Nonclassical MHC Class Research Articles

Molecular and Structural Characterization of MHC Class II \u03b2 Genes Reveals High Diversity in the Cold-Adapted Icefish Chionodraco hamatus

This study reports the presence of two distinct MHC class II β genes in the Antarctic icefish Chionodraco hamatus, belonging to the classical (ChhaDAB) and nonclassical (ChhaDBB) evolutionary lineages. By the application of targeted sequencing approach, a remarkable molecular diversity in the exon 2 sequence of the highly expressed gene ChhaDAB has been observed, resulting in an estimate of 92 different variants translated in 87 different peptides from 54 analysed icefish individuals. A highly conservative estimate, based on a 95% sequence identity threshold clustering, translate this variability in 41 different peptide clusters belonging to four different clades and showing the signature of different kinds of selection. In stark contrast, the poorly expressed ChhaDBB gene displayed a very low level of molecular diversity within exon 2, in agreement with expectations for a nonclassical MHC class II β gene.

Effect of calorific intake on proteomic composition of colostrum in dairy cows

The amount of concentrated feed supplied to a dairy cow affects milk yield. However, there is no evidence of a relationship between the colostrum proteomic composition and energy intake. We supplied 30 heifers (4–24 months old, two groups of 15 heifers each) with either a normal diet and high-energy diet to investigate the correlation between energy intake and colostrum protein composition. Colostrum milk proteins were analysed on the day of calving and on the third day following calving using two-dimensional gel electrophoresis (2-DE) and peptide mass fingerprinting (PMF). Five proteins were identified as differentially expressed between the two feeding groups in the colostrum on the day of calving. The levels of αS2-casein precursor and β-casein was higher in the colostrum from the high-energy diet group (HEG), whereas the levels of IgG3 heavy chain constant region, non-classical MHC class I antigen isoform X2, and β-casein A2 variant were higher in the normal-diet group (NEG) colostrum. Twelve differential proteins were identified on the third day: β-lactoglobulin, αS2-casein, zinc-α2-glycoprotein, lactoferrin, fibrinogen gamma-B chain isoform X1, non-classical MHC class I antigen isoform X2, complement C3, gelsolin isoform A precursor, vitamin D-binding protein isoform X1, immunoglobulin gamma 1 heavy chain constant region, IgG3 heavy chain constant region and polymeric immunoglobulin receptor. All were present at higher levels in the normal-diet group colostrum than in the high-energy diet group colostrum, although the milk yield from mature cows was lower in the normal-diet group. In conclusion, a high-energy diet can enhance milk production; however, the levels of immune-related factors are higher in the colostrum of cows fed a normal diet.

Diversification of Bw4 Specificity and Recognition of a Nonclassical MHC Class I Molecule Implicated in Maternal-Fetal Tolerance by Killer Cell Ig-like Receptors of the Rhesus Macaque.

The rhesus macaque is an important animal model for AIDS and other infectious diseases; however, studies to address NK cell function in this species have been limited by the lack of defined ligands for killer cell Ig-like receptors (KIRs). To identify ligands for rhesus macaque KIRs, we adopted a novel approach based on a pair of stable cell lines. NFAT-responsive luciferase reporter cell lines expressing the extracellular domains of macaque KIRs fused to the transmembrane and cytoplasmic domains of CD28 and CD3ζ were incubated with target cells expressing individual MHC class I molecules, and ligand recognition was detected by the MHC class I-dependent upregulation of luciferase. Using this approach, we found that Mamu-KIR3DL01, -KIR3DL06, -KIR3DL08, and -KIR3DSw08 all recognize Mamu-Bw4 molecules but with differing allotype specificity. In contrast, Mamu-KIR3DL05 recognizes Mamu-A and Mamu-A-related molecules, including Mamu-A1*002 and -A3*13, Mamu-B*036, the product of a recombinant Mamu-B allele with α1 and α2 domain sequences derived from a MHC-A gene, and Mamu-AG*01, a nonclassical molecule expressed on placental trophoblasts that originated from an ancestral duplication of a MHC-A gene. These results reveal an expansion of the lineage II KIRs in macaques that recognize Bw4 ligands and identify a nonclassical molecule implicated in placental development and pregnancy as a ligand for Mamu-KIR3DL05. In addition to offering new insights into KIR-MHC class I coevolution, these findings provide an important foundation for investigating the role of NK cells in the rhesus macaque as an animal model for infectious diseases and reproductive biology.

