Classical Major Histocompatibility Complex Class Research Articles

Genetic Diversity and Sequence Conservation of Peptide-Binding Regions of MHC Class I Genes in Pig, Cattle, Chimpanzee, and Human.

Comparative analyses of MHC gene diversity and evolution across different species could offer valuable insights into the evolution of MHC genes. Intra- and inter-species sequence diversity and conservation of 12 classical major histocompatibility complex (MHC) class I genes from cattle, chimpanzees, pigs, and humans was analyzed using 20 representative allelic groups for each gene. The combined analysis of paralogous loci for each species revealed that intra-locus amino-acid sequence variations in the peptide-binding region (PBR) of MHC I genes did not differ significantly between species, ranging from 8.44% for SLA to 10.75% for BoLA class I genes. In contrast, intraspecies differences in the non-PBRs of these paralogous genes were more pronounced, varying from 4.59% for SLA to 16.89% for HLA. Interestingly, the Shannon diversity index and rate of nonsynonymous substitutions for PBR were significantly higher in SLA and BoLA than those in Patr and HLA. Analysis of peptide-binding pockets across all analyzed MHC class I genes of the four species indicated that pockets A and E showed the lowest and highest diversity, respectively. The estimated divergence times suggest that primate and artiodactyl MHC class I genes diverged 60.41 Mya, and BoLA and SLA genes diverged 35.34 Mya. These results offer new insights into the conservation and diversity of MHC class I genes in various mammalian species.

An Ancestral Major Histocompatibility Complex Organization in Cartilaginous Fish: Reconstructing MHC Origin and Evolution.

Cartilaginous fish (sharks, rays, and chimeras) comprise the oldest living jawed vertebrates with a mammalian-like adaptive immune system based on immunoglobulins (Ig), T-cell receptors (TCRs), and the major histocompatibility complex (MHC). Here, we show that the cartilaginous fish "adaptive MHC" is highly regimented and compact, containing (i) a classical MHC class Ia (MHC-Ia) region containing antigen processing (antigen peptide transporters and immunoproteasome) and presenting (MHC-Ia) genes, (ii) an MHC class II (MHC-II) region (with alpha and beta genes) with linkage to beta-2-microglobulin (β2m) and bromodomain-containing 2, (iii) nonclassical MHC class Ib (MHC-Ib) regions with 450 million-year-old lineages, and (iv) a complement C4 associated with the MHC-Ia region. No MHC-Ib genes were found outside of the elasmobranch MHC. Our data suggest that both MHC-I and MHC-II genes arose after the second round of whole-genome duplication (2R) on a human chromosome (huchr) 6 precursor. Further analysis of MHC paralogous regions across early branching taxa from all jawed vertebrate lineages revealed that Ig/TCR genes likely arose on a precursor of the huchr9/12/14 MHC paralog. The β2m gene is linked to the Ig/TCR genes in some vertebrates suggesting that it was present at 1R, perhaps as the donor of C1 domain to the primordial MHC gene. In sum, extant cartilaginous fish exhibit a conserved and prototypical MHC genomic organization with features found in various vertebrates, reflecting the ancestral arrangement for the jawed vertebrates.

Cryptic MHC-E epitope from influenza elicits a potent cytolytic T cell response.

The extent to which unconventional forms of antigen presentation drive T cell immunity is unknown. By convention, CD8 T cells recognize viral peptides, or epitopes, in association with classical major histocompatibility complex (MHC) class I, or MHC-Ia, but immune surveillance can, in some cases, be directed against peptides presented by nonclassical MHC-Ib, in particular the MHC-E proteins (Qa-1 in mice and HLA-E in humans); however, the overall importance of nonclassical responses in antiviral immunity remains unclear. Similarly uncertain is the importance of 'cryptic' viral epitopes, defined as those undetectable by conventional mapping techniques. Here we used an immunopeptidomic approach to search for unconventional epitopes that drive T cell responses in mice infected with influenza virus A/Puerto Rico/8/1934. We identified a nine amino acid epitope, termed M-SL9, that drives a co-immunodominant, cytolytic CD8 T cell response that is unconventional in two major ways: first, it is presented by Qa-1, and second, it has a cryptic origin, mapping to an unannotated alternative reading frame product of the influenza matrix gene segment. Presentation and immunogenicity of M-SL9 are dependent on the second AUG codon of the positive sense matrix RNA segment, suggesting translation initiation by leaky ribosomal scanning. During influenza virus A/Puerto Rico/8/1934 infection, M-SL9-specific T cells exhibit a low level of egress from the lungs and strong differentiation into tissue-resident memory cells. Importantly, we show that M-SL9/Qa-1-specific T cells can be strongly induced by messenger RNA vaccination and that they can mediate antigen-specific cytolysis in vivo. Our results demonstrate that noncanonical translation products can account for an important fraction of the T cell repertoire and add to a growing body of evidence that MHC-E-restricted T cells could have substantial therapeutic value.

