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Sample records for mhc ii expression

  1. Characterization and expression of MHC class II alpha and II beta genes in mangrove red snapper (Lutjanus argentimaculatus).

    PubMed

    Wang, Tianyan; Tan, Shangjin; Cai, Zhonghua

    2015-12-01

    The major histocompatibility complex (MHC) class II plays a key role in adaptive immunity by presenting foreign peptides to CD4(+) T cells and by triggering the adaptive immune response. While the structure and function of MHC class II have been well characterized in mammalian, limited research has been done on fishes. In this study, we characterized the gene structure and expression of MHC class II α (Lunar-DAA) and II β (Lunar-DAB) of mangrove red snapper (Lutjanus argentimaculatus). Both genes shared, respectively, a high similarity and typical features with other vertebrate MHC class II α and II β. The phylogenetic analysis of the deduced peptides revealed that both Lunar-DAA and Lunar-DAB were located in the teleost subclass. Western blotting analyses indicated that both MHC class II α and II β were expressed ubiquitously in immune-related cells, tissues and organs, and that MHC class II α and II β chains existed mainly as heterodimers. While it was highly expressed in gills, thymus, head kidney (HK), spleen, head kidney macrophage and spleen leucocytes, MHC class II β chain was expressed with a low abundance in skin, intestine, stomach and heart. The highest expression of MHC class II β in thymus confirmed the conclusion that thymus is one of the primary lymphoid organs in fishes. The detection of MHC class II αβ dimers in HK macrophages and spleen leucocytes indicated that HK macrophages and spleen leucocytes play a critical role in the adaptive immunity in fishes. All these results provide valuable information for understanding the structure of MHC class II α and II β and their function in immune responses. PMID:26454477

  2. MHC II expression in the CNS after long-term demyelination

    SciTech Connect

    Cannella, B.; Aquino, D.A.; Raine, C.S.

    1995-07-01

    The ability of chronically demyelinated central nervous system (CNS) tissue to express major histocompatibility complex (MHC) class II molecules has been measured in mouse spinal cord cultures exposed for 1 and 3 weeks to demyelinating anti-white matter (WM) serum. From previous studies, It was known that after 3 weeks of demyelination in vitro, such cultures are incapable of remyelination. In the present report, MHC II levels were evaluated by immunocytochemistry and by Western and Northern blots. The results have shown that after both 1 and 3 weeks of exposure to myelinotoxic anti-WM serum, the cultures retained the ability to express MHC II and this could be further upregulated by incubation with interferon {gamma} (IFN{gamma}). Control groups showed increased expression of MHC II with age. By immunocytochemistry, all groups of cultures expressed high levels of MHC II and all groups showed upregulation after IFN{gamma} treatment. Anti-WM-treated cultures demonstrated slightly higher levels of MHC II than controls. Morphologically, the MHC II expression was associated with the surface of astrocytes. Semiquantitative analysis by Western blotting confirmed the increase in class II MHC expression in the long-term treated cultures after IFN{gamma} exposure, revealing no differences between anti-WM-treated and complement-treated cultures. This was also supported by Northern blotting which showed similar mRNA levels in both groups. These findings suggest that long-term demyelinated CNS tissue still possesses the ability to interact with CD4{sup +} T cells, observations of significance to the expansion of the chronic multiple sclerosis lesion. 50 refs., 6 figs., 2 tabs.

  3. Expression of the MHC Class II Transactivator (CIITA) type IV promoter in B lymphocytes and regulation by IFN-γ

    PubMed Central

    Piskurich, Janet F.; Gilbert, Carolyn A.; Ashley, Brittany D.; Zhao, Mojun; Chen, Han; Wu, Jian; Wright, Kenneth L.

    2006-01-01

    The MHC class II transactivator (CIITA), the master regulator of MHC class II (MHC II) expression, is a co-activator that controls MHC II transcription. Human B lymphocytes express MHC II constitutively due to persistent activity of CIITA promoter III (pIII), one of the four potential promoters (pI-pIV) of this gene. Although increases in MHC II expression in B cells in response to cytokines have been observed and induction of MHC II and CIITA by IFN-γ has been studied in a number of different cell types, the specific effects of IFN-γ on CIITA expression in B cells have not been studied. To investigate the regulation of CIITA expression by IFN-γ in B cells, RT-PCR, in vivo and in vitro protein/DNA binding studies, and functional promoter analyses were performed. Both MHC II and CIITA type IV-specific RNAs increased in human B lymphocytes in response to IFN-γ treatment. CIITA promoter analysis confirmed that pIV is IFN-γ inducible in B cells and that the GAS and IRF-E sites are necessary for full induction. DNA binding of IRF-1 and IRF-2, members of the IFN regulatory factor family, was up-regulated in B cells in response to IFN-γ and increased the activity of CIITA pIV. In vivo genomic footprint analysis demonstrated proteins binding at the GAS, IRF-E and E box sites of CIITA pIV. Although CIITA pIII is considered to be the hematopoietic-specific promoter of CIITA, these findings demonstrate that pIV is active in B lymphocytes and potentially contributes to the expression of CIITA and MHC II in these cells. PMID:15950283

  4. IL-33 promotes MHC class II expression in murine mast cells

    PubMed Central

    Ito, Tomonobu; Egusa, Chizu; Maeda, Tatsuo; Numata, Takafumi; Nakano, Nobuhiro; Nishiyama, Chiharu; Tsuboi, Ryoji

    2015-01-01

    Mast cells (MCs), recognized as tissue-resident cells of hematopoietic origin, are involved in cellular and pathological manifestations of allergic disorders including atopic dermatitis. IL-33, a member of the IL-1 cytokine family, activates Th2-type immune responses, and promotes the degranulation and maturation of MCs. However, it is uncertain whether IL-33 treatment induces mature mast cells to acquire the characteristics of the monocyte-dendritic cell lineage.We investigated the effect of IL-33 on the MHC class II expression and function of murine mast cells. IL-33-treated mature murine bone marrow-derived mast cells (BMMCs) were analyzed by FACS, real-time PCR, chromatin immunoprecipitation (ChIP) assay, and Western blotting. The morphology and degranulation activity of BMMCs and T-cell activation by BMMCs were also examined. BMMCs treated with IL-33 for 10 days induced cell surface expression of the MHC class II protein, whereas the expression of FcεRI and c-kit was not affected by IL-33. The expression of CIITA, driven from pIII and pIV, was up-regulated in IL-33-treated BMMCs. The amount of PU.1 mRNA and protein significantly increased in IL-33-treated BMMCs. The ChIP assay showed PU.1 binding to CIITA pIII, and enhanced histone acetylation due to IL-33 treatment. Syngeneic T cells were activated by co-culture with IL-33-treated BMMCs, although the expression of the co-stimulatory molecules, CD40, CD80, CD86, and PDL-1, was not detected. Mast cells express MHC class II after prolonged exposure to IL-33, probably due to enhanced recruitment of PU.1 to CIITA pIII, resulting in transactivation of CIITA and MHC class II. IL-33 is an important cytokine in allergic disorders. Mast cells have the ability to express MHC class II after prolonged exposure to IL-33 in a murine model. IL-33 holds a key to understanding the etiology of atopic dermatitis. PMID:26417437

  5. IL-33 promotes MHC class II expression in murine mast cells.

    PubMed

    Ito, Tomonobu; Egusa, Chizu; Maeda, Tatsuo; Numata, Takafumi; Nakano, Nobuhiro; Nishiyama, Chiharu; Tsuboi, Ryoji

    2015-09-01

    Mast cells (MCs), recognized as tissue-resident cells of hematopoietic origin, are involved in cellular and pathological manifestations of allergic disorders including atopic dermatitis. IL-33, a member of the IL-1 cytokine family, activates Th2-type immune responses, and promotes the degranulation and maturation of MCs. However, it is uncertain whether IL-33 treatment induces mature mast cells to acquire the characteristics of the monocyte-dendritic cell lineage.We investigated the effect of IL-33 on the MHC class II expression and function of murine mast cells. IL-33-treated mature murine bone marrow-derived mast cells (BMMCs) were analyzed by FACS, real-time PCR, chromatin immunoprecipitation (ChIP) assay, and Western blotting. The morphology and degranulation activity of BMMCs and T-cell activation by BMMCs were also examined. BMMCs treated with IL-33 for 10 days induced cell surface expression of the MHC class II protein, whereas the expression of FcεRI and c-kit was not affected by IL-33. The expression of CIITA, driven from pIII and pIV, was up-regulated in IL-33-treated BMMCs. The amount of PU.1 mRNA and protein significantly increased in IL-33-treated BMMCs. The ChIP assay showed PU.1 binding to CIITA pIII, and enhanced histone acetylation due to IL-33 treatment. Syngeneic T cells were activated by co-culture with IL-33-treated BMMCs, although the expression of the co-stimulatory molecules, CD40, CD80, CD86, and PDL-1, was not detected. Mast cells express MHC class II after prolonged exposure to IL-33, probably due to enhanced recruitment of PU.1 to CIITA pIII, resulting in transactivation of CIITA and MHC class II. IL-33 is an important cytokine in allergic disorders. Mast cells have the ability to express MHC class II after prolonged exposure to IL-33 in a murine model. IL-33 holds a key to understanding the etiology of atopic dermatitis. PMID:26417437

  6. Role of PU.1 in MHC Class II Expression via CIITA Transcription in Plasmacytoid Dendritic Cells

    PubMed Central

    Miura, Ryosuke; Kasakura, Kazumi; Nakano, Nobuhiro; Hara, Mutsuko; Maeda, Keiko; Okumura, Ko; Ogawa, Hideoki; Yashiro, Takuya; Nishiyama, Chiharu

    2016-01-01

    The cofactor CIITA is a master regulator of MHC class II expression and several transcription factors regulating the cell type-specific expression of CIITA have been identified. Although the MHC class II expression in plasmacytoid dendritic cells (pDCs) is also mediated by CIITA, the transcription factors involved in the CIITA expression in pDCs are largely unknown. In the present study, we analyzed the role of a hematopoietic lineage-specific transcription factor, PU.1, in CIITA transcription in pDCs. The introduction of PU.1 siRNA into mouse pDCs and a human pDC cell line, CAL-1, reduced the mRNA levels of MHC class II and CIITA. When the binding of PU.1 to the 3rd promoter of CIITA (pIII) in CAL-1 and mouse pDCs was analyzed by a chromatin immunoprecipitation assay, a significant amount of PU.1 binding to the pIII was detected, which was definitely decreased in PU.1 siRNA-transfected cells. Reporter assays showed that PU.1 knockdown reduced the pIII promoter activity and that three Ets-motifs in the human pIII promoter were candidates of cis-enhancing elements. By electrophoretic mobility shift assays, it was confirmed that two Ets-motifs, GGAA (-181/-178) and AGAA (-114/-111), among three candidates, were directly bound with PU.1. When mouse pDCs and CAL-1 cells were stimulated by GM-CSF, mRNA levels of PU.1, pIII-driven CIITA, total CIITA, MHC class II, and the amount of PU.1 binding to pIII were significantly increased. The GM-CSF-mediated up-regulation of these mRNAs was canceled in PU.1 siRNA-introduced cells. Taking these results together, we conclude that PU.1 transactivates the pIII through direct binding to Ets-motifs in the promoter in pDCs. PMID:27105023

  7. Equine bone marrow-derived mesenchymal stromal cells are heterogeneous in MHC class II expression and capable of inciting an immune response in vitro

