Non-classical Major Histocompatibility Complex Research Articles

Biological Characteristics of HLA-G and Its Role in Solid Organ Transplantation.

Organ transplantation is a lifesaving option for patients with advanced diseases. Rejection is regarded as one of the most severe risk factors post-transplantation. A molecule that contributes to immune tolerance and resisting rejection is human leukocyte antigen (HLA)-G, which belongs to the non-classical major histocompatibility complex class (MHC) I family. HLA-G was originally found to play a role during pregnancy to maintain immune tolerance between mother and child. It is expressed in the placenta and detected in several body fluids as soluble factor as well as different membrane isoforms on cells. Recent findings on HLA-G show that it can also play multifaceted roles during transplantation. This review will explain the general characteristics and biological function of HLA-G and summarize the views supporting the tolerogenic and other roles of HLA-G to better understand its role in solid organ transplantation (SOT) and its complications. Finally, we will discuss potential future research on the role of HLA-G in prevention, diagnosis, and treatment in SOT.

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.

Regulation of the BCR signalosome by the class II peptide editor, H2-M, affects the development and repertoire of innate-like B cells.

The non-classical Major Histocompatibility Complex class II (MHCII) protein, H2-M, edits peptides bound to conventional MHCII in favor of stable peptide/MHCII (p/MHCII) complexes. Here, we show that H2-M deficiency affects B-1 cell survival, reduces cell renewal capacity, and alters immunoglobulin repertoire, allowing for the selection of cells specific for highly abundant epitopes, but not low-frequency epitopes. H2-M-deficient B-1 cells have shorter CDR3 length, higher content of positively charged amino acids, shorter junctional regions, less mutation frequency, and a skewed clonal distribution. Mechanistically, H2-M loss reduces plasma membrane p/MHCII association with B cell receptors (BCR) on B-1 cells and diminishes integrated BCR signal strength, a key determinant of B-1 cell selection, maturation, and maintenance. Thus, H2-M:MHCII interaction serves as a cell-intrinsic regulator of BCR signaling and influences the selection of the B-1 cell clonal repertoire.

The HLA-G Immune Checkpoint Plays a Pivotal Role in the Regulation of Immune Response in Autoimmune Diseases.

The human G-leukocyte antigen (HLA-G) molecule is a non-classical major histocompatibility complex (MHC) class I molecule. The pertinence of HLA-G has been investigated in numerous studies which have sought to elucidate the relevance of HLA-G in pathologic conditions, such as autoimmune diseases, cancers, and hematologic malignancies. One of the main goals of the current research on HLA-G is to use this molecule in clinical practice, either in diagnostics or as a therapeutic target. Since HLA-G antigens are currently considered as immunomodulatory molecules that are involved in reducing inflammatory and immune responses, in this review, we decided to focus on this group of antigens as potential determinants of progression in autoimmune diseases. This article highlights what we consider as recent pivotal findings on the immunomodulatory function of HLA-G, not only to establish the role of HLA-G in the human body, but also to explain how these proteins mediate the immune response.

Identification of Non-classical Follicular T Cells.

Nonclassical T cells are a heterogeneous group of T lymphocytes that are activated during the early stages of infection and act as a bridge between the innate and adaptive immune system. Among them, Natural Killer T (NKT) cells have been extensively studied in the last two decades due to their unique ability to recognize foreign/self-lipid antigens in the context of CD1d, a nonclassical major histocompatibility complex molecule. In this chapter, we describe our protocols to track murine NKT cells in lymph nodes by flow cytometry and confocal microscopy.

HLA-G/sHLA-G and HLA-G-Bearing Extracellular Vesicles in Cancers: Potential Role as Biomarkers.

As a non-classic major histocompatibility complex (MHC) class I molecule, human leukocyte antigen G (HLA-G) is expressed in fetal-maternal interface and immunoprivileged site only in healthy condition, and in pathological conditions such as cancer, it can be de novo expressed. It is now widely accepted that HLA-G is a key molecule in the process of immune escape of cancer cells, which is ubiquitously expressed in the tumor environment. This raises the possibility that it may play an adverse role in tumor immunity. The expression level of HLA-G has been demonstrated to be highly correlated with clinical parameters in many tumors, and its potential significance in the diagnosis and prognosis of cancer has been postulated. However, because HLA-G itself has up to seven different subtypes, and for some subtypes, detected antibodies are few or absent, it is hard to evaluate the actual expression of HLA-G in tumors. In the present work, we described (a) the structure and three main forms of HLA-G, (b) summarized the mechanism of HLA-G in the immune escape of tumor cells, (c) discussed the potential role of HLA-G as a tumor marker, and reviewed (d) the methods for detecting and quantifying HLA-G.