Major histocompatibility complex class IIα and IIβ of pufferfish (Takifugu obscurus): Identification and functional characterization in response to bacterial challenge.

Classical major histocompatibility complex (MHC) class II molecules play an essential role in immune system. In this study, MHC IIα (Pf-MHC IIα) and MHC IIβ (Pf-MHC IIβ) homology genes from pufferfish (Takifugu obscurus) were cloned and their functional characterization in response to bacterial challenge was identified. The nucleotide sequences of the open reading frames (ORFs) of pufferfish Pf-MHC IIα and Pf-MHC IIβ were 708 bp and 750 bp, encoding 235 aa and 249 aa, respectively. The structure of Pf-MHC IIα or Pf-MHC IIβ contained a signal peptide, an α1/β1 domain, an α2/β2 domain, a transmembrane region and a cytoplasmic region. Multiple sequence alignment and phylogenetic analysis showed that Pf-MHC IIα and Pf-MHC IIβ molecules had the highest similarity with Fugu rubripes (Takifugu rubripes). Cellular localization analysis indicated that the distribution of Pf-MHC IIα and Pf-MHC IIβ was in the lymphocyte membrane and cytoplasm. qRT-PCR results showed that Pf-MHC IIα and Pf-MHC IIβ expressed relatively high in skin, gills and gut. In addition, after stimulation challenge in vitro (lipopolysaccharide, or polyinosinic: polycytidylic acid) and in vivo (A. hydrophila), the mRNA expressions of Pf-MHC IIα and Pf-MHC IIβ were significantly up-regulated in lymphocytes and in tissues of skin, gills, gut and head kidney. Moreover, Pf-MHC IIα or Pf-MHC IIβ neutralization reduced the ability of A. hydrophila to induce the expressions of lymphocyte cytokines (TNF-α, IL-1β and IL-10). Overall, it is speculated that Pf-MHC IIα and Pf-MHC IIβ may play an important role in the host response against A. hydrophila in pufferfish.

Crystal Structure of a Classical MHC Class I Molecule in Dogs; Comparison of DLA-88*0 and DLA-88*5 Category Molecules.

DLA-88 is a classical major histocompatibility complex (MHC) class I gene in dogs, and allelic DLA-88 molecules have been divided into two categories named "DLA-88*0" and "DLA-88*5." The defining difference between the two categories concerns an LQW motif in the α2 domain helical region of the DLA-88*5 molecules that includes the insertion of an extra amino acid compared to MHC class I consensus length. We here show that this motif has been exchanged by recombination between different DLA-88 evolutionary lineages. Previously, with pDLA-88*508:01, the structure of a molecule of the DLA-88*5 category was elucidated. The present study is the first to elucidate a structure, using X-ray crystallography, of the DLA-88*0 category, namely DLA-88*001:04 complexed with β2m and a nonamer peptide derived from canine distemper virus (CDV). The LQW motif that distinguishes DLA-88*5 from DLA-88*0 causes a shallower peptide binding groove (PBG) and a leucine exposed at the top of the α2 domain helix expected to affect T cell selection. Peptide ligand amino acid substitution and pMHC-I complex formation and stability analyses revealed that P2 and P3 are the major anchor residue positions for binding to DLA-88*001:04. We speculate that the distribution pattern of the LQW motif among canine classical MHC class I alleles represents a strategy to enhance allogeneic rejection by T cells of transmissible cancers such as canine transmissible venereal tumor (CTVT).

Reinforcement of cell-mediated immunity driven by tumor-associated Epstein-Barr virus (EBV)-specific T cells during targeted B-cell therapy with rituximab.