    PubMed Central

    2014-01-01

    Introduction The horse is a valuable species to assess the effect of allogeneic mesenchymal stromal cells (MSCs) in regenerative treatments. No studies to date have examined recipient response to major histocompatibility complex (MHC)-mismatched equine MSCs. The purposes of this study were to immunophenotype MSCs from horses of known MHC haplotype and to compare the immunogenicity of MSCs with differing MHC class II expression. Methods MSCs and peripheral blood leukocytes (PBLs) were obtained from Thoroughbred horses (n = 10) of known MHC haplotype (ELA-A2, -A3, and -A9 homozygotes). MSCs were cultured through P8; cells from each passage (P2 to P8) were cryopreserved until used. Immunophenotyping of MHC class I and II, CD44, CD29, CD90, LFA-1, and CD45RB was performed by using flow cytometry. Tri-lineage differentiation assays were performed to confirm MSC multipotency. Recombinant equine IFN-γ was used to stimulate MHC class II negative MSCs in culture, after which expression of MHC class II was re-examined. To assess the ability of MHC class II negative or positive MSCs to stimulate an immune response, modified one-way mixed leukocyte reactions (MLRs) were performed by using MHC-matched and mismatched responder PBLs and stimulator PBLs or MSCs. Proliferation of gated CFSE-labeled CD3+ responder T cells was evaluated via CFSE attenuation by using flow cytometry and reported as the number of cells in the proliferating T-cell gate. Results MSCs varied widely in MHC class II expression despite being homogenous in terms of “stemness” marker expression and ability to undergo trilineage differentiation. Stimulation of MHC class II negative MSCs with IFN-γ resulted in markedly increased expression of MHC class II. MLR results revealed that MHC-mismatched MHC class II-positive MSCs caused significantly increased responder T-cell proliferation compared with MHC-mismatched MHC class II-negative and MHC-matched MSCs, and equivalent to that of the positive control of

  8. Differential MHC class II expression on human peripheral blood monocytes and dendritic cells.

    PubMed Central

    Brooks, C F; Moore, M

    1988-01-01

    Both monocytes (MO) and dendritic cells (DC) in human peripheral blood are of a plastic-adherent nature. The expression of the MHC class II sublocus products HLA-DP, -DQ and -DR on human peripheral blood transiently adherent cells (TA) was examined by an immunocytochemical staining technique. While most TA showed strong expression of molecules of the HLA-DR subtype, only a small proportion of cells (2-6%) showed strong HLA-DP or -DQ positivity. This strong expression of the HLA-DP and HLA-DQ sublocus products by a subset of TA was seen only after short-term culture; freshly isolated cells expressed comparatively low levels of these molecules. Enrichment for Fc receptor-negative or low-density cells from TA produced populations with strong HLA-DQ and -DP expression. Such co-enrichment of the strongly HLA-DQ+ and strongly HLA-DP+ cells suggests that the same cells express high levels of both types of MHC class II molecule. Immunocytochemical analysis of TA indicated that the strongly HLA-DQ+ cells, at least, were only weakly or non-reactive with the MO-specific monoclonal antibodies OKM1, UCHM1, MO2 and EB11. In addition, strongly HLA-DQ- or -DP-positive cells were poorly phagocytic in comparison with the majority of adherent cells. The apparent FcR-negative, low-density and weakly phagocytic nature of the strongly HLA-DQ/DP+ cells, combined with their lack of reactivity with several MO-specific antibodies, suggests that they may represent the DC component of TA. Such strong HLA-DQ/DP expression by DC may aid their positive identification in human peripheral blood and may be of relevance to DC function in antigen presentation. Images Figure 1 PMID:3350576

  9. Absence of nonhematopoietic MHC class II expression protects mice from experimental autoimmune myocarditis.

    PubMed

    Thelemann, Christoph; Haller, Sergio; Blyszczuk, Przemyslaw; Kania, Gabriela; Rosa, Muriel; Eriksson, Urs; Rotman, Samuel; Reith, Walter; Acha-Orbea, Hans

    2016-03-01

    Experimental autoimmune myocarditis (EAM) is a CD4(+) T-cell-mediated model of human inflammatory dilated cardiomyopathies. Heart-specific CD4(+) T-cell activation is dependent on autoantigens presented by MHC class II (MHCII) molecules expressed on professional APCs. In this study, we addressed the role of inflammation-induced MHCII expression by cardiac nonhematopoietic cells on EAM development. EAM was induced in susceptible mice lacking inducible expression of MHCII molecules on all nonhematopoietic cells (pIV-/- K14 class II transactivator (CIITA) transgenic (Tg) mice) by immunization with α-myosin heavy chain peptide in CFA. Lack of inducible nonhematopoietic MHCII expression in pIV-/- K14 CIITA Tg mice conferred EAM resistance. In contrast, cardiac pathology was induced in WT and heterozygous mice, and correlated with elevated cardiac endothelial MHCII expression. Control mice with myocarditis displayed an increase in infiltrating CD4(+) T cells and in expression of IFN-γ, which is the major driver of nonhematopoietic MHCII expression. Mechanistically, IFN-γ neutralization in WT mice shortly before disease onset resulted in reduced cardiac MHCII expression and pathology. These findings reveal a previously overlooked contribution of IFN-γ to induce endothelial MHCII expression in the heart and to progress cardiac pathology during myocarditis. PMID:26621778

  10. Organizing MHC Class II Presentation

    PubMed Central

    Fooksman, David R.

    2014-01-01

    Major histocompatibility complex (MHC) class II molecules are ligands for CD4+ T cells and are critical for initiating the adaptive immune response. This review is focused on what is currently known about MHC class II organization at the plasma membrane of antigen presenting cells and how this affects antigen presentation to T cells. The organization and diffusion of class II molecules have been measured by a variety of biochemical and microscopic techniques. Membrane lipids and other proteins have been implicated in MHC class II organization and function. However, when compared with the organization of MHC class I or TCR complexes, much less is known about MHC class II. Since clustering of T cell receptors occurs during activation, the organization of MHC molecules prior to recognition and during synapse formation may be critical for antigen presentation. PMID:24782863

  11. Presence of specific MHC Class II expressed alleles associates with clinical disease in ovine progressive pneumonia virus (OPPV) infected sheep

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A genetic tool hypothesized to predict which OPPV infected sheep will progress to debilitating clinical disease is MHC Class II Ovis aries (Ovar)-DRB1. Previously, fifteen Ovar-DRB1 beta 1 expressed alleles were identified in a ewe-lamb flock of 32 originating from an Idaho flock using RT-PCR, clon...

  12. Expressed MHC class II genes in sea otters (Enhydra lutris) from geographically disparate populations

    USGS Publications Warehouse

    Bowen, L.; Aldridge, B.M.; Miles, A.K.; Stott, J.L.

    2006-01-01

    The major histocompatibility complex (MHC) is central to maintaining the immunologic vigor of individuals and populations. Classical MHC class II genes were targeted for partial sequencing in sea otters (Enhydra lutris) from populations in California, Washington, and Alaska. Sequences derived from sea otter peripheral blood leukocyte mRNAs were similar to those classified as DQA, DQB, DRA, and DRB in other species. Comparisons of the derived amino acid compositions supported the classification of these as functional molecules from at least one DQA, DQB, and DRA locus and at least two DRB loci. While limited in scope, phylogenetic analysis of the DRB peptide-binding region suggested the possible existence of distinct clades demarcated by geographic region. These preliminary findings support the need for additional MHC gene sequencing and expansion to a comprehensive study targeting additional otters. ?? 2006 Blackwell Munksgaard.

  13. Salmonella Typhimurium induces SPI-1 and SPI-2 regulated and strain dependent downregulation of MHC II expression on porcine alveolar macrophages

    PubMed Central

    2012-01-01

    Foodborne salmonellosis is one of the most important bacterial zoonotic diseases worldwide. Salmonella Typhimurium is the serovar most frequently isolated from persistently infected slaughter pigs in Europe. Circumvention of the host’s immune system by Salmonella might contribute to persistent infection of pigs. In the present study, we found that Salmonella Typhimurium strain 112910a specifically downregulated MHC II, but not MHC I, expression on porcine alveolar macrophages in a Salmonella pathogenicity island (SPI)-1 and SPI-2 dependent way. Salmonella induced downregulation of MHC II expression and intracellular proliferation of Salmonella in macrophages were significantly impaired after opsonization with Salmonella specific antibodies prior to inoculation. Furthermore, the capacity to downregulate MHC II expression on macrophages differed significantly among Salmonella strains, independently of strain specific differences in invasion capacity, Salmonella induced cytotoxicity and altered macrophage activation status. The fact that strain specific differences in MHC II downregulation did not correlate with the extent of in vitro SPI-1 or SPI-2 gene expression indicates that other factors are involved in MHC II downregulation as well. Since Salmonella strain dependent interference with the pig’s immune response through downregulation of MHC II expression might indicate that certain Salmonella strains are more likely to escape serological detection, our findings are of major interest for Salmonella monitoring programs primarily based on serology. PMID:22694285

  14. Interplay among coactivator-associated arginine methyltransferase 1, CBP, and CIITA in IFN-gamma-inducible MHC-II gene expression.

    PubMed

    Zika, Eleni; Fauquier, Lucas; Vandel, Laurence; Ting, Jenny P-Y

    2005-11-01

    Class II major histocompatibility (MHC-II) genes are prototype targets of IFN-gamma. IFN-gamma activates the expression of the non-DNA-binding master regulator of MHC-II, class II transactivator (CIITA), which is crucial for enhanceosome formation and gene activation. This report shows the importance of the histone methyltransferase, coactivator-associated arginine methyltransferase (CARM1/PRMT4), during IFN-gamma-induced MHC-II gene activation. It also demonstrates the coordinated regulation of CIITA, CARM1, and the acetyltransferase cyclic-AMP response element binding (CREB)-binding protein (CBP) during this process. CARM1 synergizes with CIITA in activating MHC-II transcription and synergy is abrogated when an arginine methyltransferase-defective CARM1 mutant is used. Protein-arginine methyltransferase 1 has much less effect on MHC-II transcription. Specific RNA interference reduced CARM1 expression as well as MHC-II expression. The recruitment of CARM1 to the promoter requires endogenous CIITA and results in methylation of histone H3-R17; hence, CIITA is an upstream regulator of histone methylation. Previous work has shown that CARM1 can methylate CBP at three arginine residues. Using wild-type CBP and a mutant of CBP lacking the CARM1-targeted arginine residues (R3A), we show that arginine methylation of CBP is required for IFN-gamma induction of MHC-II. A kinetic analysis shows that CIITA, CARM1, and H3-R17 methylation all precede CBP loading on the MHC-II promoter during IFN-gamma treatment. These results suggest functional and temporal relationships among CIITA, CARM1, and CBP for IFN-gamma induction of MHC-II. PMID:16254053

  15. Melanoma-specific MHC-II expression represents a tumour-autonomous phenotype and predicts response to anti-PD-1/PD-L1 therapy

    PubMed Central

    Johnson, Douglas B.; Estrada, Monica V.; Salgado, Roberto; Sanchez, Violeta; Doxie, Deon B.; Opalenik, Susan R.; Vilgelm, Anna E.; Feld, Emily; Johnson, Adam S.; Greenplate, Allison R.; Sanders, Melinda E.; Lovly, Christine M.; Frederick, Dennie T.; Kelley, Mark C.; Richmond, Ann; Irish, Jonathan M.; Shyr, Yu; Sullivan, Ryan J.; Puzanov, Igor; Sosman, Jeffrey A.; Balko, Justin M.