Mechanisms of Immunomodulation and Cytoprotection Conferred to Pancreatic Islet by Human Amniotic Epithelial Cells

Inhibiting pro-inflammatory cytokine activity can reverse inflammation mediated dysfunction of islet grafts. Human amniotic epithelial cells (hAECs) possess regenerative, immunomodulatory and anti-inflammatory properties. We hypothesized that hAECs could protect islets from cellular damage induced by pro-inflammatory cytokines. To verify our hypothesis, hAEC monocultures, rat islets (RI), or RI-hAEC co-cultures where exposed to a pro-inflammatory cytokine cocktail (Interferon γ: IFN-γ, Tumor necrosis factor α: TNF-α and Interleukin-1β: IL-1β). The secretion of anti-inflammatory cytokines and gene expression changes in hAECs and viability and function of RI were evaluated. The expression of non-classical Major Histocompatibility Complex (MHC) class I molecules by hAECs cultured with various IFN-γ concentrations were assessed. Exposure to the pro-inflammatory cocktail significantly increased the secretion of the anti-inflammatory cytokines IL6, IL10 and G-CSF by hAECs, which was confirmed by upregulation of IL6, and IL10 gene expression. HLA-G, HLA-E and PDL-1 gene expression was also increased. This correlated with an upregulation of STAT1, STAT3 and NF-κB1gene expression levels. RI co-cultured with hAECs maintained normal function after cytokine exposure compared to RI cultured alone, and showed significantly lower apoptosis rate. Our results show that exposure to pro-inflammatory cytokines stimulates secretion of anti-inflammatory and immunomodulatory factors by hAECs through the JAK1/2 – STAT1/3 and the NF-κB1 pathways, which in turn protects islets against inflammation-induced damages. Integrating hAECs in islet transplants appears as a valuable strategy to achieve to inhibit inflammation mediated islet damage, prolong islet survival, improve their engraftment and achieve local immune protection allowing reducing systemic immunosuppressive regimens.Graphical This study focuses on the cytoprotective effect of isolated hAECs on islets exposed to pro-inflammatory cytokines in vitro. Exposure to pro-inflammatory cytokines stimulated secretion of anti-inflammatory and immunomodulatory factors by hAECs putatively through the JAK1/2 – STAT1/3 and the NF-κB1 pathways. This had protective effect on islets against inflammation-induced damages. Taken together our results indicate that incorporating hAECs in islet transplants could be a valuable strategy to inhibit inflammation mediated islet damage, prolong islet survival, improve their engraftment and achieve local immune protection allowing to reduce systemic immunosuppressive regimens.

Molecular characterization, expression patterns, and subcellular localization of a classical and a novel nonclassical MHC class I α molecules from Japanese eel Anguilla japonica