In immunocompromised patients, Epstein-Barr virus (EBV) infection or reactivation is associated with increased morbidity and mortality, including the development of B-cell lymphomas. The first-line treatment consists of reduction of immunosuppression and administration of rituximab (anti-CD20 antibody). Furthermore, the presence of EBV-specific T cells against latent EBV proteins is crucial for the control of EBV-associated diseases. Therefore, in addition to effective treatment strategies, appropriate monitoring of T cells of high-risk patients is of great importance for improving clinical outcome. In this study, we hypothesized that rituximab-mediated lysis of malignant EBV-infected B cells leads to the release and presentation of EBV-associated antigens and results in an augmentation of EBV-specific effector memory T-cell responses. EBV-infected B lymphoblastoid cell lines (B-LCLs) were used as a model for EBV-associated lymphomas, which are capable of expressing latency stage II and III EBV proteins present in all known EBV-positive malignant cells. Rituximab was administered to obtain cell lysates containing EBV antigens (ACEBV). Efficiency of cross-presentation of EBV-antigen by B-LCLs compared to cross-presentation by professional antigen presenting cells (APCs) such as dendritic cells (DCs) and B cells was investigated by in vitro T-cell immunoassays. Deep T-cell profiling of the tumor-reactive EBV-specific T cells in terms of activation, exhaustion, target cell killing, and cytokine profile was performed, assessing the expression of T-cell differentiation and activation markers as well as regulatory and cytotoxic molecules by interferon-γ (IFN-γ) EliSpot assay, multicolor flow cytometry, and multiplex analyses. By inhibiting parts of the cross-presentation pathway, B-LCLs were shown to cross-present obtained exogenous ACEBV-derived antigens mainly through major histocompatibility complex (MHC) class I molecules. This mechanism is comparable to that for DCs and B cells and resulted in a strong EBV-specific CD8+ cytotoxic T-cell response. Stimulation with ACEBV-loaded APCs also led to the activation of CD4+ T helper cells, suggesting that longer peptide fragments are processed via the classical MHC class II pathway. In addition, B-LCLs were also found to be able to take up exogenous antigens from surrounding cells by endocytosis leading to induction of EBV-specific T-cell responses although to a much lesser extent than cross-presentation of ACEBV-derived antigens. Increased expression of activation markers CD25, CD71 and CD137 were detected on EBV-specific T cells stimulated with ACEBV-loaded APCs, which showed high proliferative and cytotoxic capacity as indicated by enhanced EBV-specific frequencies and increased secretion levels of cytotoxic effector molecules (e.g. IFN-γ, granzyme B, perforin, and granulysin). Expression of the regulatory proteins PD-1 and Tim-3 was induced but had no negative impact on effector T-cell functions. In this study, we showed for the first time that rituximab-mediated lysis of EBV-infected tumor cells can efficiently boost EBV-specific endogenous effector memory T-cell responses through cross-presentation of EBV-derived antigens. This promotes the restoration of antiviral cellular immunity and presents an efficient mechanism to improve the treatment of CD20+ EBV-associated malignancies. This effect is also conceivable for other therapeutic antibodies or even for therapeutically applied unmodified or genetically modified T cells, which lead to the release of tumor antigens after specific cell lysis.

Placental macrophage responses to viral and bacterial ligands and the influence of fetal sex.

Bacterial and viral infections of the placenta are associated with inflammation and adverse pregnancy outcomes. Hofbauer cells (HBCs) are fetal-origin macrophages in the placenta, proposed to protect the fetus from vertical pathogen transmission. We performed quantitative proteomics on term HBCs under resting conditions and following exposure to bacterial and viral pathogen-associated molecular patterns (PAMPs), and investigated the contribution of fetal sex. Resting HBCs expressed proteins pertinent to macrophage function, including chemokines, cytokines, Toll-like receptors, and major histocompatibility complex class I and II molecules. HBCs mounted divergent responses to bacterial versus viral PAMPs but exhibited protein expression changes suggestive of a more pro-inflammatoryphenotype. A comparison between male and female HBCs showed that the latter mounted a stronger and wider response. Here, we provide a comprehensive understanding of the sex-dependent responses of placental macrophages to infectious triggers, which were primarily associated with lipid metabolism in males and cytoskeleton organization in females.

Repair of Retinal Degeneration by Human Amniotic Epithelial Stem Cell-Derived Photoreceptor-like Cells.