    2016-01-01

    Anti-PD-1 therapy yields objective clinical responses in 30–40% of advanced melanoma patients. Since most patients do not respond, predictive biomarkers to guide treatment selection are needed. We hypothesize that MHC-I/II expression is required for tumour antigen presentation and may predict anti-PD-1 therapy response. In this study, across 60 melanoma cell lines, we find bimodal expression patterns of MHC-II, while MHC-I expression was ubiquitous. A unique subset of melanomas are capable of expressing MHC-II under basal or IFNγ-stimulated conditions. Using pathway analysis, we show that MHC-II(+) cell lines demonstrate signatures of ‘PD-1 signalling', ‘allograft rejection' and ‘T-cell receptor signalling', among others. In two independent cohorts of anti-PD-1-treated melanoma patients, MHC-II positivity on tumour cells is associated with therapeutic response, progression-free and overall survival, as well as CD4+ and CD8+ tumour infiltrate. MHC-II+ tumours can be identified by melanoma-specific immunohistochemistry using commercially available antibodies for HLA-DR to improve anti-PD-1 patient selection. PMID:26822383

  16. Human umbilical cord mesenchymal stromal cells suppress MHC class II expression on rat vascular endothelium and prolong survival time of cardiac allograft

    PubMed Central

    Qiu, Ying; Yun, Mark M; Han, Xia; Zhao, Ruidong; Zhou, Erxia; Yun, Sheng

    2014-01-01

    Background: Human umbilical cord mesenchymal stromal cells (UC-MSCs) have low immunogenicity and immune regulation. To investigate immunomodulatory effects of human UC-MSCs on MHC class II expression and allograft, we transplanted heart of transgenic rats with MHC class II expression on vascular endothelium. Methods: UC-MSCs were obtained from human umbilical cords and confirmed with flow cytometry analysis. Transgenic rat line was established using the construct of human MHC class II transactivator gene (CIITA) under mouse ICAM-2 promoter control. The induced MHC class II expression on transgenic rat vascular endothelial cells (VECs) was assessed with immunohistological staining. And the survival time of cardiac allograft was compared between the recipients with and without UC-MSC transfusion. Results: Flow cytometry confirmed that the human UC-MSCs were positive for CD29, CD44, CD73, CD90, CD105, CD271, and negative for CD34 and HLA-DR. Repeated infusion of human UC-MSCs reduced MHC class II expression on vascular endothelia of transplanted hearts, and increased survival time of allograft. The UC-MSCs increased regulatory cytokines IL10, transforming growth factor (TGF)-β1 and suppressed proinflammatory cytokines IL2 and IFN-γ in vivo. The UC-MSC culture supernatant had similar effects on cytokine expression, and decreased lymphocyte proliferation in vitro. Conclusions: Repeated transfusion of the human UC-MSCs reduced MHC class II expression on vascular endothelia and prolonged the survival time of rat cardiac allograft. PMID:25126177

  17. Heligmosomoides polygyrus infection is associated with lower MHC class II gene expression in Apodemus flavicollis: indication for immune suppression?

    PubMed

    Axtner, Jan; Sommer, Simone

    2011-12-01

    Due to their key role in recognizing foreign antigens and triggering the subsequent immune response the genes of the major histocompatibility complex (MHC) provide a potential target for parasites to attack in order to evade detection and expulsion from the host. A diminished MHC gene expression results in less activated T cells and might serve as a gateway for pathogens and parasites. Some parasites are suspected to be immune suppressors and promote co-infections of other parasites even in other parts of the body. In our study we found indications that the gut dwelling nematode Heligmosomoides polygyrus might exert a systemic immunosuppressive effect in yellow-necked mice (Apodemus flavicollis). The amount of hepatic MHC class II DRB gene RNA transcripts in infected mice was negatively associated with infection intensity with H. polygyrus. The hepatic expression of immunosuppressive cytokines, such as transforming growth factor β and interleukin 10 was not associated with H. polygyrus infection. We did not find direct positive associations of H. polygyrus with other helminth species. But the prevalence and infection intensity of the nematodes Syphacia stroma and Trichuris muris were higher in multiple infected individuals. Furthermore, our data indicated antagonistic effects in the helminth community of A. flavicollis as cestode infection correlated negatively with H. polygyrus and helminth species richness. Our study shows that expression analyses of immune relevant genes can also be performed in wildlife, opening new aspects and possibilities for future ecological and evolutionary research. PMID:21983561

  18. Analysis of MHC class I and II expression in relation to presence of HPV genotypes in premalignant and malignant cervical lesions.

    PubMed Central

    Cromme, F. V.; Meijer, C. J.; Snijders, P. J.; Uyterlinde, A.; Kenemans, P.; Helmerhorst, T.; Stern, P. L.; van den Brule, A. J.; Walboomers, J. M.

    1993-01-01

    Cervical intraepithelial neoplasia (CIN) grades I to III lesions (n = 94) and squamous cell carcinomas of the uterine cervix (n = 27) were analysed for MHC class I and II expression and presence of HPV genotypes. MHC class I and II expression was studied by immunohistochemistry and HPV typing was performed by general primer- and type-specific primer mediated PCR (GP/TS PCR). Both techniques were performed on paraffin embedded tissue sections. Results show disturbed MHC class I heavy chain expression in CIN I to CIN III, as well as in cervical carcinomas. Upregulated MHC class II expression on dysplastic epithelial cells was also found in the different CIN groups and carcinomas. Prevalence of HPV genotypes increased with the severity of the lesion, mainly due to the contribution of the HPV types 16 and 18. No correlation could be established between the presence of specific HPV genotypes and any MHC expression pattern in the different CIN groups or cervical carcinomas. In some cases these data were confirmed by RNA in situ hybridisation showing HPV 16 E7 transcripts in the same dysplastic/neoplastic cells from which MHC status was determined. The results indicate that local differences may exist in the type of cellular immune response to HPV induced lesions. Images Figure 1 Figure 2 Figure 4 PMID:8390286

  19. ZBTB32 is an early repressor of the class II transactivator and MHC class II gene expression during B cell differentiation to plasma cells1

    PubMed Central

    Yoon, Hyesuk; Scharer, Christopher D.; Majumder, Parimal; Davis, Carl W.; Butler, Royce; Zinzow-Kramer, Wendy; Skountzou, Ioanna; Koutsonanos, Dimitrios G.; Ahmed, Rafi; Boss, Jeremy M.

    2012-01-01

    The MHC class II transactivator (CIITA) and MHC class II expression is silenced during the differentiation of B cells to plasma cells. When B cell differentiation is carried out ex vivo, CIITA silencing occurs rapidly but the factors contributing to this event are not known. ZBTB32, also known as repressor of GATA3, was identified as an early repressor of CIITA in an ex vivo plasma cell differentiation model. ZBTB32 activity occurred at a time when Blimp-1, the regulator of plasma cell fate and suppressor of CIITA, was minimally induced. Ectopic expression of ZBTB32 suppressed CIITA and I-A gene expression in B cells. ShRNA depletion of ZBTB32 in a plasma cell line resulted in reexpression of CIITA and I-A. Compared to conditional Blimp-1 knock out and wild-type B cells, B cells from ZBTB32/ROG-knock out mice displayed delayed kinetics in silencing CIITA during ex vivo plasma cell differentiation. ZBTB32 was found to bind to the CIITA gene, suggesting that ZBTB32 directly regulates CIITA. Lastly, ZBTB32 and Blimp-1 coimmunoprecipitated, suggesting that the two repressors may ultimately function together to silence CIITA expression. These results introduce ZBTB32 as a novel regulator of MHC-II gene expression and a potential regulatory partner of Blimp-1 in repressing gene expression. PMID:22851713

  20. Cohesin regulates major histocompatibility complex class II genes through interactions with MHC-II insulators1

    PubMed Central

    Majumder, Parimal; Boss, Jeremy M.

    2011-01-01

    Cohesin is a multiprotein ringed complex that is most well known for its role in stabilizing the association of sister chromatids between S phase and M. More recently cohesin was found to be associated with transcriptional insulators, elements that are associated with the organization of chromatin into regulatory domains. The human major histocompatibility complex class II (MHC-II) locuscontains ten intergenic elements, termed MHC-II insulators, which bind the transcriptional insulator protein CCCTC transcription factor (CTCF). MHC-II insulators interact with each other forming a base architecture of discrete loops and potential regulatory domains. When MHC-II genes are expressed, their proximal promoter regulatory regions reorganize to the foci established by the interacting MHC-II insulators. MHC-II insulators also bind cohesin, but the functional role of cohesin in regulating this system is not known. Here we show that the binding of cohesin to MHC-II insulators occurred irrespective of MHC-II expression but was required for optimal expression of the HLA-DR and HLA-DQ genes. In a DNA dependent manner, cohesin subunits interacted with CTCF and the MHC-II specific transcription factors RFX and CIITA. Intriguingly, cohesin subunits were important for DNA looping interactions between the HLA-DRA promoter region and a 5’ MHC-II insulator but were not required for interactions between the MHC-II insulators themselves. This latter observation introduces cohesin as a regulator of MHC-II expression by initiating or stabilizing MHC-II promoter regulatory element interactions with the MHC-II insulator elements; events which are required for maximal MHC-II transcription. PMID:21911605

  1. Non-neutral evolution and reciprocal monophyly of two expressed Mhc class II B genes in Leach's storm-petrel.

    PubMed

    Dearborn, Donald C; Gager, Andrea B; Gilmour, Morgan E; McArthur, Andrew G; Hinerfeld, Douglas A; Mauck, Robert A

    2015-02-01

    The major histocompatibility complex (Mhc) is subject to pathogen-mediated balancing selection and can link natural selection with mate choice. We characterized two Mhc class II B loci in Leach's storm-petrel, Oceanodroma leucorhoa, focusing on exon 2 which encodes the portion of the protein that binds pathogen peptides. We amplified and sequenced exon 2 with locus-specific nested PCR and Illumina MiSeq using individually barcoded primers. Repeat genotyping of 78 single-locus genotypes produced identical results in 77 cases (98.7%). Sequencing of messenger RNA (mRNA) from three birds confirmed expression of both loci, consistent with the observed absence of stop codons or frameshifts in all alleles. In 48 birds, we found 9 and 12 alleles at the two loci, respectively, and all 21 alleles translated to unique amino acid sequences. Unlike many studies of duplicated Mhc genes, alleles of the two loci clustered into monophyletic groups. Consistent with this phylogenetic result, interlocus gene conversion appears to have affected only two short fragments of the exon. As predicted under a paradigm of pathogen-mediated selection, comparison of synonymous and non-synonymous substitution rates found evidence of a history of positive selection at putative peptide binding sites. Overall, the results suggest that the gene duplication event leading to these two loci is not recent and that point mutations and positive selection on the peptide binding sites may be the predominant forces acting on these genes. Characterization of these loci sets the stage for population-level work on the evolutionary ecology of Mhc in this species. PMID:25416539

  2. MHC-class-II are expressed in a subpopulation of human neural stem cells in vitro in an IFNγ-independent fashion and during development.

    PubMed

    Vagaska, B; New, S E P; Alvarez-Gonzalez, C; D'Acquisto, F; Gomez, S G; Bulstrode, N W; Madrigal, A; Ferretti, P

    2016-01-01

    Expression of major histocompatibility antigens class-2 (MHC-II) under non-inflammatory conditions is not usually associated with the nervous system. Comparative analysis of immunogenicity of human embryonic/fetal brain-derived neural stem cells (hNSCs) and human mesenchymal stem cells with neurogenic potential from umbilical cord (UC-MSCs) and paediatric adipose tissue (ADSCs), while highlighting differences in their immunogenicity, led us to discover subsets of neural cells co-expressing the neural marker SOX2 and MHC-II antigen in vivo during human CNS development. MHC-II proteins in hNSCs are functional, and differently regulated upon differentiation along different lineages. Mimicking an inflammatory response using the inflammatory cytokine IFNγ induced MHC-II up-regulation in both astrocytes and hNSCs, but not in UC-MSCs and ADSCs, either undifferentiated or differentiated, though IFNγ receptor expression was comparable. Together, hypoimmunogenicity of both UC-MSCs and ADSCs supports their suitability for allogeneic therapy, while significant immunogenicity of hNSCs and their progeny may at least in part underlie negative effects reported in some patients following embryonic neural cell grafts. Crucially, we show for the first time that MHC-II expression in developing human brains is not restricted to microglia as previously suggested, but is present in discrete subsets of neural progenitors and appears to be regulated independently of inflammatory stimuli. PMID:27080443

  3. MHC-class-II are expressed in a subpopulation of human neural stem cells in vitro in an IFNγ–independent fashion and during development

    PubMed Central

    Vagaska, B.; New, S. E. P.; Alvarez-Gonzalez, C.; D’Acquisto, F.; Gomez, S. G.; Bulstrode, N. W.; Madrigal, A.; Ferretti, P.

