:01:01:01 by a single nucleotide exchange in nucleotide -33(T->C).

Dimorphism of HLA-E and its Disease Association.

HLA-E is a nonclassical member of the major histocompatibility complex class I gene locus. HLA-E protein shares a high level of homology with MHC Ia classical proteins: it has similar tertiary structure, associates with β2-microglobulin, and is able to present peptides to cytotoxic lymphocytes. The main function of HLA-E under normal conditions is to present peptides derived from the leader sequences of classical HLA class I proteins, thus serving for monitoring of expression of these molecules performed by cytotoxic lymphocytes. However, opposite to multiallelic classical MHC I genes, HLA-E in fact has only two alleles—HLA-E*01:01 and HLA-E*01:03—which differ by one nonsynonymous amino acid substitution at position 107, resulting in an arginine in HLA-E*01:01 (HLA-ER) and glycine in HLA-E*01:03 (HLA-EG). In contrast to HLA-ER, HLA-EG has higher affinity to peptide, higher surface expression, and higher thermal stability of the corresponding protein, and it is more ancient than HLA-ER, though both alleles are presented in human populations in nearly equal frequencies. In the current review, we aimed to uncover the reason of the expansion of the younger allele, HLA-ER, by analysis of associations of both HLA-E alleles with a number of diseases, including viral and bacterial infections, cancer, and autoimmune disorders.

P063 Global distribution of HLA-E alleles based on next-generation sequencing of 1.8 million individuals sampled from 5 national DKMS donor centers

P063 Global distribution of HLA-E alleles based on next-generation sequencing of 1.8 million individuals sampled from 5 national DKMS donor centers

The Role of HLA-E Polymorphism in Acute Leukemia Patients Receiving a 10/10 HLA Matched Unrelated HSCT

Introduction: HLA-E is a class I HLA molecule that is involved in NK and CTL functions. It has limited polymorphism but can present unconventional peptides if it is aberrantly expressed under stress conditions. Donor-recipient (D/R) mismatching may potentiate NK-mediated alloreactivity. The functional role of HLA-E in HSCT is uncertain. We used CIBMTR data and samples to investigate the effect of HLA-E polymorphism on HSCT outcome in a 10/10 HLA matched unrelated acute leukemia setting by addressing the following questions: 1) Does D/R HLA-E match status affect HSCT outcome? 2) Does a specific D/R HLA-E genotype correlate with outcome?Methods: The study included 1840 adult AML/ALL patients in complete remission who received first T-cell replete, 10/10 high resolution HLA matched transplants from unrelated donors between 2000 to 2015. Cohort characteristics are summarized in Table 1. Both D/R were HLA-E genotyped by NGS using a validated protocol on an Illumina Miseq platform. Overall survival (OS), leukemia free survival (LFS), relapse (RI), transplant related mortality (TRM), acute GvHD (aGvHD) and chronic GvHD (cGvHD) were evaluated; p< 0.01 was considered significant. Factors violating the proportional hazards assumption were adjusted via stratification, while a stepwise model building approach was used to select variables related to a given outcome with a threshold of 0.05 for both entry and retention in the model. HLA-E genotype effects in D/R were tested separately by comparing 0, 1 or 2 copies of a particular HLA-E allele (i.e. HLA-E*01:03). Transplant pairs with HLA-E genotypes including other HLA-E alleles (i.e. other than HLA-E*01:01 or *01:03) were excluded from this analysis. The joint effect of D/R HLA-E genotype was also explored through a 4-level analysis of D/R HLA-E*01:03 homozygous vs other.Results: HLA-E*01:01 and *01:03 frequencies in both D/R as well as the rate of HLA-E discrepant transplant pairs (32.7%) were in accordance with previously reported data. Distribution of HLA-E matched vs HLA-E mismatched cases was balanced for all significant clinical predictors, while no significant interactions between the HLA-E matching status variable and the adjusted covariates were detected. HLA-E mismatch had no significant association with any of the outcome endpoints in both univariate and multivariate models (data not shown). With respect to the HLA-E genotype effect on outcome, 1827/1840 transplant pairs were considered for analysis, as 13 transplant pairs were excluded due to new or rare HLA-E genotypes. When tested separately, both D/R HLA-E*01:03/01:03 genotypes were associated with significantly lower LFS (HR=1.28, p=0.0027 and HR=1.31, p=0.0017 respectively). While the impact of HLA-E genotype on other outcome endpoints was not statistically significant, relapse rates were higher in cases with D/R HLA-E*01:03/01:03 genotype. Joint D/R HLA-E genotype analysis suggest that donor HLA-E*01:03/01:03 is driving this effect. The results of LFS and RI multivariate models are summarized in Table 2. No significant interactions were observed between the HLA-E genotype variable and the adjusted covariates in the multivariate models.Conclusion: This is the largest study investigating the effect of HLA-E polymorphism on HSCT outcome. We found no apparent effect of HLA-E mismatch on HSCT outcome in AML/ALL patients in complete remission, while donor HLA-E*01:03/01:03 genotype was associated with lower LFS, contrary to some previous reports. Differential efficiency of the two HLA allelic forms in inducing the CD94/NKG2A inhibitory pathway may account for their divergent effect on leukemia control, although this hypothesis is yet to be confirmed by functional assays. Our results suggest that avoidance of HLA-E*01:03/01:03 donors may improve the outcome of HSCT from 10/10 matched unrelated grafts.This project was funded by the Else Kröner-Fresenius-Foundation and the National Marrow Donor Program® (NMDP). [Display omitted] DisclosuresLee:Kadmon: Research Funding; Amgen: Consultancy, Research Funding; Mallinckrodt: Honoraria; Incyte: Consultancy; Pfizer: Consultancy; Onyx: Research Funding; Takeda: Research Funding. Fleischhauer:GENDX: Research Funding. Paczesny:Viracor IBT Laboratories: Patents & Royalties. Schrezenmeier:Alexion Pharmaceuticals, Inc.: Honoraria, Research Funding.