Structure of MHC class I-like MILL2 reveals heparan-sulfate binding and interdomain flexibility

The MILL family, composed of MILL1 and MILL2, is a group of nonclassical MHC class I molecules that occur in some orders of mammals. It has been reported that mouse MILL2 is involved in wound healing; however, the molecular mechanisms remain unknown. Here, we determine the crystal structure of MILL2 at 2.15 Å resolution, revealing an organization similar to classical MHC class I. However, the α1-α2 domains are not tightly fixed on the α3-β2m domains, indicating unusual interdomain flexibility. The groove between the two helices in the α1-α2 domains is too narrow to permit ligand binding. Notably, an unusual basic patch on the α3 domain is involved in the binding to heparan sulfate which is essential for MILL2 interactions with fibroblasts. These findings suggest that MILL2 has a unique structural architecture and physiological role, with binding to heparan sulfate proteoglycans on fibroblasts possibly regulating cellular recruitment in biological events.

HLA-F on HLA-Null 721.221 Cells Activates Primary NK Cells Expressing the Activating Killer Ig-like Receptor KIR3DS1.

NK cells elicit important responses against transformed and virally infected cells. Carriage of the gene encoding the activating killer Ig-like receptor KIR3DS1 is associated with slower time to AIDS and protection from HIV infection. Recently, open conformers of the nonclassical MHC class Ib Ag HLA-F were identified as KIR3DS1 ligands. In this study, we investigated whether the interaction of KIR3DS1 on primary NK cells with HLA-F on the HLA-null cell line 721.221 (221) stimulated KIR3DS1+ NK cells. We used a panel of Abs to detect KIR3DS1+CD56dim NK cells that coexpressed the inhibitory NK cell receptors KIR2DL1/L2/L3, 3DL2, NKG2A, and ILT2; the activating NK cell receptors KIR2DS1/S2/S3/S5; and CCL4, IFN-γ, and CD107a functions. We showed that both untreated and acid-pulsed 221 cells induced a similar frequency of KIR3DS1+ cells to secrete CCL4/IFN-γ and express CD107a with a similar intensity. A higher percentage of KIR3DS1+ than KIR3DS1- NK cells responded to 221 cells when either inclusive or exclusive (i.e., coexpressing none of the other inhibitory NK cell receptors and activating NK cell receptors detected by the Ab panel) gating strategies were employed to identify these NK cell populations. Blocking the interaction of HLA-F on 221 cells with KIR3DS1-Fc chimeric protein or anti-HLA-F Abs on exclusively gated KIR3DS1+ cells reduced the frequency of functional cells compared with that of unblocked conditions for stimulated KIR3DS1+ NK cells. Thus, ligation of KIR3DS1 activates primary NK cells for several antiviral functions.

HLA-G: Facts and Fictions

Human leukocyte antigen (HLA)-G is a nonclassical MHC class I molecule with modulatory effects on NK and T cells. Unlike classical HLA class I molecules, HLA-G has seven isoforms, three of which are soluble. Soluble HLA-G molecules are reportedly able to transduce negative signals to immune cells after interacting with their corresponding receptors. The expression of these molecules plays significant roles in maternal tolerance against semi-allogenic fetuses. Overexpression of HLA-G in tumors and increased serum levels of soluble HLA-G have been reported in different malignancies, and these changes may be involved in tumoral immune evasion and cancer progression. To improve immune responses against tumor cells, the downmodulation of HLA-G by siRNA or blocking monoclonal antibodies can be helpful in cancer immunotherapy. Additionally, HLA-G can be considered a potential biomarker for the diagnosis and/or prognosis of certain cancers. Although polymorphism of the HLA-G gene-coding region is more limited than in classical HLA class I, some genetic variations in regulatory regions of the gene control the expression level of this molecule. Furthermore, epigenetic factors such as infections may affect the expression of HLA-G in infection-related cancers.

Self-reactive T cells restricted by non-classical MHC Ib are associated with metabolic disease

Abstract The normal peptide repertoire presented by classical and non-classical MHC class I molecules is regulated by ERAAP, the ER aminopeptidase associated with antigen processing. Loss of ERAAP’s peptide trimming function in cells causes dramatic changes in the peptide repertoire. The changes in the peptide repertoire enhance the immunogenicity of ERAAP-deficient cells and elicit potent immune responses in otherwise syngeneic wild-type mice. Normal ERAAP function is monitored by an unusual subset of semi-invariant CD8+ T cells that recognize the QFL ligand. The QFL ligand consists of a peptide encoded by the highly conserved FAM49a/b genes that is presented by Qa-1b, a non-classical MHC Ib molecule only on surface of ERAAP-deficient cells. We show that these QFL-specific CD8+ T cells (QFL-T cells) bear unique and semi-invariant αβ TCRs. Genetic manipulation of the expression of the self-QFL ligand and functional characteristics of QFL-T cells shows that in addition to monitoring ERAAP function, QFL-T cells may also regulate metabolic activity.