    2016-01-01

    Expression of major histocompatibility antigens class-2 (MHC-II) under non-inflammatory conditions is not usually associated with the nervous system. Comparative analysis of immunogenicity of human embryonic/fetal brain-derived neural stem cells (hNSCs) and human mesenchymal stem cells with neurogenic potential from umbilical cord (UC-MSCs) and paediatric adipose tissue (ADSCs), while highlighting differences in their immunogenicity, led us to discover subsets of neural cells co-expressing the neural marker SOX2 and MHC-II antigen in vivo during human CNS development. MHC-II proteins in hNSCs are functional, and differently regulated upon differentiation along different lineages. Mimicking an inflammatory response using the inflammatory cytokine IFNγ induced MHC-II up-regulation in both astrocytes and hNSCs, but not in UC-MSCs and ADSCs, either undifferentiated or differentiated, though IFNγ receptor expression was comparable. Together, hypoimmunogenicity of both UC-MSCs and ADSCs supports their suitability for allogeneic therapy, while significant immunogenicity of hNSCs and their progeny may at least in part underlie negative effects reported in some patients following embryonic neural cell grafts. Crucially, we show for the first time that MHC-II expression in developing human brains is not restricted to microglia as previously suggested, but is present in discrete subsets of neural progenitors and appears to be regulated independently of inflammatory stimuli. PMID:27080443

  4. HLA-G and MHC Class II Protein Expression in Diffuse Large B-Cell Lymphoma.

    PubMed

    Jesionek-Kupnicka, Dorota; Bojo, Marcin; Prochorec-Sobieszek, Monika; Szumera-Ciećkiewicz, Anna; Jabłońska, Joanna; Kalinka-Warzocha, Ewa; Kordek, Radzisław; Młynarski, Wojciech; Robak, Tadeusz; Warzocha, Krzysztof; Lech-Maranda, Ewa

    2016-06-01

    The expression of human leukocyte antigen-G (HLA-G) and HLA class II protein was studied by immunohistochemical staining of lymph nodes from 148 patients with diffuse large B-cell lymphoma (DLBCL) and related to the clinical course of the disease. Negative HLA-G expression was associated with a lower probability of achieving a complete remission (p = 0.04). Patients with negative HLA-G expression tended towards a lower 3-year overall survival (OS) rate compared to those with positive expression of HLA-G (p = 0.08). When restricting the analysis to patients receiving chemotherapy with rituximab, the estimated 3-year OS rate of patients with positive HLA-G expression was 73.3 % compared with 47.5 % (p = 0.03) in those with negative expression. Patients with negative HLA class II expression presented a lower 3-year OS rate compared to subjects with positive expression (p = 0.04). The loss of HLA class II expression (p = 0.05) and belonging to the intermediate high/high IPI risk group (p = 0.001) independently increased the risk of death. HLA class II expression also retained its prognostic value in patients receiving rituximab; the 3-year OS rate was 65.3 % in patients with positive HLA class II expression versus 29.6 % (p = 0.04) in subjects that had loss of HLA class II expression. To our knowledge, for the first time, the expression of HLA-G protein in DLBCL and its association with the clinical course of the disease was demonstrated. Moreover, the link between losing HLA class II protein expression and poor survival of patients treated with immunochemotherapy was confirmed. PMID:26667793

  5. Loss in CD4 T-cell responses to multiple epitopes in influenza due to expression of one additional MHC class II molecule in the host

    PubMed Central

    Nayak, Jennifer L; Sant, Andrea J

    2012-01-01

    An understanding of factors controlling CD4 T-cell immunodominance is needed to pursue CD4 T-cell epitope-driven vaccine design, yet our understanding of this in humans is limited by the complexity of potential MHC class II molecule expression. In the studies described here, we took advantage of genetically restricted, well-defined mouse strains to better understand the effect of increasing MHC class II molecule diversity on the CD4 T-cell repertoire and the resulting anti-influenza immunodominance hierarchy. Interferon-γ ELISPOT assays were implemented to directly quantify CD4 T-cell responses to I-Ab and I-As restricted peptide epitopes following primary influenza virus infection in parental and F1 hybrid strains. We found striking and asymmetric declines in the magnitude of many peptide-specific responses in F1 animals. These declines could not be accounted for by the lower surface density of MHC class II on the cell or by antigen-presenting cells failing to stimulate T cells with lower avidity T-cell receptors. Given the large diversity of MHC class II expressed in humans, these findings have important implications for the rational design of peptide-based vaccines that are based on the premise that CD4 T-cell epitope specificity can be predicted by a simple cataloguing of an individual’s MHC class II genotype. PMID:22747522

  6. The melting pot of the MHC II peptidome.

    PubMed

    Stern, Lawrence J; Santambrogio, Laura

    2016-06-01

    Recent advances in mass spectrometry technology have facilitated detailed examination of MHC-II immunopeptidomes, for example the repertoires of peptides bound to MHC-II molecules expressed in antigen presenting cells. These studies have deepened our view of MHC-II presentation. Other studies have broadened our view of pathways leading up to peptide loading. Here we review these recent studies in the context of earlier work on conventional and non-conventional MHC-II processing. The message that emerges is that sources of antigen beyond conventional endosomal processing of endocytosed proteins are important for generation of cellular immune responses to pathogens and maintenance of central and peripheral tolerance. The multiplicity of pathways results in a broad MHC II immunopeptidome that conveys the sampled environment to patrolling T cells. PMID:27018930

  7. Characterization of expressed class II MHC sequences in the banner-tailed kangaroo rat (Dipodomys spectabilis) reveals multiple DRB loci.

    PubMed

    Busch, Joseph D; Waser, Peter M; DeWoody, J Andrew

    2008-11-01

    Genes of the major histocompatibility complex (MHC) are exceptionally polymorphic due to the combined effects of natural and sexual selection. Most research in wild populations has focused on the second exon of a single class II locus (DRB), but complete gene sequences can provide an illuminating backdrop for studies of intragenic selection, recombination, and organization. To this end, we characterized class II loci in the banner-tailed kangaroo rat (Dipodomys spectabilis). Seven DRB-like sequences (provisionally named MhcDisp-DRB*01 through *07) were isolated from spleen cDNA and most likely comprise > or =5 loci; this multiformity is quite unlike the situation in muroid rodents such as Mus, Rattus, and Peromyscus. In silico translation revealed the presence of important structural residues for glycosylation sites, salt bonds, and CD4+ T-cell recognition. Amino-acid distances varied widely among the seven sequences (2-34%). Nuclear DNA sequences from the Disp-DRB*07 locus (approximately 10 kb) revealed a conventional exon/intron structure as well as a number of microsatellites and short interspersed nuclear elements (B4, Alu, and IDL-Geo subfamilies). Rates of nucleotide substitution at Disp-DRB*07 are similar in both exons and introns (pi = 0.015 and 0.012, respectively), which suggests relaxed selection and may indicate that this locus is an expressed pseudogene. Finally, we performed BLASTn searches against Dipodomys ordii genomic sequences (unassembled reads) and find 90-97% nucleotide similarity between the two kangaroo rat species. Collectively, these data suggest that class II diversity in heteromyid rodents is based on polylocism and departs from the muroid architecture. PMID:18836711

  8. FOXP1 suppresses immune response signatures and MHC class II expression in activated B-cell-like diffuse large B-cell lymphomas.

    PubMed

    Brown, P J; Wong, K K; Felce, S L; Lyne, L; Spearman, H; Soilleux, E J; Pedersen, L M; Møller, M B; Green, T M; Gascoyne, D M; Banham, A H

    2016-03-01

    The FOXP1 (forkhead box P1) transcription factor is a marker of poor prognosis in diffuse large B-cell lymphoma (DLBCL). Here microarray analysis of FOXP1-silenced DLBCL cell lines identified differential regulation of immune response signatures and major histocompatibility complex class II (MHC II) genes as some of the most significant differences between germinal center B-cell (GCB)-like DLBCL with full-length FOXP1 protein expression versus activated B-cell (ABC)-like DLBCL expressing predominantly short FOXP1 isoforms. In an independent primary DLBCL microarray data set, multiple MHC II genes, including human leukocyte antigen DR alpha chain (HLA-DRA), were inversely correlated with FOXP1 transcript expression (P<0.05). FOXP1 knockdown in ABC-DLBCL cells led to increased cell-surface expression of HLA-DRA and CD74. In R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone)-treated DLBCL patients (n=150), reduced HLA-DRA (<90% frequency) expression correlated with inferior overall survival (P=0.0003) and progression-free survival (P=0.0012) and with non-GCB subtype stratified by the Hans, Choi or Visco-Young algorithms (all P<0.01). In non-GCB DLBCL cases with <90% HLA-DRA, there was an inverse correlation with the frequency (P=0.0456) and intensity (P=0.0349) of FOXP1 expression. We propose that FOXP1 represents a novel regulator of genes targeted by the class II MHC transactivator CIITA (MHC II and CD74) and therapeutically targeting the FOXP1 pathway may improve antigen presentation and immune surveillance in high-risk DLBCL patients. PMID:26500140

  9. FOXP1 suppresses immune response signatures and MHC class II expression in activated B-cell-like diffuse large B-cell lymphomas

    PubMed Central

    Brown, P J; Wong, K K; Felce, S L; Lyne, L; Spearman, H; Soilleux, E J; Pedersen, L M; Møller, M B; Green, T M; Gascoyne, D M; Banham, A H

    2016-01-01

    The FOXP1 (forkhead box P1) transcription factor is a marker of poor prognosis in diffuse large B-cell lymphoma (DLBCL). Here microarray analysis of FOXP1-silenced DLBCL cell lines identified differential regulation of immune response signatures and major histocompatibility complex class II (MHC II) genes as some of the most significant differences between germinal center B-cell (GCB)-like DLBCL with full-length FOXP1 protein expression versus activated B-cell (ABC)-like DLBCL expressing predominantly short FOXP1 isoforms. In an independent primary DLBCL microarray data set, multiple MHC II genes, including human leukocyte antigen DR alpha chain (HLA-DRA), were inversely correlated with FOXP1 transcript expression (P<0.05). FOXP1 knockdown in ABC-DLBCL cells led to increased cell-surface expression of HLA-DRA and CD74. In R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone)-treated DLBCL patients (n=150), reduced HLA-DRA (<90% frequency) expression correlated with inferior overall survival (P=0.0003) and progression-free survival (P=0.0012) and with non-GCB subtype stratified by the Hans, Choi or Visco–Young algorithms (all P<0.01). In non-GCB DLBCL cases with <90% HLA-DRA, there was an inverse correlation with the frequency (P=0.0456) and intensity (P=0.0349) of FOXP1 expression. We propose that FOXP1 represents a novel regulator of genes targeted by the class II MHC transactivator CIITA (MHC II and CD74) and therapeutically targeting the FOXP1 pathway may improve antigen presentation and immune surveillance in high-risk DLBCL patients. PMID:26500140

  10. Quantifying Significance of MHC II Residues.

    PubMed

    Fan, Ying; Lu, Ruoshui; Wang, Lusheng; Andreatta, Massimo; Li, Shuai Cheng

    2014-01-01

    The major histocompatibility complex (MHC), a cell-surface protein mediating immune recognition, plays important roles in the immune response system of all higher vertebrates. MHC molecules are highly polymorphic and they are grouped into serotypes according to the specificity of the response. It is a common belief that a protein sequence determines its three dimensional structure and function. Hence, the protein sequence determines the serotype. Residues play different levels of importance. In this paper, we quantify the residue significance with the available serotype information. Knowing the significance of the residues will deepen our understanding of the MHC molecules and yield us a concise representation of the molecules. In this paper we propose a linear programming-based approach to find significant residue positions as well as quantifying their significance in MHC II DR molecules. Among all the residues in MHC II DR molecules, 18 positions are of particular significance, which is consistent with the literature on MHC binding sites, and succinct pseudo-sequences appear to be adequate to capture the whole sequence features. When the result is used for classification of MHC molecules with serotype assigned by WHO, a 98.4 percent prediction performance is achieved. The methods have been implemented in java (http://code.google.com/p/quassi/). PMID:26355503

  11. Upregulation and induction of surface antigens with special reference to MHC class II expression in microglia in postnatal rat brain following intravenous or intraperitoneal injections of lipopolysaccharide.