Major histocompatibility complex (MHC) molecules play a critical role in the immune response of vertebrate animals by presenting foreign antigens to T lymphocytes. In this study, we first cloned and identified a classical and a novel nonclassical MHC I α molecules from Japanese eel (Anguilla japonica), named as AjMHC I-UBA and AjMHC I-L, respectively. The full-length cDNA of AjMHC I-UBA contains an open reading frame (ORF) of 1047 bp encoding a predicted protein of 348 amino acids, while AjMHC I-L 1089 bp encodes 362 amino acids. The multiple alignment of the amino acid sequence showed that AjMHC I-UBA and AjMHC I-L consist of an N-terminal MHC I superfamily domain within α1 and α2 helices, an IgC-MHC I α3 domain, and a transmembrane region close to the C-terminal, which are similar to other fish and mammal species. Molecular polymorphism analysis showed that eight different major alleles of AjMHC I-UBA, named as AjMHC I-UBA*0101～1001, were identified from six Japanese eel individuals. Furthermore, a distinguishing signature of the nonclassical L-lineage genes specific motif “HINMTL”, including an N-glycosylation site (NXS/T), was present in the α3 domain of AjMHC I-L sequences. Although the predicted three-dimensional structures of AjMHC I-UBA and AjMHC I-L are similar to that of human MHC I α, phylogenetic analysis showed that these two protein molecules belong to classical MHC I UBA gene of U-lineage and nonclassical MHC I gene of L-lineage, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the highest expression of AjMHC I-UBA and AjMHC I-L was found in the intestine and the expression level of AjMHC I-UBA in all of the examined tissues was significantly higher than that of AjMHC I-L. The expressions of AjMHC I-UBA and AjMHC I-L in liver, kidney and spleen were significantly induced following injection with the viral mimic poly I:C, LPS, and Aeromonas hydrophila infection. In vitro, the AjMHC I-UBA and AjMHC I-L transcripts of Japanese eel liver cells were significantly enhanced by the treatment of poly I:C or the stimulation of the high concentration of A. hydrophila. Subcellular localization showed that under natural conditions, AjMHC I-UBA and AjMHC I-L were uniformly distributed in the cytoplasm, and aggregated into spots or flakes in the cytoplasm after the stimulation of poly I:C or LPS. These results collectively suggested that AjMHC I-UBA and AjMHC I-L are important components possibly involved in Japanese eel defense against viral and bacterial infection. Taken together, these findings provide valuable insight into the immune function of MHC class I in the immune system of teleost.

Prognostic significance of high circulating sHLA-G in ovarian carcinoma.

HLA-G is a non-classical major histocompatibility complex class Ib molecule. Its expression has been described in various cancer types, including ovarian cancer. HLA-G molecule has been implicated in immune escape and in progression of ovarian tumor cells. Our goal was to assess if total soluble (s)HLA-G molecules or HLA-G5 and sHLA-G1 isoforms could be considered as circulating ovarian tumor biomarkers, we measured the concentration of these molecules in ovarian carcinoma patients stratified according with their clinicopathological parameters. sHLA-G, sHLA-G1 and HLA-G5 concentrations were dosed in plasma samples by sandwich-ELISA. The sHLA-G dimerization was analyzed after immunoprecipitation and SDS-PAGE migration. Total sHLA-G and sHLA-G1 levels were significantly represented in plasma of ovarian carcinoma patients compared to healthy controls. sHLA-G1 isoform concentration was highly represented in ovarian carcinoma compared to HLA-G5 isoforms. Additionally, high sHLA-G molecules have been found in aged patients, as well as in patients with advanced stages, and those with metastatic lymph nodes and those with distant metastasis. Elsewhere, sHLA-G monomers were highly represented in ovarian carcinoma patients compared to controls. sHLA-G plasmatic protein was highly represented in ovarian carcinoma. In effect, HLA-G might be considered as a new checkpoint molecule that could be used to assess progression and recurrence of the disease, thus placing it as a potential biomarker for advanced and complicated ovarian carcinoma.

NKG2A Represents an Important Immune Checkpoint for Human Cytokine-Induced Memory-like NK Cells in Patients with AML