Non-classical Immunity Controls Microbiota Impact on Skin Immunity and Tissue Repair

Mammalian barrier surfaces are constitutively colonized by numerous microorganisms. We explored how the microbiota was sensed by the immune system and the defining properties of such responses. Here, we show that a skin commensal can induce Tcell responses in a manner that is restricted to non-classical MHC class I molecules. These responses are uncoupled from inflammation and highly distinct from pathogen-induced cells. Commensal-specific Tcells express a defined gene signature that is characterized by expression of effector genestogether with immunoregulatory and tissue-repairsignatures. As such, non-classical MHCI-restricted commensal-specific immune responses not only promoted protection to pathogens, but also accelerated skin wound closure. Thus, the microbiota can induce a highly physiological and pleiotropic form of adaptive immunity that couples antimicrobial function with tissue repair. Our work also reveals that non-classical MHC class I molecules, an evolutionarily ancient arm of the immune system, can promote homeostatic immunity to the microbiota.

The Role of MHC-E in T Cell Immunity Is Conserved among Humans, Rhesus Macaques, and Cynomolgus Macaques.

MHC-E is a highly conserved nonclassical MHC class Ib molecule that predominantly binds and presents MHC class Ia leader sequence-derived peptides for NK cell regulation. However, MHC-E also binds pathogen-derived peptide Ags for presentation to CD8+ T cells. Given this role in adaptive immunity and its highly monomorphic nature in the human population, HLA-E is an attractive target for novel vaccine and immunotherapeutic modalities. Development of HLA-E-targeted therapies will require a physiologically relevant animal model that recapitulates HLA-E-restricted T cell biology. In this study, we investigated MHC-E immunobiology in two common nonhuman primate species, Indian-origin rhesus macaques (RM) and Mauritian-origin cynomolgus macaques (MCM). Compared to humans and MCM, RM expressed a greater number of MHC-E alleles at both the population and individual level. Despite this difference, human, RM, and MCM MHC-E molecules were expressed at similar levels across immune cell subsets, equivalently upregulated by viral pathogens, and bound and presented identical peptides to CD8+ T cells. Indeed, SIV-specific, Mamu-E-restricted CD8+ T cells from RM recognized antigenic peptides presented by all MHC-E molecules tested, including cross-species recognition of human and MCM SIV-infected CD4+ T cells. Thus, MHC-E is functionally conserved among humans, RM, and MCM, and both RM and MCM represent physiologically relevant animal models of HLA-E-restricted T cell immunobiology.

Human mucosal-associated invariant T (MAIT) cells possess capacity for B cell help.

Mucosal-associated invariant T (MAIT) cells are an innate-like T cell subset, restricted by the nonclassic MHC class I-related protein MR1 and enriched at mucosal sites. Human studies have shown an association between MAIT cells and pathogen-specific antibody responses. In this study, we investigate the effect of human MAIT cells on B cells ex vivo. We found that supernatants from microbe- or cytokine-stimulated MAIT cells, when added to purified autologous B cells, increase frequencies of plasmablasts and promote IgA, IgG, and IgM production. We found effects to be mostly MR1-dependent and that the increases in plasmablasts are likely a result of increased differentiation from memory B cells. Furthermore, microbe-activated MAIT cell supernatant contains multiple cytokines known to stimulate B cells, including IL-6, -10, and -21. This study thus provides the first direct evidence of a newly identified role of MAIT cells in providing help to B cells.

Neutralizing Antibody Responses to Viral Infections Are Linked to the Non-classical MHC Class II Gene H2-Ob

Select humans and animals control persistent viral infections via adaptive immune responses that include production of neutralizing antibodies. The precise genetic basis for the control remains enigmatic. Here, we report positional cloning of the gene responsible for production of retrovirus-neutralizing antibodies in mice of the I/LnJ strain. It encodes the beta subunit of the non-classical major histocompatibility complex class II (MHC-II)-like molecule H2-O, a negative regulator of antigen presentation. The recessive and functionally null I/LnJ H2-Ob allele supported the production of virus-neutralizing antibodies independently of the classical MHC haplotype. Subsequent bioinformatics and functional analyses of the human H2-Ob homolog, HLA-DOB, revealed both loss- and gain-of-function alleles, which could affect the ability of their carriers to control infections with human hepatitis B (HBV) and C (HCV) viruses. Thus, understanding of the previously unappreciated role of H2-O (HLA-DO) in immunity to infections may suggest new approaches in achieving neutralizing immunity to viruses.