    PubMed Central

    Xu, J; Ling, E A

    1994-01-01

    The effects of bacterial lipopolysaccharide (LPS) on the expression of surface antigens including major histocompatibility complex (MHC) and complement type 3 (CR3) receptors on microglial cells in the corpus callosum in postnatal rat brain were investigated. When LPS was injected intravenously (i.v.) in 1-d-old rats, the immunostaining of callosal amoeboid microglial cells with OX-18 directed against MHC class I antigen was enhanced 24 h after the injection in comparison with the controls. The expression of MHC class II (Ia) antigen on the same cell type as shown by its immunoreactivity with OX-6 was also elicited especially after 2 intraperitoneal (i.p.) injections of LPS. Thus 7 d after a single i.p. injection of LPS into 1-d-old rats, only a few OX-6 positive cells showing a moderate staining reaction were observed in the corpus callosum. The immunoreactivity diminished 14 d after the injection. However, in rats receiving 2 successive i.p. injections of LPS at 1 and 4 d of age and killed 7 d after the 1st injection, a significant number of intensely stained OX-6 positive amoeboid microglial cells were observed in the corpus callosum. The expression of MHC class II antigens induced by 2 injections of LPS was sustained at least until d 14 when the callosal ramified microglial cells, known to be derived from gradual metamorphic transformation of amoeboid microglia, still exhibited intense immunoreactivity with OX-6. The effect of LPS on the expression of CR3 on amoeboid microglial cells was not obvious after a single injection, but the immunoreactivity with OX-42 was also augmented in rats given 2 i.p. administration of LPS into rats at 1 an 4 d of age. It is concluded from this study that the expression of MHC class I and class II antigens on amoeboid microglial cells in corpus callosum was upregulated and induced respectively after i.v. or i.p. injection of LPS into early postnatal rats. Although relatively fewer in number when compared with OX-18 and OX-42

  12. The arginine methyltransferase PRMT5 regulates CIITA-dependent MHC II transcription.

    PubMed

    Fan, Zhiwen; Kong, Xiaocen; Xia, Jun; Wu, Xiaoyan; Li, He; Xu, Huihui; Fang, Mingming; Xu, Yong

    2016-05-01

    Class II major histocompatibility complex (MHC II) dependent antigen presentation serves as a key step in mammalian adaptive immunity and host defense. In antigen presenting cells (e.g., macrophages), MHC II transcription can be activated by interferon gamma (IFN-γ) and mediated by class II transactivator (CIITA). The underlying epigenetic mechanism, however, is not completely understood. Here we report that following IFN-γ stimulation, symmetrically dimethylated histone H3 arginine 2 (H3R2Me2s) accumulated on the MHC II promoter along with CIITA. IFN-γ augmented expression, nuclear translocation, and promoter binding of the protein arginine methyltransferase PRMT5 in macrophages. Over-expression of PRMT5 potentiated IFN-γ induced activation of MHC II transcription in an enzyme activity-dependent manner. In contrast, PRMT5 silencing or inhibition of PRMT5 activity by methylthioadenosine (MTA) suppressed MHC II transactivation by IFN-γ. CIITA interacted with and recruited PRMT5 to the MHC II promoter and mediated the synergy between PRMT5 and ASH2/WDR5 to activate MHC II transcription. PRMT5 expression was down-regulated in senescent and H2O2-treated macrophages rendering ineffectual induction of MHC II transcription by IFN-γ. Taken together, our data reveal a pathophysiologically relevant role for PRMT5 in MHC II transactivation in macrophages. PMID:26972221

  13. MHC class II super-enhancer increases surface expression of HLA-DR and HLA-DQ and affects cytokine production in autoimmune vitiligo

    PubMed Central

    Cavalli, Giulio; Hayashi, Masahiro; Jin, Ying; Yorgov, Daniel; Santorico, Stephanie A.; Holcomb, Cherie; Rastrou, Melinda; Erlich, Henry; Tengesdal, Isak W.; Dagna, Lorenzo; Neff, C. Preston; Palmer, Brent E.; Spritz, Richard A.; Dinarello, Charles A.

    2016-01-01

    Genetic risk for autoimmunity in HLA genes is most often attributed to structural specificity resulting in presentation of self-antigens. Autoimmune vitiligo is strongly associated with the MHC class II region. Here, we fine-map vitiligo MHC class II genetic risk to three SNPs only 47 bp apart, located within a predicted super-enhancer in an intergenic region between HLA-DRB1 and HLA-DQA1, localized by a genome-wide association study of 2,853 Caucasian vitiligo patients. The super-enhancer corresponds to an expression quantitative trait locus for expression of HLA-DR and HLA-DQ RNA; we observed elevated surface expression of HLA-DR (P = 0.008) and HLA-DQ (P = 0.02) on monocytes from healthy subjects homozygous for the high-risk SNP haplotype. Unexpectedly, pathogen-stimulated peripheral blood mononuclear cells from subjects homozygous for the high-risk super-enhancer haplotype exhibited greater increase in production of IFN-γ and IL-1β than cells from subjects homozygous for the low-risk haplotype. Specifically, production of IFN-γ on stimulation of dectin-1, mannose, and Toll-like receptors with Candida albicans and Staphylococcus epidermidis was 2.5- and 2.9-fold higher in high-risk subjects than in low-risk subjects, respectively (P = 0.007 and P = 0.01). Similarly, production of IL-1β was fivefold higher in high-risk subjects than in low-risk subjects (P = 0.02). Increased production of immunostimulatory cytokines in subjects carrying the high-risk haplotype may act as an “adjuvant” during the presentation of autoantigens, tying together genetic variation in the MHC with the development of autoimmunity. This study demonstrates that for risk of autoimmune vitiligo, expression level of HLA class II molecules is as or more important than antigen specificity. PMID:26787888

  14. In vitro digestion with proteases producing MHC class II ligands.

    PubMed

    Tohmé, Mira; Maschalidi, Sophia; Manoury, Bénédicte

    2013-01-01

    Proteases generate peptides that bind to MHC class II molecules to interact with a wide diversity of CD4(+) T cells. They are expressed in dedicated organelles: endosomes and lysosomes of professional antigen presenting cells (pAPCs) such as B cells, macrophages, and dendritic cells. The identification of endosomal proteases which produce antigenic peptides is important, for example, for better vaccination and to prevent autoimmune diseases. Here, we describe a panel of technics (in vitro digestion assays of protein with recombinant proteases or purified endosomes/lysosomes, T cell stimulation) to monitor the production of MHC class II ligands. PMID:23329510

  15. A novel regulatory pathway for autoimmune disease: Binding of partial MHC class II constructs to monocytes reduces CD74 expression and induces both specific and bystander T-cell tolerance

    PubMed Central

    Andrew, Shayne; Huan, Jianya; Chou, Yuan K.; Buenafe, Abigail C.; Dahan, Rony; Reiter, Yoram; Mooney, Jeffery L.; Offner, Halina; Burrows, Gregory G.

    2012-01-01

    Treatment with partial (p)MHC class II-β1α1 constructs (also referred to as recombinant T-cell receptor ligands – RTL) linked to antigenic peptides can induce T-cell tolerance, inhibit recruitment of inflammatory cells and reverse autoimmune diseases. Here we demonstrate a novel regulatory pathway that involves RTL binding to CD11b+ mononuclear cells through a receptor comprised of MHC class II invariant chain (CD74), cell-surface histones and MHC class II itself for treatment of experimental autoimmune encephalomyelitis (EAE). Binding of RTL constructs with CD74 involved a previously unrecognized MHC class II-α1/CD74 interaction that inhibited CD74 expression, blocked activity of its ligand, macrophage migration inhibitory factor, and reduced EAE severity. These findings implicate binding of RTL constructs to CD74 as a key step in both antigen-driven and bystander T-cell tolerance important in treatment of inflammatory diseases. PMID:23026773

  16. An ATF/CREB binding motif is required for aberrant constitutive expression of the MHC class II DR alpha promoter and activation by SV40 T-antigen.

    PubMed Central

    Cox, P M; Goding, C R

    1992-01-01

    Constitutive expression of major histocompatibility complex class II (MHC II) antigens normally occurs in B-lymphocytes and antigen presenting cells of the monocyte/macrophage lineage. However, many malignant tumours and transformed cells express these proteins aberrantly. We demonstrate here that the MHC II DR alpha promoter is constitutively active both in the SV40 large T antigen transformed cell line, COS, and in CV1 cells from which they are derived. As an approach to understanding the molecular mechanisms underlying aberrant DR alpha expression we have examined the cis- and trans-acting requirements for DR alpha transcription in these cell types. Electrophoretic mobility shift assays showed that the region immediately 3' to the X-box was bound by a member of the ATF/CREB family of transcription factors. Using deletions and point mutations in the DR alpha promoter we demonstrate that, in contrast to B-cells, the octamer motif and conserved X- and Y-boxes make only a minor contribution to promoter function while single point mutations in the ATF/CREB motif reduced transcription up to 20-fold. In addition, we show that the DR alpha promoter is activated by SV40 large T-antigen and that activation requires an intact ATF/CREB motif. Similar data were obtained using B16 melanoma cells. These results suggest that the ATF/CREB motif may be a target for transcription deregulation in several transformed cell types. Images PMID:1329030

  17. Differential regulation of expression of the MHC class II molecules RT1.B and RT1.D on rat B lymphocytes: effects of interleukin-4, interleukin-13 and interferon-gamma.

    PubMed Central

    Roos, A; Schilder-Tol, E J; Chand, M A; Claessen, N; Lakkis, F G; Pascual, D W; Weening, J J; Aten, J

    1998-01-01

    Susceptibility to induction of both T helper 1- (Th1) and Th2-mediated autoimmunity is multifactorial and involves genetic linkage to the major histocompatibility complex (MHC) class II haplotype. Brown Norway (BN) rats exposed to mercuric chloride develop a Th2-dependent systemic autoimmunity, whereas Lewis rats, which are highly susceptible to Th1-mediated autoimmunity, develop immune suppression after mercuric chloride exposure. Exposure to mercuric chloride is known to enhance B-lymphocyte expression of the MHC class II molecule RT1.B, predominantly in BN rats. We demonstrate that, in contrast, expression of RT1.D was unmodified on these B cells, whereas both RT1.B and RT1.D were up-regulated on epithelial cells. Regulation of B-cell MHC class II isotype expression was further studied in vitro, using BN rat lymph node (LN) cells. Interleukin-4 (IL-4) strongly enhanced B-cell expression of RT1.B (2.8-fold), whereas RT1.D expression was only slightly, although significantly, modified (1.2-fold). B cells from Lewis rats showed a similar IL-4-induced enhancement of RT1.B expression (2.5-fold), whereas, in contrast, RT1.D expression was unmodified. Exposure of LN cells from BN rats to interferon-gamma induced a moderate increase of B-cell MHC class II expression, predominantly of RT1.B. Strong and rapid enhancement of B-cell RT1.D expression was observed after stimulation by phorbol 12-myristate 13-acetate and ionomycin. Rat IL-13 did not modify B-cell MHC class II expression; however, it induced typical morphological changes in peritoneal macrophages. These experiments demonstrate isotype-specific and strain-dependent regulation of MHC class II expression on rat B lymphocytes, which may be of pathophysiological relevance for the strain-dependent susceptibility for Th1- or Th2-mediated autoimmunity. Images Figure 1 Figure 5 PMID:9536116

  18. A Case of Probable MHC Class II Deficiency with Disseminated BCGitis.

    PubMed

    Alyasin, Soheyla; Abolnezhadian, Farhad; Khoshkhui, Maryam

    2015-09-01

    Major histocompatibility complex (MHC) class II deficiency is a primary immunodeficiency disease characterized by abnormality of MHC class II molecules surface expression on peripheral blood lymphocytes and monocytes. Clinical manifestations include extreme susceptibility to viral, bacterial, and fungal infections but the immunodeficiency is not as severe as SCID (severe combined immunodeficiency), as evidenced by failure to develop disseminated infection after BCG vaccination. Therefore, MHC II deficiency with BCGosis, that is disseminated BCGitis, is not reported commonly. We report an interesting case of BCGosis after vaccination that was diagnosed to have probable MHC II deficiency. PMID:26412640

  19. Endogenous Antigen Presentation of MHC Class II Epitopes through Non-Autophagic Pathways

    PubMed Central

    Leung, Carol S. K.