Natural killer (NK) cells are an emerging cellular immunotherapy for patients with acute myeloid leukemia (AML); however, the best approach to maximize NK cell anti-leukemia potential is unclear. Paradigm-shifting reports have shown that NK cells exhibit “memory-like” properties following hapten exposure, virus infection, or combined cytokine pre-activation. We and others have established that human cytokine-induced memory-like NK cells display enhanced re-stimulation responses to numerous activating stimuli, including tumor target cells. We have translated this finding into a potential cellular therapy for rel/ref AML patients, and have completed the dose escalation of a phase 1/2 study. We demonstrated that donor memory-like NK cells expand in patients' blood and bone marrow and retain enhanced functionality ex vivo. Further, of the 11 evaluable AML patients, we have observed 7 clinical responses, including 6 patients with CR/CRi. Since NK cell recognition depends on signals from multiple activating and inhibitory receptors, we developed mass cytometry panels to immunophenotype and track the diversity and effector functions of these human in vivo-differentiated memory-like NK cells.Mass cytometry revealed that in vivo-differentiated memory-like (ML) NK cells were distinct from baseline (BL) NK cells from the same donor, as well as NK cells from normal donor PBMC. Multidimensional analyses revealed a memory-like phenotype: CD56hi CD11blo CD62L+ NKG2Ahi NKp30hi Ki-67+ (baseline: 2.7% ± 0.45% v memory-like: 87.4% ± 5.3%, mean ± SEM within memory-like gate). Furthermore, Citrus analyses revealed that higher NKG2A expression was significantly correlated with treatment failure [TF median NKG2A= 89.2 ± 25.0; responder median NKG2A= 8.1 ± 2.9 (mean ± SEM); p = 0.007, n = 5 each group; SAM FDR <0.1]. NKG2A is a C-type lectin receptor with two immunoreceptor tyrosine-based inhibitory motifs. Signaling through NKG2A is achieved when it engages its ligand, HLA-E. HLA-E is a non-classical major histocompatibility complex class I molecule that is expressed abundantly on many normal tissue types as well as tumors, including AML. Based on these findings that NKG2A is upregulated on memory-like NK cells and the intensity of NKG2A on memory-like NK cells correlates with patient responses, we hypothesized that NKG2A/HLA-E interactions in vivo represent a major barrier to memory-like NK cell responses.In order to interrogate the role NKG2A may play in limiting cytokine-induced memory-like NK cell responses, we stimulated in vitro-differentiated ML cells with the K562 AML tumor line displaying forced expression of HLA-E (K562-E) in the presence of isotype or anti-NKG2A blocking antibody. Increased IFN-γ (p = 0.02) and TNF (p = 0.05) production by NKG2A-blocked ML NK cells was detected, compared to isotype-treated ML NK cells. We performed a similar stimulation assay using primary AML blasts as targets, which express a variable level of HLA-E. Again, ML NK cells treated with NKG2A blockade produced significantly more IFN-γ than isotype-treated ML NK cells (p = 0.001). Studies examining NKG2A checkpoint blockade on in vivo-differentiated ML NK cells from AML patients are ongoing. In conclusion, in vivo-differentiated cytokine-induced memory-like NK cells are phenotypically distinct from conventional NK cells. Further, targeting NKG2A expressed on ML NK cells in vivo has the potential to enhance the clinical efficacy of memory-like NK cell therapy for patients. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Evidence for a shift in placental HLA-G allelic dominance and the HLA-G isoform profile during a healthy pregnancy and preeclampsia†.

Human leukocyte antigen (HLA)-G, which belongs to a nonclassical class Ib major histocompatibility complex gene family expressed by placental trophoblast cells, plays a central role in establishing tolerance to the semiallogeneic fetus and in placentation. HLA-G exists in different soluble or membrane-bound isoforms. Preeclampsia, a major cause of fetal and maternal morbidity and mortality, has been linked to insufficient placentation and an altered immune response in pregnancy, including altered HLA-G expression. The 14bp insertion/deletion polymorphism in the 3' untranslated region of the gene and the isoform profile may affect HLA-G expression. The aim of the current pilot study was to characterize the expression patterns of HLAG mRNA, protein, and isoform profile in uncomplicated term pregnancies and in cases of preeclampsia. Maternal sHLA-G mRNA and protein levels were slightly reduced in preeclampsia. No difference was found for placental blood, and no correlation between peripheral and placental sHLA-G levels was found. We observed no association between neither fetal nor maternal HLA-G 14bp insertion/deletion genotypes and preeclampsia, nor a significant difference in isoform profiles. However, in HLA-G 14bp insertion/deletion heterozygous placental samples, we observed abundant HLA-G1 14bp insertion allele expression in the term placentae, which is contrary to previous findings in first trimester trophoblast. Increased HLA-G1 14bp insertion allele expression in the placenta was associated with reduced levels of placental sHLA-G and an altered isoform profile with increased relative levels of HLA-G1 and -G5 and reduced levels of HLA-G3. The results indicate that an allelic shift in heterozygous individuals could represent a novel regulatory pathway.

The Role of HLA-G in Tumor Escape: Manipulating the Phenotype and Function of Immune Cells.

Human leukocyte antigen-G (HLA-G) is a non-classical major histocompatibility complex class I (MHC I) molecule, and under physiological conditions, its expression is strictly restricted to the maternal–fetal interface and immune-privileged organs where HLA-G is expected to contribute to establishment and maintenance of immune tolerance. However, the expression of HLA-G has been found in various types of tumors, and the level of its expression frequently correlates with high-grade histology and poor prognosis, raising the possibility that it may play a negative role in tumor immunity. ILT2 and ILT4, present on a broad of immune cells, have been identified as the main receptors engaging HLA-G, and their interactions have been found to allow the conversion of effectors like NK cells and T cells to anergic or unresponsive state, activated DCs to tolerogenic state, and to drive the differentiation of T cells toward suppressive phenotype. Therefore, tumors can employ HLA-G to modulate the phenotype and function of immune cells, allowing them to escape immune attack. In this review, we discuss the mechanism underlying HLA-G expression and function, its role played in each step of the tumor-immunity cycle, as well as the potential to target it for therapeutic benefit.