HLA-E regulatory and coding region variability and haplotypes in a Brazilian population sample

The HLA-E gene is characterized by low but wide expression on different tissues. HLA-E is considered a conserved gene, being one of the least polymorphic class I HLA genes. The HLA-E molecule interacts with Natural Killer cell receptors and T lymphocytes receptors, and might activate or inhibit immune responses depending on the peptide associated with HLA-E and with which receptors HLA-E interacts to. Variable sites within the HLA-E regulatory and coding segments may influence the gene function by modifying its expression pattern or encoded molecule, thus, influencing its interaction with receptors and the peptide. Here we propose an approach to evaluate the gene structure, haplotype pattern and the complete HLA-E variability, including regulatory (promoter and 3′UTR) and coding segments (with introns), by using massively parallel sequencing. We investigated the variability of 420 samples from a very admixed population such as Brazilians by using this approach. Considering a segment of about 7kb, 63 variable sites were detected, arranged into 75 extended haplotypes. We detected 37 different promoter sequences (but few frequent ones), 27 different coding sequences (15 representing new HLA-E alleles) and 12 haplotypes at the 3′UTR segment, two of them presenting a summed frequency of 90%. Despite the number of coding alleles, they encode mainly two different full-length molecules, known as E*01:01 and E*01:03, which corresponds to about 90% of all. In addition, differently from what has been previously observed for other non classical HLA genes, the relationship among the HLA-E promoter, coding and 3′UTR haplotypes is not straightforward because the same promoter and 3′UTR haplotypes were many times associated with different HLA-E coding haplotypes. This data reinforces the presence of only two main full-length HLA-E molecules encoded by the many HLA-E alleles detected in our population sample. In addition, this data does indicate that the distal HLA-E promoter is by far the most variable segment. Further analyses involving the binding of transcription factors and non-coding RNAs, as well as the HLA-E expression in different tissues, are necessary to evaluate whether these variable sites at regulatory segments (or even at the coding sequence) may influence the gene expression profile.

P087 A next generation sequencing based genotyping strategy for HLA-E

Aim In 2013, we implemented a short amplicon based Next Generation Sequencing (NGS) high-throughput workflow for HLA donor registry typing. Data analysis is performed by the in-house typing software neXtype. Currently, over 30,000 samples per week can be typed for 6 HLA loci, ABO, RhD, CCR5, and KIR with this cost-efficient workflow in our lab. As HLA-E has been reported to impact the outcome of stem cell transplantations in certain settings, we evaluated the feasibility of low-cost high-throughput HLA-E typing. On a clinically relevant antigen recognition domain (ARD) level, there are currently 9 groups of HLA-E alleles known (Fig. 1). Methods HLA-E is amplified by PCR with two primers spanning exons 2 and 3. This amplification product is subject to sequencing on Illumina MiSeq or HiSeq 2500 instruments without fragmentation. Thereby, information about the phasing between exon 2 and exon 3 is preserved, which is an important advantage of this method since it allows genotyping at ARD-level resolution. As shown in Fig. 1, alleles HLA-E∗01:01:02, 01:03:03, 01:04, 01:05, 01:07 and 01:08N can be resolved to the allele level. The remaining alleles fall into three distinguishable G-groups, namely HLA-E∗01:01:01G, 01:03:01G and 01:03:02G. Results The primer set was tested using 384 samples. All successfully sequenced samples corresponded to the G-groups HLA-E∗01:01:01G, 01:03:01G and 01:03:02G. Validation of the workflow against pre-typed samples is projected. Conclusions Sequencing HLA-E on allelic level has been show feasible in a cost efficient high throughput workflow. As the impact of HLA-E on the transplantation outcome becomes more settled, it could be considered for inclusion into the recruitment typing profile. Download high-res image (244KB) Download full-size image

A novel HLA-E allele, HLA-E*01:01:01:06, identified in a Chinese Leukemia patient.

A novel HLA-E allele, HLA-E*01:01:01:06, was identified in a Chinese leukemia patient.

New insights in HLA-E polymorphism by refined analysis of the full-length gene.

Human leukocyte antigen (HLA)-E is a non-classical HLA class I molecule that plays a role in both the innate and the adaptive immune response through interaction with receptors on natural killer- and T-cells. The HLA-E gene is characterized by limited polymorphism compared with the classical HLA loci on chromosome 6. At the start of this study, only 13 variable sites had been identified (IPD-IMGT/HLA Database v3.18.0). While most previous studies focused on polymorphism in exons 2 and 3 or specific gene regions, polymorphism in the other exons and introns could influence protein expression and function as well. Studies that investigate extended HLA-E polymorphism are therefore needed to better understand the functional relevance of HLA-E in health and disease. The aim of this study was to examine the variability of the full-length HLA-E gene region in individuals originating from different populations. A total of 7 new HLA-E alleles were identified using full-length HLA-E sequencing of 123 individuals from Asian, Dutch or Hunan Han origin. Furthermore, genome variation analysis of the third phase of the 1000 genomes database showed 107 new variable sites in 2504 individuals originating from 26 different populations. Our study demonstrates that the nucleotide variability of the HLA-E gene is much higher than previously known, albeit in only a limited number of individuals. Overall only 2 variants, HLA-E*01:01 and *01:03, are frequently present worldwide, suggesting that balancing selection is acting on HLA-E.