The loss of photoreceptors is a major event of retinal degeneration that accounts for most cases of untreatable blindness globally. To date, there are no efficient therapeutic approaches to treat this condition. In the present study, we aimed to investigate whether human amniotic epithelial stem cells (hAESCs) could serve as a novel seed cell source of photoreceptors for therapy. Here, a two–step treatment with combined Wnt, Nodal, and BMP inhibitors, followed by another cocktail of retinoic acid, taurine, and noggin induced photoreceptor–like cell differentiation of hAESCs. The differentiated cells demonstrated the morphology and signature marker expression of native photoreceptor cells and, intriguingly, bore very low levels of major histocompatibility complex (MHC) class II molecules and a high level of non–classical MHC class I molecule HLA–G. Importantly, subretinal transplantation of the hAESCs–derived PR–like cells leads to partial restoration of visual function and retinal structure in Royal College of Surgeon (RCS) rats, the classic preclinical model of retinal degeneration. Together, our results reveal hAESCs as a potential source of functional photoreceptor cells; the hAESCs–derived photoreceptor–like cells could be a promising cell–replacement candidate for therapy of retinal degeneration diseases.

Intra-specific copy number variation of MHC class II genes in the Siamese fighting fish.

Duplicates of genes for major histocompatibility complex (MHC) molecules can be subjected to selection independently and vary markedly in their evolutionary rates, sequence polymorphism, and functional roles. Therefore, without a thorough understanding of their copy number variation (CNV) in the genome, the MHC-dependent fitness consequences within a species could be misinterpreted. Studying the intra-specific CNV of this highly polymorphic gene, however, has long been hindered by the difficulties in assigning alleles to loci and the lack of high-quality genomic data. Here, using the high-quality genome of the Siamese fighting fish (Betta splendens), a model for mate choice studies, and the whole-genome sequencing (WGS) data of 17 Betta species, we achieved locus-specific amplification of their three classical MHC class II genes - DAB1, DAB2, and DAB3. By performing quantitative PCR and depth-of-coverage analysis using the WGS data, we revealed intra-specific CNV at the DAB3 locus. We identified individuals that had two allelic copies (i.e., heterozygous or homozygous) or one allele (i.e., hemizygous) and individuals without this gene. The CNV was due to the deletion of a 20-kb-long genomic region harboring both the DAA3 and DAB3 genes. We further showed that the three DAB genes were under different modes of selection, which also applies to their corresponding DAA genes that share similar pattern of polymorphism. Our study demonstrates a combined approach to study CNV within a species, which is crucial for the understanding of multigene family evolution and the fitness consequences of CNV.

Functional analysis of a novel MHC-Iα genotype in orange-spotted grouper: Effects on Singapore grouper iridovirus (SGIV) replication and apoptosis

The classical major histocompatibility complex class I (MHC-Ⅰ) molecule plays a key role in vertebrate immune response for its important functions in antigen presentation and immune regulation. MHC pathway is closely related to many diseases involving autoimmunity, antigen intrusion and inflammation. However, rare literatures about the effect of MHC-I on fish cells apoptosis were reported. In this study, a novel type of MHC-Ⅰα genotype from orange-spotted grouper (named EcMHC-ⅠA*01) were cloned and characterized. It shared a 77% identity to its Epinephelus coioides MHC-Iα homology that has been uploaded to NCBI (ACZ97571.1). Molecular characterization analysis showed that EcMHC-ⅠA*01 encodes a 357-amino-acid protein, containing a signal peptide，α1，α2，α3, Cytoplasmic (Cyt) and Transmembrane (TM) domains. Tissue expression pattern showed that EcMHC-ⅠA*01 was extensively distributed in twelve selected tissues, with higher expression in the gill, intestine and skin. The expression of EcMHC-ⅠA*01 in grouper liver and spleen tissues were significantly induced by different stimuli (Zymosan A, LPS, Ploy I:C, RGNNV and SGIV). Comparing with the EcMHC-ⅠA*01 expression levels induced by Zymosan A, Ploy I:C and RGNNV, the effects induced by SGIV and LPS were more significant. Subcellular localization analysis showed that EcMHC-ⅠA*01 localizes throughout the cytoplasm appeared both diffuse and focal intracellular expression pattern. Overexpression of EcMHC-ⅠA*01 inhibited the CPE progression, the mRNA expression of the SGIV related genes (MCP, LITAF, ICP-18 and VP19) and the protein expression of MCP. Meanwhile, qRT-PCR result showed that EcMHC-ⅠA*01 overexpression upregulated the expression of interferon signaling molecules (IFN-γ, ISG56, MDA5 and MXI) and inflammatory cytokines (IL-1β, IL-6, TNF-α and TRAF6). In addition, our results showed that overexpression of EcMHC-ⅠA*01 promoted the apoptosis of normal fathead minnow (FHM) cells as well as the apoptosis of FHM cells induced by SGIV. However, there was no significant change in the activity of caspase 3 between control group and EcMHC-ⅠA*01 overexpression group, suggesting that EcMHC-ⅠA*01-induced apoptosis may not depend on the caspase 3 pathway. Taken together, these data in our study provide new insights into the role of MHC-I in antiviral immune response and apoptosis in fish.