Abstract 2999: NKG2A checkpoint receptor expression on tumor-infiltrating CD8+ T cells restrains efficacy of immunotherapy

Abstract The inhibitory receptor CD94-NKG2A is expressed on subsets of NK cells and cytotoxic CD8+ T cells, but not CD4+ T helper cells. It’s ligand is the non-classical MHC class I molecule HLA-E, commonly over-expressed in human cancers. We previously demonstrated that high HLA-E expression in ovarian carcinoma and NSCLC may neutralize the survival benefit of T cell infiltration, suggesting that NKG2A could represent an immune checkpoint that tumors may exploit for immune evasion. We thus examined NKG2A expression on human and mouse Tumor Infiltrating Lymphocyte (TIL) populations and tested blockade of this receptor in mouse tumor models. Multicolor flow cytometry analysis showed higher frequency of NKG2A+CD8+ T cells in TIL samples obtained from biopsies of human head- and neck- carcinomas (n=17) compared to peripheral blood CD8+ T cells. Frequency of NKG2A+ TIL was higher in patients with measurable immune reactivity to HPV16-viral antigens than in patients without tumor immunity (14.8% vs 4.7%, respectively). Interestingly, only subsets of the NKG2A+ T cells co-expressed other inhibitory receptors, including PD-1 and TIM3, suggesting the existence of a ‘dedicated’ T cell subset for NKG2A. In our mouse tumor models we observed similar findings and, moreover, demonstrated that therapeutic vaccines comprising tumor antigens increased the frequency of NKG2A+-positive CD8+ TIL up to 80%. Similarly, expression of Qa-1, the mouse homologue of HLA-E, on tumor cells also increased after therapeutic vaccination. As a proof of concept that the NKG2A - Qa-1 axis limits the efficacy of immunotherapy, NKG2A blocking antibodies were applied in the TC-1 model together with therapeutic peptide vaccination. Combination treatment with vaccine and NKG2A blockade significantly improved the therapeutic efficacy compared to the vaccine alone, both in terms of survival (P<0.05, median survival 44 vs 31 days) as well as overall response rate, defined as decreased tumor size of more than 30% (93.8% vs 43.8%, respectively). Similar results were obtained in Qa-1-deficient mice, suggesting that Qa-1 on tumor cells is of importance. In conclusion, inhibitory receptor NKG2A is enriched on CD8+ TIL and functions as an immune checkpoint that restrains therapeutic efficacy of a cancer vaccine. Citation Format: Thorbald van Hall, Nadine van Montfoort, Sytse Piersma, Marjolein Sluijter, Marij J. Welters, Pascale André, Nicolai Wagtmann, Sjoerd H. van der Burg. NKG2A checkpoint receptor expression on tumor-infiltrating CD8+ T cells restrains efficacy of immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2999. doi:10.1158/1538-7445.AM2017-2999

Comparison of 6B11 mAb and α-GalCer-loaded CD1d dextramers for detection of iNKT cells by flow cytometry

Invariant natural killer T (iNKT) cells are a small population of thymus-derived T cells that are restricted by non-classical MHC class I molecule CD1d and express an evolutionary conserved TCR with an invariant α-chain. The frequency of iNKT cells in peripheral blood is very low, thus, accurate methods to identify and enumerate iNKT cells are needed. The aim of the study was to compare 6B11 mAb or α-GalCer-loaded CD1d dextramers usage in iNKT cell detection. The frequency of CD3+CD56+ lymphocytes is much higher, with statistical significance (p<0,001), than real iNKT cells detected by 6B11 mAb or α-GalCer-loaded CD1d dextramers. The frequency of iNKT cells, recognized by 6B11 mAb or α-GalCer-loaded CD1d dextramers, was in a similar range. Nonetheless, when we compared whether 6B11+ and α-GalCer-loaded CD1d dextramers+ are the same populations, it turned out that by this approach we were able to identify three distinct subsets of iNKT cells: i) 6B11+/α-GalCer-loaded dextramer− cells, ii) 6B11+/α-GalCer-loaded dextramer+ cells, and iii) 6B11−/α-GalCer-loaded dextramer+. Thus, although 6B11 mAb and α-GalCer-loaded dextramers may identify not exactly the same cells, application of these methods seems to give similar results of iNKT cell frequency in peripheral blood. It seems that both approaches for iNKT detection can be used for precise identification of these cells. Moreover, our results indicate that CD3+CD56+ lymphocytes are a heterogeneous population of T cells, expressing activation markers of both NK and T lymphocytes, yet with not well characterized properties.