Revisiting nonclassical HLA II functions in antigen presentation: Peptide editing and its modulation.

The nonclassical major histocompatibility complex of class II molecules (ncMHCII) HLA-DM (DM) and HLA-DO (DO) feature essential functions for the selection of the peptides that are displayed by classical MHCII proteins (MHCII) for CD4+ Th cell surveillance. Thus, although the binding groove of classical MHCII dictates the main features of the peptides displayed, ncMHCII function defines the preferential loading of peptides from specific cellular compartments and the extent to which they are presented. DM acts as a chaperone for classical MHCII molecules facilitating peptide exchange and thereby favoring the binding of peptide-MHCII complexes of high kinetic stability mostly in late endosomal compartments. DO on the other hand binds to DM blocking its peptide-editing function in B cells and thymic epithelial cells, limiting DM activity in these cellular subsets. DM and DO distinct expression patterns therefore define specific antigen presentation profiles that select unique peptide pools for each set of antigen presenting cell. We have come a long way understanding the mechanistic underpinnings of such distinct editing profiles and start to grasp the implications for ncMHCII biological function. DM acts as filter for the selection of immunodominant, pathogen-derived epitopes while DO blocks DM activity under certain physiological conditions to promote tolerance to self. Interestingly, recent findings have shown that the unexplored and neglected ncMHCII genetic diversity modulates retroviral infection in mouse, and affects human ncMHCII function. This review aims at highlighting the importance of ncMHCII function for CD4+ Th cell responses while integrating and evaluating what could be the impact of distinct editing profiles because of natural genetic variations.

Characterization of two MHC II genes (DOB, DRB) in white-tailed deer (Odocoileus virginianus)

BackgroundThe major histocompatibility complex (MHC) is responsible for detecting and addressing foreign pathogens inside the body. While the general structure of MHC genes is relatively well conserved among mammalian species, it is notably different among ruminants due to a chromosomal inversion that splits MHC type II genes into two subregions (IIa, IIb). Recombination rates are reportedly high between these subregions, and a lack of linkage has been documented in domestic ruminants. However, no study has yet examined the degree of linkage between these subregions in a wild ruminant. The white-tailed deer (Odocoileus virginianus), a popular ruminant of the Cervidae family, is habitually plagued by pathogens in its natural environment (e.g. Haemonchus contortus, Elaeophora). Due to the association between MHC haplotypes and disease susceptibility, a deeper understanding of MHC polymorphism and linkage between MHC genes can further aid in this species’ successful management. We sequenced MHC-DRB exon 2 (IIa) and MHC-DOB exon 2 (IIb) on the MiSeq platform from an enclosed white-tailed deer population located in Alabama.ResultsWe identified 12 new MHC-DRB alleles, and resampled 7 alleles, which along with other published alleles brings the total number of documented alleles in white-tailed deer to 30 for MHC-DRB exon 2. The first examination of MHC-DOB in white-tailed deer found significantly less polymorphism (11 alleles), as was expected of a non-classical MHC gene. While MHC-DRB was found to be under positive, diversifying selection, MHC-DOB was found to be under purifying selection for white-tailed deer. We found no significant linkage disequilibrium between MHC-DRB and MHC-DOB, suggesting that these loci are unlikely to be closely linked.ConclusionsOverall, this study identified 12 new MHC-DRB exon 2 alleles and characterized a new, non-classical, MHC II gene (MHC-DOB) for white-tailed deer. We also found a lack of significant linkage between these two loci, which supports previous findings of a chromosomal inversion within the MHC type II gene region in ruminants, and suggests that white-tailed deer may have a recombination hotspot between these MHC regions similar to that found for Bos taurus.