The dominantly expressed class II molecule from a resistant MHC haplotype presents only a few Marek's disease virus peptides by using an unprecedented binding motif.

Viral diseases pose major threats to humans and other animals, including the billions of chickens that are an important food source as well as a public health concern due to zoonotic pathogens. Unlike humans and other typical mammals, the major histocompatibility complex (MHC) of chickens can confer decisive resistance or susceptibility to many viral diseases. An iconic example is Marek's disease, caused by an oncogenic herpesvirus with over 100 genes. Classical MHC class I and class II molecules present antigenic peptides to T lymphocytes, and it has been hard to understand how such MHC molecules could be involved in susceptibility to Marek's disease, given the potential number of peptides from over 100 genes. We used a new in vitro infection system and immunopeptidomics to determine peptide motifs for the 2 class II molecules expressed by the MHC haplotype B2, which is known to confer resistance to Marek's disease. Surprisingly, we found that the vast majority of viral peptide epitopes presented by chicken class II molecules arise from only 4 viral genes, nearly all having the peptide motif for BL2*02, the dominantly expressed class II molecule in chickens. We expressed BL2*02 linked to several Marek's disease virus (MDV) peptides and determined one X-ray crystal structure, showing how a single small amino acid in the binding site causes a crinkle in the peptide, leading to a core binding peptide of 10 amino acids, compared to the 9 amino acids in all other reported class II molecules. The limited number of potential T cell epitopes from such a complex virus can explain the differential MHC-determined resistance to MDV, but raises questions of mechanism and opportunities for vaccine targets in this important food species, as well as providing a basis for understanding class II molecules in other species including humans.

TAP dysfunction in dendritic cells enables noncanonical cross-presentation for T cell priming.

Classic MHC-I presentation relies on shuttling cytosolic peptides into the endoplasmic reticulum (ER) by the transporter associated with antigen processing (TAP). Viruses disable TAP to block MHC-I presentation and evade cytotoxic CD8+ T cells. Priming CD8+ T cells against these viruses is thought to rely solely on cross-presentation by uninfected TAP-functional dendritic cells (DCs). We found that protective CD8+ T cells could be mobilized during viral infection even when TAP was absent in all hematopoietic cells. TAP blockade depleted the endosomal recycling compartment (ERC) of MHC-I and as such impaired Toll-like receptor-regulated cross-presentation. Instead, MHC-I accumulated in ER-Golgi intermediate compartments (ERGIC), sequestered away from Toll-like receptor control, and coopted ER-SNARE Sec22b-mediated vesicular traffic to intersect with internalized antigen and rescue cross-presentation. Thus, when classic MHC-I presentation and ERC-dependent cross-presentation are impaired in DCs, cell-autonomous non-canonical cross-presentation relying on ERGIC-derived MHC-I counters TAP dysfunction to nevertheless mediate CD8+ T cell priming.