    2015-01-01

    Antigenic peptides presented by major histocompatibility complex (MHC) class II molecules are generally derived from exogenous proteins acquired by antigen presenting cells. However, in some circumstances, MHC class II molecules can present intracellular proteins expressed within the antigen-presenting cells. There are several described pathways by which endogenous antigens are degraded and gain access to MHC class II molecules. These include autophagy and other non-autophagic pathways; the latter category includes the MHC class I-like pathways, heat shock protein 90-mediated pathways, and internalization from the plasma membrane. This review will summarize and discuss the non-autophagic pathways. PMID:26441969

  20. Secreted Toxoplasma gondii molecules interfere with expression of MHC-II in interferon gamma-activated macrophages.

    PubMed

    Leroux, Louis-Philippe; Dasanayake, Dayal; Rommereim, Leah M; Fox, Barbara A; Bzik, David J; Jardim, Armando; Dzierszinski, Florence S

    2015-04-01

    The obligate intracellular protozoan parasite Toxoplasma gondii interferes with major histocompatibility complex class II antigen presentation to dampen host CD4(+) T cell responses. While it is known that T. gondii inhibits major histocompatibility complex class II gene transcription and expression in infected host cells, the mechanism of this host manipulation is unknown. Here, we show that soluble parasite proteins inhibit IFNγ-induced expression of major histocompatibility complex class II on the surface of the infected cell in a dose-dependent response that was abolished by protease treatment. Subcellular fractionation of T. gondii tachyzoites revealed that the major histocompatibility complex class II inhibitory activity co-partitioned with rhoptries and/or dense granules. However, parasite mutants deleted for single rhoptries or dense granules genes (ROP1, 4/7, 14, 16 and 18 or GRA 2-9 and 12 knock-out strains) retained the ability to inhibit expression of major histocompatibility complex class II. In addition, excreted/secreted antigens released by extracellular tachyzoites displayed immunomodulatory activity characterized by an inhibition of major histocompatibility complex class II expression, and reduced expression and release of TNFα by macrophages. Tandem MS analysis of parasite excreted/secreted antigens generated a list of T. gondii secreted proteins that may participate in major histocompatibility complex class II inhibition and the modulation of host immune functions. PMID:25720921

  1. CITA/NLRC5: A critical transcriptional regulator of MHC class I gene expression.

    PubMed

    Downs, Isaac; Vijayan, Saptha; Sidiq, Tabasum; Kobayashi, Koichi S

    2016-07-01

    Major histocompatibility complex (MHC) class I and class II molecules play essential roles in the development and activation of the human adaptive immune system. An NLR protein, CIITA (MHC class II transactivator) has been recognized as a master regulator of MHC class II gene expression, albeit knowledge about the regulatory mechanism of MHC class I gene expression had been limited. Recently identified MHC class I transactivator (CITA), or NLRC5, also belongs to the NLR protein family and constitutes a critical regulator for the transcriptional activation of MHC class I genes. In addition to MHC class I genes, CITA/NLRC5 induces the expression of β2 -microglobulin, TAP1 and LMP2, essential components of the MHC class I antigen presentation pathway. Therefore, CITA/NLRC5 and CIITA are transcriptional regulators that orchestrate the concerted expression of critical components in the MHC class I and class II pathways, respectively. © 2016 BioFactors, 42(4):349-357, 2016. PMID:27087581

  2. RAT CYTOMEGALOVIRUS INFECTION DEPLETES MHC II IN BONE MARROW DERIVED DENDRITIC CELLS

    PubMed Central

    Baca Jones, Carmen C.; Kreklywich, Craig N.; Messaoudi, Ilhem; Vomaske, Jennifer; McCartney, Erin; Orloff, Susan L.; Nelson, Jay A.; Streblow, Daniel N.

    2009-01-01

    While cytomegalovirus (CMV) infects and replicates in a multitude of cell types, the ability of the virus to replicate in antigen presenting cells (APCs) is believed to play a critical role in the viral dissemination and latency. CMV infection of APCs and manipulation of their function is an important area of investigation. CMV down regulation of MHC II is reportedly mediated by the HCMV proteins US2, US3, UL83, UL111a (vIL10) or through the induction of cellular IL10. In this study, we demonstrate that rat CMV (RCMV) significantly reduces MHC II expression by mechanisms that do not involve orthologues of the known HCMV genes nor by an increase in cellular IL10. Rat bone marrow derived dendritic cells (BMDC) were highly susceptible to infection with RCMV and a recombinant RCMV expressing eGFP. RCMV infection of BMDCs depleted both surface and intracellular MHC II to nearly undetectable levels as well as reduced surface expression of MHC I. The effect on MHC II only occurred in the infected GFP positive cells and is mediated by an immediate early or early viral gene product. Furthermore, treatment of uninfected immature DCs with virus-free conditioned supernatants from infected cells failed to down regulate MHC II. RCMV depletion of MHC II was sensitve to treatment with lysosomal inhibitors but not proteasomal inhibitors suggesting that the mechanism of RCMV mediated down-regulation of MHC II occurs through endocytic degradation. Since RCMV does not encode homologues of US2, US3, UL83 or UL111a, these data indicate a novel mechanism for RCMV depletion of MHC II. PMID:19349057

  3. FCRL6 is an MHC class II receptor1

    PubMed Central

    Schreeder, Daniel M.; Cannon, John P.; Wu, Jiongru; Li, Ran; Shakhmatov, Mikhail A.; Davis, Randall S.

    2016-01-01

    Receptors for the Fc portion (FCR) of Ig have been extensively characterized and are known to regulate humoral responses, but members of the closely related FCR-like (FCRL) family have not been found to bind Ig and to date no ligand has been identified for any FCRL. Using a cell-based GFP reporter system and a recombinant Fc chimeric protein, we show that human FCRL6, a receptor selectively expressed by cytotoxic T and NK cells, directly binds HLA-DR, a major histocompatibility complex (MHC) class II molecule. Given the similarity among constant regions of Ig and MHC molecules, these findings suggest that representatives of the FCR and FCRL multigene families may have independently evolved to engage two ancestral elements fundamental to adaptive immunity. This discovery may offer new insight into the interaction between cytotoxic lymphocytes and antigen presenting cells and may have important implications for better understanding HLA disease susceptibility and pathogenesis. PMID:20519654

  4. MHC class II antigen presentation pathway in murine tumours: tumour evasion from immunosurveillance?

    PubMed Central

    Walter, W; Lingnau, K; Schmitt, E; Loos, M; Maeurer, M J

    2000-01-01

    Qualitative differences in the MHC class II antigen processing and presentation pathway may be instrumental in shaping the CD4+ T cell response directed against tumour cells. Efficient loading of many MHC class II alleles with peptides requires the assistance of H2-M, a heterodimeric MHC class II-like molecule. In contrast to the HLA-DM region in humans, the β-chain locus is duplicated in mouse, with the H2-Mb1 (Mb1β-chain distal to H2-Mb2 (Mb2) and the H2-Ma (Ma) α-chain gene). Here, we show that murine MHC class II and H2-M genes are coordinately regulated in murine tumour cell lines by T helper cell 1 (IFN-γ) and T helper cell 2 (IL-4 or IL-10) cytokines in the presence of the MHC class II-specific transactivator CIITA as determined by mRNA expression and Western blot analysis. Furthermore, Mαβ1 and Mαβ2 heterodimers are differentially expressed in murine tumour cell lines of different histology. Both H2-M isoforms promote equally processing and presentation of native protein antigens to H2-Ad- and H2-Ed-restricted CD4+ T cells. Murine tumour cell lines could be divided into three groups: constitutive MHC class II and CIITA expression; inducible MHC class II and CIITA expression upon IFN-γ-treatment; and lack of constitutive and IFN-γ-inducible MHC class II and CIITA expression. These differences may impact on CD4+ T cell recognition of cancer cells in murine tumour models. © 2000 Cancer Research Campaign PMID:11027433

  5. Constitutive induction of intestinal Tc17 cells in the absence of hematopoietic cell-specific MHC class II expression.

    PubMed

    Rubino, Stephen J; Geddes, Kaoru; Magalhaes, Joao G; Streutker, Catherine; Philpott, Dana J; Girardin, Stephen E

    2013-11-01

    The enteric pathogen Citrobacter rodentium induces a mucosal IL-17 response in CD4(+) T helper (Th17) cells that is dependent on the Nod-like receptors Nod1 and Nod2. Here, we sought to determine whether this early Th17 response required antigen presentation by major histocompatibility complex class II (MHCII) for full induction. At early phases of C. rodentium infection, we observed that the intestinal mucosal Th17 response was fully blunted in irradiated mice reconstituted with MHCII-deficient (MHCII(-/-) →WT) hematopoietic cells. Surprisingly, we also observed a substantial increase in the relative frequency of IL-17(+) CD8(+) CD4(-) TCR-β(+) cells (Tc17 cells) and FOXP3(+) CD8(+) CD4(-) TCR-β(+) cells in the lamina propria and intraepithelial lymphocyte compartment of MHCII(-/-) →WT mice compared with that in WT→WT counterparts. Moreover, MHCII(-/-) →WT mice displayed increased susceptibility, increased bacterial translocation to deeper organs, and more severe colonic histopathology after infection with C. rodentium. Finally, a similar phenotype was observed in mice deficient for CIITA, a transcriptional regulator of MHCII expression. Together, these results indicate that MHCII is required to mount early mucosal Th17 responses to an enteric pathogen, and that MHCII regulates the induction of atypical CD8(+) T-cell subsets, such as Tc17 cells and FOXP3(+) CD8(+) cells, in vivo. PMID:23881368

  6. Protection against H1N1 influenza challenge by a DNA vaccine expressing H3/H1 subtype hemagglutinin combined with MHC class II-restricted epitopes

    PubMed Central

    2010-01-01

    Background Multiple subtypes of avian influenza viruses have crossed the species barrier to infect humans and have the potential to cause a pandemic. Therefore, new influenza vaccines to prevent the co-existence of multiple subtypes within a host and cross-species transmission of influenza are urgently needed. Methods Here we report a multi-epitope DNA vaccine targeted towards multiple subtypes of the influenza virus. The protective hemagglutinin (HA) antigens from H5/H7/H9 subtypes were screened for MHC II class-restricted epitopes overlapping with predicted B cell epitopes. We then constructed a DNA plasmid vaccine, pV-H3-EHA-H1, based on HA antigens from human influenza H3/H1 subtypes combined with the H5/H7/H9 subtype Th/B epitope box. Results Epitope-specific IFN-γ ELISpot responses were significantly higher in the multi-epitope DNA group than in other vaccine and control groups (P < 0.05). The multi-epitope group significantly enhanced Th2 cell responses as determined by cytokine assays. The survival rate of mice given the multi-epitope vaccine was the highest among the vaccine groups, but it was not significantly different compared to those given single antigen expressing pV-H1HA1 vaccine and dual antigen expressing pV-H3-H1 vaccine (P > 0.05). No measurable virus titers were detected in the lungs of the multi-epitope immunized group. The unique multi-epitope DNA vaccine enhanced virus-specific antibody and cellular immunity as well as conferred complete protection against lethal challenge with A/New Caledonia/20/99 (H1N1) influenza strain in mice. Conclusions This approach may be a promising strategy for developing a universal influenza vaccine to prevent multiple subtypes of influenza virus and to induce long-term protective immune against cross-species transmission. PMID:21134292

  7. Blocking MHC class II on human endothelium mitigates acute rejection

    PubMed Central

    Abrahimi, Parwiz; Qin, Lingfeng; Chang, William G.; Bothwell, Alfred L.M.; Tellides, George; Saltzman, W. Mark; Pober, Jordan S.