Tumour and placenta establishment: The importance of antigen processing and presentation

Classical and non-classical MHC class I (MHC I) molecules displayed at the cell surface are essential for the induction of innate and adaptive immune responses. Classical MHC I present endogenously derived peptides to CD8+ T cells for immunosurveillance of infected or malignant cells. By contrast, non-classical MHC I, in particular HLA-G, also display peptides, but primarily act as immunomodulatory ligands for the innate immune response and are an important component for extravillous trophoblast invasion to form the placenta in pregnancy. Endoplasmic Reticulum AminoPeptidase 1 (ERAP1), which trims peptides in the ER to generate ligands for MHC I loading, is a key regulator of the peptide repertoire and has a significant impact on the formation of stable MHC I at the cell surface. ERAP1 also plays a role in angiogenesis, cell cycle progression and migration, events that are shared between tumour cells and placenta formation. Here we discuss the similarities between tumour and extravillous trophoblast cells in their immune modulatory, invasion, migration and proliferation properties in the context of ERAP1 and its role in establishment of solid tumours and placenta formation.

HLA-G: A Versatile Biomarker

HLA-G is a non-classical MHC Class I molecule that is usually restricted to cytotrophoblasts and other fetal cells in order to protect the fetus from maternal immune responses. However, HLA-G is expressed during viral infections and cancer. The presence of soluble HLA-G, sHLA-G, in serum is noted in various instances of human diseases. Serum HLA-G can serve as a biomarker, which makes it a unique tool in developing confirmatory diagnostics for infectious diseases and cancer. An accurate and early diagnosis of deadly diseases is really important for an early intervention or treatment. The immunosuppressive capabilities of HLA-G make it not only a biomarker for diagnosis but also an excellent target for developing therapeutics. Here, I discuss HLA-G’s potential to be an invaluable tool as a biomarker for diagnostics and a target for therapeutics.

CD8αα+MHC Class II+ Cell with the Capacity To Terminate Autoimmune Inflammation Is a Novel Antigen-Presenting NK-like Cell in Rats.

Discovery of immune tolerance mechanisms, which inhibit pre-existing autoimmune inflammation, may provide us with new strategies for treating autoimmune diseases. We have identified a CD8αα+MHC class II+ cell with professional APC capacity during our investigation on spontaneous recovery from autoimmune glomerulonephritis in a rat model. This cell actively invades inflamed target tissue and further terminates an ongoing autoimmune inflammation by selective killing of effector autoreactive T cells. In this study, we show that this cell used a cytotoxic machinery of Ly49s+ NK cells in killing of target T cells. Thus, this CD8αα+MHC class II+ cell was a dually functional Ag-presenting NK-like (AP-NK) cell. Following its coupling with target T cells through Ag presentation, killing stimulatory receptor Ly49s6 and coreceptor CD8αα on this cell used rat nonclassic MHC class I C/E16 on the target T cells as a ligand to initiate killing. Thus, activated effector T cells with elevated expression of rat nonclassic MHC class I C/E16 were highly susceptible to the killing by the CD8αα+ AP-NK cell. Granule cytolytic perforin/granzyme C from this cell subsequently mediated cytotoxicity. Thus, inhibition of granzyme C effectively attenuated the killing. As it can recognize and eliminate effector autoreactive T cells in the inflamed target tissue, the CD8αα+ AP-NK cell not only represents a new type of immune cell involved in immune tolerance, but it also is a potential candidate for developing a cell-based therapy for pre-existing autoimmune diseases.

Gene conversion of the major histocompatibility complex class I Caja-G in common marmosets (Callithrix jacchus).