MHC class I evolution; from Northern pike to salmonids

BackgroundSalmonids are of major importance both as farmed and wild animals. With the changing environment comes changes in pathogenic pressures so understanding the immune system of all salmonid species is of essence. Major histocompatibility complex (MHC) genes are key players in the adaptive immune system signalling infection to responding T-cells populations. Classical MHC class I (MHCI) genes, defined by high polymorphism, broad expression patterns and peptide binding ability, have a key role in inducing immunity. In salmonids, the fourth whole genome duplication that occurred 94 million years ago has provided salmonids with duplicate MHCI regions, while Northern Pike, a basal sister clade to salmonids, represent a species which has not experienced this whole genome duplication.ResultsComparing the gene organization and evolution of MHC class I gene sequences in Northern pike versus salmonids displays a complex picture of how many of these genes evolved. Regional salmonid Ia and Ib Z lineage gene duplicates are not orthologs to the Northern pike Z lineage sequences. Instead, salmonids have experienced unique gene duplications in both duplicate regions as well as in the Salmo and Oncorhynchus branch. Species-specific gene duplications are even more pronounced for some L lineage genes.ConclusionsAlthough both Northern pike as well as salmonids have expanded their U and Z lineage genes, these gene duplications occurred separately in pike and in salmonids. However, the similarity between these duplications suggest the transposable machinery was present in a common ancestor. The salmonid MHCIa and MHCIb regions were formed during the 94 MYA since the split from pike and before the Oncorhynchus and Salmo branch separated. As seen in tetrapods, the non-classical U lineage genes are diversified duplicates of their classical counterpart. One MHCI lineage, the L lineage, experienced massive species-specific gene duplications after Oncorhynchus and Salmo split approximately 25 MYA. Based on what we currently know about L lineage genes, this large variation in number of L lineage genes also signals a large functional diversity in salmonids.

Human Tissue-Resident Memory T Cells in the Maternal–Fetal Interface. Lost Soldiers or Special Forces?

The immune system plays a critical role during pregnancy, but the specific mechanisms and immune cell function needed to support pregnancy remain incompletely understood. Despite decades of research efforts, it is still unclear how the immune system maintains tolerance of fetal-derived tissues, which include most cells of the placenta and of course the fetus itself, without forfeiting the ability to protect against harmful infections. T cells recognize antigen in the context of major histocompatibility complex (MHC) encoded proteins, but classical MHC class I and II expression are diminished in fetal-derived cells. Can T cells present at the maternal–fetal interface (MFI) protect these cells from infection? Here we review what is known in regard to tissue-resident memory T (Trm) cells at the MFI. We mainly focus on how Trm cells can contribute to protection in the context of the unique features of the MFI, such as limited MHC expression as well as the temporary nature of the MFI, that are not found in other tissues.

Crystal structure of the ternary complex of TCR, MHC class I and lipopeptides.

The covalent conjugation of a 14-carbon fatty acid (myristic acid) to the N-terminal Gly residue, termed N-myristoylation, occurs in some viral proteins to dictate their pathological function. This protein lipidation reaction, however, is monitored by host cytotoxic T lymphocytes that are capable of recognizing N-terminal lipopeptide fragments in the context of major histocompatibility complex (MHC) class I molecules. In a rhesus model of human AIDS, for example, the classical MHC class I allomorph, Mamu-B*05104, was shown to bind SIV Nef-derived 4-mer lipopeptides (myristic acid-Gly-Gly-Ala-Ile; C14nef4) and present them to the CD8+ T-cell line, SN45. These lipopeptides accommodated in MHC class I molecules expose much shorter peptide chains than conventional MHC class I-presented 8-10-mer peptides, and the molecular mechanisms by which αβ T-cell receptors (TCRs) recognize lipopeptides currently remain unclear. An X-ray crystallographic analysis of the SN45 TCR α and β heterodimer in a form that was co-crystallized with the C14nef4-bound Mamu-B*05104 complex indicated that the amide group of the N-myristoylated glycine residue offered a primary T-cell epitope by establishing a sole hydrogen bond between its nitrogen atom and the side chain of Glu at position 101 of CDR3β. Accordingly, the Glu to Ala mutation at this position resulted in the loss of lipopeptide recognition. On the other hand, TCRs were positioned remotely from the peptide portion of C14nef4, and strong interactions were not observed. Thus, these observations provide novel structural insights into lipopeptide recognition by TCRs, which contrast sharply with the general molecular principle of peptide recognition.

Targeting the coding sequence: opposing roles in regulating classical and non-classical MHC class I molecules by miR-16 and miR-744.