    2016-01-01

    Acute allograft rejection is mediated by host CD8+ cytotoxic T lymphocytes (CTL) targeting graft class I major histocompatibility complex (MHC) molecules. In experimental rodent models, rejection requires differentiation of naive CD8+ T cells into alloreactive CTL within secondary lymphoid organs, whereas in humans, CTL may alternatively develop within the graft from circulating CD8+ effector memory T cells (TEM) that recognize class I MHC molecules on graft endothelial cells (EC). This latter pathway is poorly understood. Here, we show that host CD4+ TEM, activated by EC class II MHC molecules, provide critical help for this process. First, blocking HLA-DR on EC lining human artery grafts in immunodeficient mice reduces CD8+ CTL development within and acute rejection of the artery by adoptively transferred allogeneic human lymphocytes. Second, siRNA knockdown or CRISPR/Cas9 ablation of class II MHC molecules on EC prevents CD4+ TEM from helping CD8+ TEM to develop into CTL in vitro. Finally, implanted synthetic microvessels, formed from CRISPR/Cas9-modified EC lacking class II MHC molecules, are significantly protected from CD8+ T cell–mediated destruction in vivo. We conclude that human CD8+ TEM–mediated rejection targeting graft EC class I MHC molecules requires help from CD4+ TEM cells activated by recognition of class II MHC molecules. PMID:26900601

  8. DNA sequence of the Peromyscus leucopus MHC class II gene Aa (MhcPeleAa)

    SciTech Connect

    Crew, M.D.; Bates, L.M.

    1996-09-01

    The genus Peromyscus has been extensively studied by populations biologists and ecologists for over eighty years, with P. leucopus (the white-footed mouse) being one of the most intensively investigated species. Polymorphic major histocompatibility complex (MHC) genes have proven useful in population genetic studies and might be helpful in understanding the population dynamics of Peromyscus species which are ubiquitously distributed over North and Central America. Polymorphism of P. leucopus MHC (MhcPele) class II genes was evident by restriction fragment length polymorphism (RFLP) analyses using human and mouse probes and Pele class II loci exhibited degrees of polymorphism similar to H2 class II genes (A-like>E-like). 8 refs., 2 figs.

  9. MHC evolution in three salmonid species: a comparison between class II alpha and beta genes.

    PubMed

    Gómez, Daniela; Conejeros, Pablo; Marshall, Sergio H; Consuegra, Sofia

    2010-08-01

    The genes of the major histocompatibility complex (MHC) are amongst the most variable in vertebrates and represent some of the best candidates to study processes of adaptive evolution. However, despite the number of studies available, most of the information on the structure and function of these genes come from studies in mammals and birds in which the MHC class I and II genes are tightly linked and class II alpha exhibits low variability in many cases. Teleost fishes are among the most primitive vertebrates with MHC and represent good organisms for the study of MHC evolution because their class I and class II loci are not physically linked, allowing for independent evolution of both classes of genes. We have compared the diversity and molecular mechanisms of evolution of classical MH class II alpha and class II beta loci in farm populations of three salmonid species: Oncorhynchus kisutch, Oncorhynchus mykiss and Salmo salar. We found single classical class II loci and high polymorphism at both class II alpha and beta genes in the three species. Mechanisms of evolution were common for both class II genes, with recombination and point mutation involved in generating diversity and positive selection acting on the peptide-binding residues. These results suggest that the maintenance of variability at the class IIalpha gene could be a mechanism to increase diversity in the MHC class II in salmonids in order to compensate for the expression of one single classical locus and to respond to a wider array of parasites. PMID:20521040

  10. Immunological Functions of the Membrane Proximal Region of MHC Class II Molecules

    PubMed Central

    Harton, Jonathan; Jin, Lei; Hahn, Amy; Drake, Jim

    2016-01-01

    Major histocompatibility complex (MHC) class II molecules present exogenously derived antigen peptides to CD4 T cells, driving activation of naïve T cells and supporting CD4-driven immune functions. However, MHC class II molecules are not inert protein pedestals that simply bind and present peptides. These molecules also serve as multi-functional signaling molecules delivering activation, differentiation, or death signals (or a combination of these) to B cells, macrophages, as well as MHC class II-expressing T cells and tumor cells. Although multiple proteins are known to associate with MHC class II, interaction with STING (stimulator of interferon genes) and CD79 is essential for signaling. In addition, alternative transmembrane domain pairing between class II α and β chains influences association with membrane lipid sub-domains, impacting both signaling and antigen presentation. In contrast to the membrane-distal region of the class II molecule responsible for peptide binding and T-cell receptor engagement, the membrane-proximal region (composed of the connecting peptide, transmembrane domain, and cytoplasmic tail) mediates these “non-traditional” class II functions. Here, we review the literature on the function of the membrane-proximal region of the MHC class II molecule and discuss the impact of this aspect of class II immunobiology on immune regulation and human disease. PMID:27006762

  11. MHC2SKpan: a novel kernel based approach for pan-specific MHC class II peptide binding prediction

    PubMed Central

    2013-01-01

    Background Computational methods for the prediction of Major Histocompatibility Complex (MHC) class II binding peptides play an important role in facilitating the understanding of immune recognition and the process of epitope discovery. To develop an effective computational method, we need to consider two important characteristics of the problem: (1) the length of binding peptides is highly flexible; and (2) MHC molecules are extremely polymorphic and for the vast majority of them there are no sufficient training data. Methods We develop a novel string kernel MHC2SK (MHC-II String Kernel) method to measure the similarities among peptides with variable lengths. By considering the distinct features of MHC-II peptide binding prediction problem, MHC2SK differs significantly from the recently developed kernel based method, GS (Generic String) kernel, in the way of computing similarities. Furthermore, we extend MHC2SK to MHC2SKpan for pan-specific MHC-II peptide binding prediction by leveraging the binding data of various MHC molecules. Results MHC2SK outperformed GS in allele specific prediction using a benchmark dataset, which demonstrates the effectiveness of MHC2SK. Furthermore, we evaluated the performance of MHC2SKpan using various benckmark data sets from several different perspectives: Leave-one-allele-out (LOO), 5-fold cross validation as well as independent data testing. MHC2SKpan has achieved comparable performance with NetMHCIIpan-2.0 and outperformed NetMHCIIpan-1.0, TEPITOPEpan and MultiRTA, being statistically significant. MHC2SKpan can be freely accessed at http://datamining-iip.fudan.edu.cn/service/MHC2SKpan/index.html. PMID:24564280

  12. Towards a systems understanding of MHC class I and MHC class II antigen presentation.

    PubMed

    Neefjes, Jacques; Jongsma, Marlieke L M; Paul, Petra; Bakke, Oddmund

    2011-12-01

    The molecular details of antigen processing and presentation by MHC class I and class II molecules have been studied extensively for almost three decades. Although the basic principles of these processes were laid out approximately 10 years ago, the recent years have revealed many details and provided new insights into their control and specificity. MHC molecules use various biochemical reactions to achieve successful presentation of antigenic fragments to the immune system. Here we present a timely evaluation of the biology of antigen presentation and a survey of issues that are considered unresolved. The continuing flow of new details into our understanding of the biology of MHC class I and class II antigen presentation builds a system involving several cell biological processes, which is discussed in this Review. PMID:22076556

  13. Heparan sulfates targeting increases MHC class I- and MHC class II-restricted antigen presentation and CD8(+) T-cell response.

    PubMed

    Knittel, Delphine; Gadzinski, Adeline; Hua, Stéphane; Denizeau, Jordan; Savatier, Alexandra; de la Rochère, Philippe; Boulain, Jean-Claude; Amigorena, Sebastian; Piaggio, Eliane; Sedlik, Christine; Léonetti, Michel

    2016-06-01

    Heparan sulfates (HS) are carbohydrate moieties of HS proteoglycans (HSPGs). They often represent alternative attachment points for proteins or microorganisms targeting receptors. HSPGs, which are ubiquitously expressed, thereby participate in numerous biological processes. We previously showed that MHC class II-restricted antigen presentation is increased when antigens are coupled to HS ligands, suggesting that HSPGs might contribute to adaptive immune responses. Here, we examined if HSPG targeting influences other aspects of immune responses. We found that coupling of an HS ligand to the antigen increases antigen presentation to CD4(+) and CD8(+) T-cells after antigen targeting to membrane immunoglobulins or to MHC-II molecules. Moreover, this increased stimulating capacity correlates with an enhanced CD8(+) immune response in mice. Last, animals control more effectively the growth of Ova-expressing tumour cells when they are immunized with an Ova construct targeting HSPGs and MHC-II molecules. Our results indicate that ubiquitous molecules can influence both MHC class I- and MHC class II-restricted antigen presentation and behave as co-receptors during T-cell stimulation. Moreover, they suggest that tumour-antigens endowed with the ability to target both HSPGs and MHC-II molecules could be of value to increase CD8(+) immune response and control tumour-growth, opening new perspectives for the design of highly immunogenic protein-based vaccines. PMID:27154391

  14. Multiple sclerosis: a role for astroglia in active demyelination suggested by class II MHC expression and ultrastructural study.

    PubMed

    Lee, S C; Moore, G R; Golenwsky, G; Raine, C S

    1990-03-01

    Central nervous system (CNS) tissue was studied by immunocytochemistry and electron microscopy from three cases of multiple sclerosis (MS) in which evidence of ongoing myelin breakdown could be documented. The study focussed upon the role of glial cells in the pathogenesis of demyelination. In acute MS, demyelination involved the vesicular dissolution of myelin from intact axons and a paucity of fibrillary astrogliosis. Foamy macrophages, many of them probably derived from transformed and recently proliferated microglia, contained recognizable myelin debris and lipid droplets and were abundant throughout the lesions. These cells formed the major phagocytic population and stained positively for class II major histocompatibility complex antigens (HLA-DR; Ia). In acute MS lesions, rounded astrocytes were encountered which possessed membrane-bound compartments enclosing phagocytosed fragments of myelin basic protein-positive debris. Despite the superficial resemblance of these cells to foamy macrophages, the presence of intermediate filaments, glycogen granules and diffuse glial fibrillary acidic protein positivity supported an astroglial identity. Astrocyte processes were involved in myelin removal and invested recently demyelinated axons. Hypertrophic fibrous astrocytes were common in chronic active lesions, were capable of myelin degradation and on occasion, contained myelin debris attached to clathrin-coated pits. These astrocytes were sometimes Ia+. Oligodendrocytes were depleted from the center of active lesions but were numerous at the lesion margin, suggesting survival and proliferation. They stained positively for myelin-associated glycoprotein, a marker for immature oligodendrocytes. However, they were invariably Ia-. The findings confirm and further support a role for the astrocyte as both an antigen presenting cell and a phagocyte in the CNS during MS. PMID:2307980

  15. MHC class II proteins contain a potential binding site for the verotoxin receptor glycolipid CD77.

    PubMed

    George, T; Boyd, B; Price, M; Lingwood, C; Maloney, M

    2001-11-01

    Globotriaosyl ceramide or CD77 functions as a cell surface receptor for toxins of the Shiga toxin/verotoxin family and as a marker for germinal center stage B-cells. The B-cell protein CD19 and the interferon-alpha receptor possess verotoxin-like amino acid sequences in their extracellular domains, and CD77 has been shown to function in CD19-mediated adhesion and interferon-induced growth inhibition. The Burkitt's lymphoma cell line, Daudi, is similar to germinal center B-cells in their expression of CD77, CD19 and MHC class II molecules. Using the multiple sequence alignment program, ClustalW, we have identified a verotoxin-like amino acid sequence on the beta-chain of human and murine MHC class II molecules. Binding of CD77 at this site could modulate the peptide-binding properties of these MHC class II molecules. Using Western blot analysis of whole cell extracts, we found that CD77-positive Daudi cells have higher levels of HLA-D proteins than VT500 cells, a Daudi-derived CD77-deficient mutant cell line. In contrast, MHC class II-mediated adhesion and surface expression are similar in the two cell lines. Therefore, CD77 could play a functional or regulatory role in MHC class II-mediated functions specifically relating to antigen presentation by B-cells to T helper cells. PMID:11838965

  16. The tetraspanin CD9 mediates lateral association of MHC class II molecules on the dendritic cell surface

    PubMed Central

    Unternaehrer, Julia J.; Chow, Amy; Pypaert, Marc; Inaba, Kayo; Mellman, Ira

    2007-01-01

    We have found that MHC class II (MHC II) molecules exhibit a distinctive organization on the dendritic cell (DC) plasma membrane. Both in DC lysates and on the surface of living cells, I-A and I-E molecules engaged in lateral interactions not observed on other antigen-presenting cells such as B blasts. Because DCs and B blasts express MHC II at comparable surface densities, the interaction was not due to simple mass action. Instead, it reflected the selective expression of the tetraspanin CD9 at the DC surface. I-A and I-E molecules coprecipitated with each other and with CD9. The association of heterologous MHC II molecules was abrogated in DCs from CD9−/− mice. Conversely, expression of exogenous CD9 in B cells induced MHC II interactions. CD9 is thus necessary for the association of heterologous MHC II, a specialization that would facilitate the formation of MHC II multimers expected to enhance T cell receptor stimulation by DCs. PMID:17190803

  17. Features of target cell lysis by class I and class II MHC restricted cytolytic T lymphocytes

    SciTech Connect

    Maimone, M.M.; Morrison, L.A.; Braciale, V.L.; Braciale, T.J.