Expression of Tolerogenic HLA-G Confer Worse Outcome in Patients with Chronic Lymphocytic Leukemia

Background: Chronic lymphocytic leukemia (CLL) is the most common hematological malignancy in adults presenting with varied clinical course. There is a higher request for founding known prognostic factors for stratifying CLL patients. HLA-G is a non-classical major histocompatibility complex (MHC) molecule. It can be expressed in membrane bound (m.HLA-G) and soluble forms (s.HLA-G). Objective: The aim of work was to investigate the expression of membrane form of HLA-G in CLL and correlate findings and a variety of clinical and laboratory variables. Patients and methods: This prospective study included a total of thirty newly diagnosed B-CLL patients, attending at Hematology Unit, Medical Oncology Department, national Cancer institute, Cairo. This study was conducted between December 2017 to July 2019. Diagnosis of CLL was confirmed by flow cytometric immunophenenotyping using standard lymphoma panel, membrane HLA-G was determined by flow cytometry. Results: The expression of HLA-G by flowcytometry was negatively correlated with the platelet count (r = -0.516, p = 0.004) and Hb (r = -0.479, p = 0.007). The expression of HLA-G was significantly higher in CD38 positive cells (p = 0.006). The expression of HLA-G was significantly higher in Rai stage 4 compared to stage 1 & stage 2 (p = 0.001). Conclusion: It could be concluded that expression of HLA-G might represent signs of immunosuppression in CLL patients which contribute to the immune escape of tumor cells. In addition, HLA-G expression by B-CLL cells correlates significantly with the known prognostic markers of disease progression, mainly Rai clinical staging, CD38 expression and worse survival, making this parameter possibly an important prognostic factors of disease progression.

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.

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.

MHC Restriction: Where Are We Now?

MHC Restriction: Where Are We Now?

Evolution of major histocompatibility complex class I genes in the sable Martes zibellina (Carnivora, Mustelidae).

The molecules encoded by major histocompatibility complex (MHC) genes play an essential role in the adaptive immune response among vertebrates. We investigated the molecular evolution of MHC class I genes in the sable Martes zibellina. We isolated 26 MHC class I sequences, including 12 putatively functional sequences and 14 pseudogene sequences, from 24 individuals from two geographic areas of northeast China. The number of putatively functional sequences found in a single individual ranged from one to five, which might be at least 1–3 loci. We found that both balancing selection and recombination contribute to evolution of MHC class I genes in M. zibellina. In addition, we identified a candidate nonclassical MHC class I lineage in Carnivora, which may have preceded the divergence (about 52–57 Mya) of Caniformia and Feliformia. This may contribute to further understanding of the origin and evolution of nonclassical MHC class I genes. Our study provides important immune information of MHC for M. zibellina, as well as other carnivores.

The murine CD94/NKG2 ligand, Qa-1b, is a high-affinity, functional ligand for the CD8αα homodimer

The immune co-receptor CD8 molecule (CD8) has two subunits, CD8α and CD8β, which can assemble into homo or heterodimers. Nonclassical (class-Ib) major histocompatibility complex (MHC) molecules (MHC-Ibs) have recently been identified as ligands for the CD8αα homodimer. This was demonstrated by the observation that histocompatibility 2, Q region locus 10 (H2-Q10) is a high-affinity ligand for CD8αα which also binds the MHC-Ib molecule H2-TL. This suggests that MHC-Ib proteins may be an extended source of CD8αα ligands. Expression of H2-T3/TL and H2-Q10 is restricted to the small intestine and liver, respectively, yet CD8αα-containing lymphocytes are present more broadly. Therefore, here we sought to determine whether murine CD8αα binds only to tissue-specific MHC-Ib molecules or also to ubiquitously expressed MHC-Ib molecules. Using recombinant proteins and surface plasmon resonance-based binding assays, we show that the MHC-Ib family furnishes multiple binding partners for murine CD8αα, including H2-T22 and the CD94/NKG2-A/B-activating NK receptor (NKG2) ligand Qa-1b We also demonstrate a hierarchy among MHC-Ib proteins with respect to CD8αα binding, in which Qa-1b > H2-Q10 > TL. Finally, we provide evidence that Qa-1b is a functional ligand for CD8αα, distinguishing it from its human homologue MHC class I antigen E (HLA-E). These findings provide additional clues as to how CD8αα-expressing cells are controlled in different tissues. They also highlight an unexpected immunological divergence of Qa-1b/HLA-E function, indicating the need for more robust studies of murine MHC-Ib proteins as models for human disease.