BackgroundTo control gene expression, microRNAs (miRNAs) are of key importance and their deregulation is associated with the development and progression of various cancer types. In this context, a discordant messenger RNA/protein expression pointing to extensive post-transcriptional regulation of major histocompatibility complex (MHC) class I molecules was already shown. However, only a very limited number of miRNAs targeting these molecules have yet been identified. Despite an increasing evidence of coding sequence (CDS)-located miRNA binding sites, there exists so far, no detailed study of the interaction of miRNAs with the CDS of MHC class I molecules.MethodsUsing an MS2-tethering approach in combination with small RNA sequencing, a number of putative miRNAs binding to the CDS of human leukocyte antigen (HLA)-G were identified. These candidate miRNAs were extensively screened for their effects in the HLA-G-positive JEG3 cell line. Due to the high sequence similarity between HLA-G and classical MHC class I molecules, the impact of HLA-G candidate miRNAs on HLA class I surface expression was also analyzed. The Cancer Genome Atlas data were used to correlate candidate miRNAs and HLA class I gene expression.ResultsTransfection of candidate miRNAs revealed that miR-744 significantly downregulates HLA-G protein levels. In contrast, overexpression of the candidate miRNAs miR-15, miR-16, and miR-424 sharing the same seed sequence resulted in an unexpected upregulation of HLA-G. Comparable results were obtained for classical MHC class I members after transfection of miRNA mimics into HEK293T cells. Analyses of The Cancer Genome Atlas data sets for miRNA and MHC class I expression further validated the results.ConclusionsOur data expand the knowledge about MHC class I regulation and showed for the first time an miRNA-dependent control of MHC class I antigens mediated by the CDS. CDS-located miRNA binding sites could improve the general use of miRNA-based therapeutic approaches as these sites are highly independent of structural variations (e.g. mutations) in the gene body. Surprisingly, miR-16 family members promoted MHC class I expression potentially in a gene activation-like mechanism.

Crystal structures of lysophospholipid-bound MHC class I molecules

Newly synthesized major histocompatibility complex (MHC) class I proteins are stabilized in the endoplasmic reticulum (ER) by binding 8-10-mer-long self-peptide antigens that are provided by transporter associated with antigen processing (TAP). These MHC class I:peptide complexes then exit the ER and reach the plasma membrane, serving to sustain the steady-state MHC class I expression on the cell surface. A novel subset of MHC class I molecules that preferentially bind lipid-containing ligands rather than conventional peptides was recently identified. The primate classical MHC class I allomorphs, Mamu-B*098 and Mamu-B*05104, are capable of binding the N-myristoylated 5-mer (C14-Gly-Gly-Ala-Ile-Ser) or 4-mer (C14-Gly-Gly-Ala-Ile) lipopeptides derived from the N-myristoylated SIV Nef protein, respectively, and of activating lipopeptide antigen-specific cytotoxic T lymphocytes. We herein demonstrate that Mamu-B*098 samples lysophosphatidylethanolamine and lysophosphatidylcholine containing up to a C20 fatty acid in the ER. The X-ray crystal structures of Mamu-B*098 and Mamu-B*05104 complexed with lysophospholipids at high resolution revealed that the B and D pockets in the antigen-binding grooves of these MHC class I molecules accommodate these lipids through a monoacylglycerol moiety. Consistent with the capacity to bind cellular lipid ligands, these two MHC class I molecules did not require TAP function for cell-surface expression. Collectively, these results indicate that peptide- and lipopeptide-presenting MHC class I subsets use distinct sources of endogenous ligands.

FMF Is Associated With a Wide Spectrum of MHC Class I- and Allied SpA Disorders but Not With Classical MHC Class II-Associated Autoimmune Disease: Insights From a Large Cohort Study.

Objectives: To test the hypothesis that familial Mediterranean fever (FMF)-associated autoinflammation may exaggerate the tendency toward adaptive immunopathology or spondyloarthritis (SpA)-associated disorders including major histocompatibility complex (MHC) class I associated disorders but not classical MHC class II-associated disorders that exhibit transplacental autoimmunity including myasthenia gravis and pemphigus.Methods: Seven thousand seven hundred forty-seven FMF patients and 10,080 age- and sex-matched controls in the Clalit Health Services medical database were identified and compared in terms of prevalence of SpA-associated disorders. We also evaluated four classical and strong MHC class II-associated disorders, namely, pemphigus vulgaris, myasthenia gravis, sarcoidosis, and pernicious anemia, to ascertain whether such associations with SpA-spectrum disease were specific or merely reflected the non-specific consequences of innate immune system activation on driving divergent types of immunity. The diagnosis of FMF was based on the medical records and not genetically proven.Results: FMF showed a strong association with MHC class I-related diseases: odds ratio (OR) of 28.58 [95% confidence interval (95% CI), 6.93–117.87; p < 0.0001] for Behçet's disease, OR of 10.33 (95% CI, 4.09–26.09; p < 0.0001) for ankylosing spondylitis, and OR of 1.67 (95% CI, 1.19–2.33; p = 0.0029) for psoriasis. For weakly MHC class I-linked diseases, an OR of 3.76 (95% CI, 2.48–5.69; p < 0.0001) for Crohn's disease and OR of 2.64 (95% CI, 1.52–4.56; p = 0.0005) for ulcerative colitis were found. No association was found between FMF and the four MHC class II-associated autoimmune disorders.Conclusion: FMF patients are associated with increased risk of SpA-related disease diagnosis including MHC-I-opathies but not MHC-II-associated autoimmune diseases, suggesting that tissue-specific dysregulation of innate immunity share between FMF and SpA spectrum disorders may drive adaptive immune MHC class I-associated conditions.