SummaryCurrently, the amount of sequenced and classified MHC class I genes of the common marmoset is limited, in spite of the wide use of this species as an animal model for biomedical research. In this study, 480 clones of MHC class I G locus (Caja‐G) cDNA sequences were obtained from 21 common marmosets. Up to 10 different alleles were detected in each common marmoset, leading to the assumption that the Caja‐G loci duplicated in the marmoset genome. In the investigated population, four alleles occurred more often, giving evidence for higher immunological advantage of these alleles. In contrast to the human non‐classical MHC class I genes, Caja‐G shows high rates of polymorphism at the relevant peptide‐binding sites, despite its phylogenetic relationship to the non‐classical HLA‐G. Our results provide information for better understanding of the immunological properties of the common marmoset and confirm the theory of a gene conversion of the Caja‐G due to its detected plasticity and the absence of any known HLA‐A equivalent.

ERAAP Shapes the Peptidome Associated with Classical and Nonclassical MHC Class I Molecules.

The peptide repertoire presented by classical as well as nonclassical MHC class I (MHC I) molecules is altered in the absence of the endoplasmic reticulum aminopeptidase associated with Ag processing (ERAAP). To characterize the extent of these changes, peptides from cells lacking ERAAP were eluted from the cell surface and analyzed by high-throughput mass spectrometry. We found that most peptides found in wild-type (WT) cells were retained in the absence of ERAAP. In contrast, a subset of "ERAAP-edited" peptides was lost in WT cells, and ERAAP-deficient cells presented a unique "unedited" repertoire. A substantial fraction of MHC-associated peptides from ERAAP-deficient cells contained N-terminal extensions and had a different molecular composition than did those from WT cells. We found that the number and immunogenicity of peptides associated with nonclassical MHC I was increased in the absence of ERAAP. Conversely, only peptides presented by classical MHC I were immunogenic in ERAAP-sufficient cells. Finally, MHC I peptides were also derived from different intracellular sources in ERAAP-deficient cells.

Exploring the functions of nonclassical MHC class Ib genes in Xenopus laevis by the CRISPR/Cas9 system

A large family of highly related and clustered Xenopus nonclassical MHC class Ib (XNC) genes influences Xenopus laevis immunity and potentially other physiological functions. Using RNA interference (RNAi) technology, we previously demonstrated that one of XNC genes, XNC10.1, is critical for the development and function of a specialized innate T (iT) cell population. However, RNAi limitation such as a variable and unstable degree of gene silencing in F0 and F1 generations is hampering a thorough functional analysis of XNC10.1 and other XNC genes. To overcome this obstacle, we adapted the CRISPR/Cas9-mediated gene editing technique for XNC genes. We efficiently and specifically generated single gene knockouts of XNC10.1, XNC11, and XNC1 as well as double gene knockouts of XNC10.1 and XNC11 in X. laevis. In single XNC10.1 knockout X. laevis tadpoles, the absence of XNC10.1 and Vα6-Jα1.43 invariant T cell receptor rearrangement transcripts indicated XNC10.1 loss-of-function and deficiency in Vα6-Jα1.43 iT cells. Notably, targeting XNC10.1 did not affect neighboring XNC genes exhibiting high sequence similarity. Furthermore, XNC1 gene disruption induced mortality during developmental stage 47, suggesting some non-immune but essential function of this gene. These data demonstrate that the CRISPR/Cas9 system can be successfully adapted for genetic analysis in F0 generation of X. laevis.

HNRNPR Regulates the Expression of Classical and Nonclassical MHC Class I Proteins.

MHC class I molecules, in addition to their role in specific activation of the CTL of adaptive immune system, function also as the main ligands for NK cell inhibitory receptors, which prevent NK cells from killing normal, healthy cells. MHC class I proteins are divided into classical and nonclassical proteins. The former group consists of hundreds of HLA-A, B, and C alleles, which are universally expressed, whereas several alleles of the latter group, such as HLA-G, manifest a restricted expression pattern. Despite the important role played by these molecules in innate and adaptive immune responses, their complex expression regulation is not fully known. In our study, we investigated the regulation processes controlling the expression of MHC class I molecules, with a particular focus on their 3' untranslated regions. We identified heterogeneous nuclear ribonucleoprotein R (HNRNPR) as an important positive regulator of classical and nonclassical MHC class I molecules. HNRNPR is a RNA-binding protein belonging to the heterogeneous nuclear ribonucleoprotein family, which has a known role in processing of precursor mRNA. We demonstrated that HNRNPR binds MHC class I mRNAs in their 3' untranslated regions and enhances their stability and consequently their expression. Furthermore, regulation by HNRNPR modulates the cytotoxic activity of NK cells. In conclusion, we show that HNRNPR acts as a general positive regulator of MHC class I expression.