Microbial Danger Signals Control Transcriptional Induction of Distinct MHC Class I L Lineage Genes in Atlantic Salmon.

Antigen processing and presentation by major histocompatibility complex (MHC) molecules is a cornerstone in vertebrate immunity. Like mammals, teleosts possess both classical MHC class I and multiple families of divergent MHC class I genes. However, while certain mammalian MHC class I-like molecules have proven to be integral in immune regulation against a broad array of pathogens, the biological relevance of the different MHC class I lineages in fish remains elusive. This work focuses on MHC class I L lineage genes and reveals unique regulatory patterns of six genes (Sasa-lia, Sasa-lda, Sasa-lca, Sasa-lga, Sasa-lha, and Sasa-lfa) in antimicrobial immunity of Atlantic salmon (Salmo salar L.). Using two separate in vivo challenge models with different kinetics and immune pathologies combined with in vitro stimulation using viral and bacterial TLR ligands, we show that de novo synthesis of different L lineage genes is distinctly regulated in response to various microbial stimuli. Prior to the onset of classical MHC class I gene expression, lia was rapidly and systemically induced in vivo by the single-stranded (ss) RNA virus salmonid alpha virus 3 (SAV3) but not in response to the intracellular bacterium Piscirickettsia salmonis. In contrast, lga expression was upregulated in response to both viral and bacterial stimuli. A role for distinct MHC class I L-lineage genes in anti-microbial immunity in salmon was further substantiated by a marked upregulation of lia and lga gene expression in response to type I IFNa stimulation in vitro. Comparably, lha showed no transcriptional induction in response to IFNa stimulation but was strongly induced in response to a variety of viral and bacterial TLR ligands. In sharp contrast, lda showed no response to viral or bacterial challenge. Similarly, induction of lca, which is predominantly expressed in primary and secondary lymphoid tissues, was marginal with the exception of a strong and transient upregulation in pancreas following SAV3 challenge Together, these findings suggest that certain Atlantic salmon MHC class I L lineage genes play important and divergent roles in early anti-microbial response and that their regulation, in response to different activation signals, represents a system for selectively promoting the expression of distinct non-classical MHC class I genes in response to different types of immune challenges.

Major Histocompatibility Complex (MHC) Genes and Disease Resistance in Fish.

Fascinating about classical major histocompatibility complex (MHC) molecules is their polymorphism. The present study is a review and discussion of the fish MHC situation. The basic pattern of MHC variation in fish is similar to mammals, with MHC class I versus class II, and polymorphic classical versus nonpolymorphic nonclassical. However, in many or all teleost fishes, important differences with mammalian or human MHC were observed: (1) The allelic/haplotype diversification levels of classical MHC class I tend to be much higher than in mammals and involve structural positions within but also outside the peptide binding groove; (2) Teleost fish classical MHC class I and class II loci are not linked. The present article summarizes previous studies that performed quantitative trait loci (QTL) analysis for mapping differences in teleost fish disease resistance, and discusses them from MHC point of view. Overall, those QTL studies suggest the possible importance of genomic regions including classical MHC class II and nonclassical MHC class I genes, whereas similar observations were not made for the genomic regions with the highly diversified classical MHC class I alleles. It must be concluded that despite decades of knowing MHC polymorphism in jawed vertebrate species including fish, firm conclusions (as opposed to appealing hypotheses) on the reasons for MHC polymorphism cannot be made, and that the types of polymorphism observed in fish may not be explained by disease-resistance models alone.