    1986-12-01

    The lytic activity of influenza virus-specific muvine cytolytic T lymphocyte (CTL) clones that are restricted by either H-2K/D (class I) or H-2I (class II) major histocompatibility (MHC) locus products was compared on an influenza virus-infected target cell expressing both K/D and I locus products. With the use of two in vitro measurements of cytotoxicity, conventional /sup 51/Cr release, and detergent-releasable radiolabeled DNA (as a measure of nuclear disintegration in the early post-lethal hit period), the authors found no difference between class I and class II MHC-restricted CTL in the kinetics of target cell destruction. In addition, class II MHC-restricted antiviral CTL failed to show any lysis of radiolabeled bystander cells. Killing of labeled specific targets by these class II MHC-restricted CTL was also efficiently inhibited by unlabeled specific competitor cells in a cold target inhibition assay. In sum, these data suggest that class I and class II MHC-restricted CTL mediate target cell destruction by an essentially similar direct mechanism.

  18. MHC class II DR allelic diversity in bighorn sheep

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We hypothesized that decreased diversity and/or unique polymorphisms in MHC class II alleles of bighorn sheep (BHS, Ovis canadensis) are responsible for lower titer of antibodies against Mannheimia haemolytica leukotoxin, in comparison to domestic sheep (DS, Ovis aries). To test this hypothesis, DRA...

  19. Reversion of a transcriptionally defective MHC class II-negative human B-cell mutant.

    PubMed Central

    Ombra, M N; Perfetto, C; Autiero, M; Anzisi, A M; Pasquinelli, R; Maffei, A; Del Pozzo, G; Guardiola, J

    1993-01-01

    RJ2.2.5, a mutant derived from the human B-lymphoma cell, Raji, is unable to express the MHC class II genes because of a recessive transcriptional defect attributed to the lack of an activator function. We report the isolation of a RJ2.2.5 revertant, namely AR, in which the expression of the mRNAs encoded by these genes is restored. Comparison of the binding of nuclear extracts or of partially purified nuclear preparations from the wild-type, the mutant and the revertant cells to a conserved MHC class II promoter element, the X-box, showed no alteration in the mobility of the complexes thus formed. However, in extracts from RJ2.2.5, and other MHC class II negative cell lines, such as HeLa, the amount of complex observed was significantly higher than in wild-type Raji cells. Furthermore, the binding activity exhibited by the AR revertant was lower than that of the RJ2.2.5 and higher than that of Raji. The use of specific monoclonal antibodies indicated that in all cases c-Jun and c-Fos or antigenically related proteins were required for binding. An inverse correlation between the level of DNA-protein complex formed and the level of MHC class II gene mRNA expressed in the three cell lines was apparent, suggesting that overexpression of a DNA binding factor forming complexes with class II promoter elements may cause repression of MHC class II transcription. A model which reconciles the previously ascertained recessivity of the phenotype of the mutation carried by RJ2.2.5 with the findings reported here is discussed. Images PMID:8441650

  20. The anti-CD74 humanized monoclonal antibody, milatuzumab, which targets the invariant chain of MHC II complexes, alters B-cell proliferation, migration, and adhesion molecule expression

    PubMed Central

    2012-01-01

    Introduction Targeting CD74 as the invariant chain of major histocompatibility complexes (MHC) became possible by the availability of a specific humanized monoclonal antibody, milatuzumab, which is under investigation in patients with hematological neoplasms. CD74 has been reported to regulate chemo-attractant migration of macrophages and dendritic cells, while the role of CD74 on peripheral naïve and memory B cells also expressing CD74 remains unknown. Therefore, the current study addressed the influence of milatuzumab on B-cell proliferation, chemo-attractant migration, and adhesion molecule expression. Methods Surface expression of CD74 on CD27- naïve and CD27+ memory B cells as well as other peripheral blood mononuclear cells (PBMCs) obtained from normals, including the co-expression of CD44, CXCR4, and the adhesion molecules CD62L, β7-integrin, β1-integrin and CD9 were studied after binding of milatuzumab using multicolor flow cytometry. The influence of the antibody on B-cell proliferation and migration was analyzed in vitro in detail. Results In addition to monocytes, milatuzumab also specifically bound to human peripheral B cells, with a higher intensity on CD27+ memory versus CD27- naïve B cells. The antibody reduced B-cell proliferation significantly but moderately, induced enhanced spontaneous and CXCL12-dependent migration together with changes in the expression of adhesion molecules, CD44, β7-integrin and CD62L, mainly of CD27- naïve B cells. This was independent of macrophage migration-inhibitory factor as a ligand of CD74/CD44 complexes. Conclusions Milatuzumab leads to modestly reduced proliferation, alterations in migration, and adhesion molecule expression preferentially of CD27- naïve B cells. It thus may be a candidate antibody for the autoimmune disease therapy by modifying B cell functions. PMID:22404985

  1. Donor MHC class II antigen is essential for induction of transplantation tolerance by bone marrow cells.

    PubMed

    Umemura, A; Monaco, A P; Maki, T

    2000-05-01

    Posttransplant infusion of donor bone marrow cells (BMC) induces tolerance to allografts in adult mice, dogs, nonhuman primates, and probably humans. Here we used a mouse skin allograft model and an allogeneic radiation chimera model to examine the role of MHC Ags in tolerance induction. Infusion of MHC class II Ag-deficient (CIID) BMC failed to prolong C57BL/6 (B6) skin grafts in ALS- and rapamycin-treated B10.A mice, whereas wild-type B6 or MHC class I Ag-deficient BMC induced prolongation. Removal of class II Ag-bearing cells from donor BMC markedly reduced the tolerogenic effect compared with untreated BMC, although graft survival was significantly longer in mice given depleted BMC than that in control mice given no BMC. Infusion of CIID BMC into irradiated syngeneic B6 or allogeneic B10.A mice produced normal lymphoid cell reconstitution including CD4+ T cells except for the absence of class II Ag-positive cells. However, irradiated B10.A mice reconstituted with CIID BMC rejected all B6 and a majority of CIID skin grafts despite continued maintenance of high degree chimerism. B10.A mice reconstituted with B6 BMC maintained chimerism and accepted both B6 and CIID skin grafts. Thus, expression of MHC class II Ag on BMC is essential for allograft tolerance induction and peripheral chimerism with cells deficient in class II Ag does not guarantee allograft acceptance. PMID:10779744

  2. Inflammatory bowel diseases influence major histocompatibility complex class I (MHC I) and II compartments in intestinal epithelial cells.

    PubMed

    Bär, F; Sina, C; Hundorfean, G; Pagel, R; Lehnert, H; Fellermann, K; Büning, J

    2013-05-01

    Antigen presentation by intestinal epithelial cells (IEC) is crucial for intestinal homeostasis. Disturbances of major histocompatibility complex class I (MHC I)- and II-related presentation pathways in IEC appear to be involved in an altered activation of CD4(+) and CD8(+) T cells in inflammatory bowel disease. However, a comprehensive analysis of MHC I- and II-enriched compartments in IEC of the small and large bowel in the healthy state as opposed to inflammatory bowel diseases is lacking. The aim of this study was to characterize the subcellular expression of MHC I and II in the endocytic pathway of IEC throughout all parts of the intestinal tract, and to identify differences between the healthy state and inflammatory bowel diseases. Biopsies were taken by endoscopy from the duodenum, jejunum, ileum and colon in healthy individuals (n = 20). In Crohn's disease (CD), biopsies were obtained from the ileum and colon and within the colon from ulcerative colitis (UC) patients (n = 15). Analysis of IEC was performed by immunoelectron microscopy. MHC I and II were identified in early endosomes and multi-vesicular, multi-lamellar, electrondense and vacuolar late endosomes. Both molecules were enriched in multi-vesicular bodies. No differences were found between the distinct parts of the gut axis. In CD and UC the expression of MHC I and II showed a shift from multi-vesicular bodies towards the basolateral membranes. Within the multi-vesicular bodies, MHC I and II moved from internal vesicles to the limiting membranes upon inflammation in CD and UC. MHC I- and II-enriched compartments in IEC were identical in all parts of the small and large bowel. CD and UC appear to modulate the MHC I- and II-related presentation pathways of exogenous antigens in IEC. PMID:23574324

  3. MHC class II transcription is associated with inflammatory responses in a wild marine mammal.

    PubMed

    Montano-Frías, Jorge E; Vera-Massieu, Camila; Álvarez-Martínez, Roberto; Flores-Morán, Adriana; Acevedo-Whitehouse, Karina

    2016-08-01

    Inflammation is one of the most important non-specific and rapid responses that a vertebrate can elicit in response to damage or a foreign insult. To date, despite increasing evidence that the innate and adaptive branches of immunity are more intricately related than previously thought, few have examined interactions between the Major Histocompatibility Complex (MHC, a polymorphic region of the vertebrate genome that is involved with antigen presentation) and inflammation, and even less is known about these interactions in an eco-immunological context. Here, we examined the effect of MHC class II DRB gene multiplicity and transcription on phytohemagglutinin (PHA)-induced inflammation during the early stages of development of California sea lions. Neither constitutive nor expressed ZacaDRB diversity was found to be associated with pup responses to PHA at any of the stages of pup development. However, for two-month-old pups, those with a specific MHC-DRB locus (ZacaDRB-A) tended to have less efficient responsive inflammation. Transcription of distinct MHC-DRB loci was also linked to PHA-induced inflammation, with patterns that varied markedly between ages, and that suggested that ongoing infectious processes could limit the capacity to respond to a secondary challenge. Life history constraints and physiological processes associated with development of California sea lions, in conjunction with their changing pathogenic environment could explain the observed effects of MHC class II transcription on PHA-induced inflammation. To our knowledge, ours is the first study to examine the importance of expressed vs. constitutive MHC loci on inflammation in a natural population. PMID:27137083

  4. MHC Class II haplotypes of Colombian Amerindian tribes

    PubMed Central

    Yunis, Juan J.; Yunis, Edmond J.; Yunis, Emilio

    2013-01-01

    We analyzed 1041 individuals belonging to 17 Amerindian tribes of Colombia, Chimila, Bari and Tunebo (Chibcha linguistic family), Embera, Waunana (Choco linguistic family), Puinave and Nukak (Maku-Puinave linguistic families), Cubeo, Guanano, Tucano, Desano and Piratapuyo (Tukano linguistic family), Guahibo and Guayabero (Guayabero Linguistic Family), Curripaco and Piapoco (Arawak linguistic family) and Yucpa (Karib linguistic family). for MHC class II haplotypes (HLA-DRB1, DQA1, DQB1). Approximately 90% of the MHC class II haplotypes found among these tribes are haplotypes frequently encountered in other Amerindian tribes. Nonetheless, striking differences were observed among Chibcha and non-Chibcha speaking tribes. The DRB1*04:04, DRB1*04:11, DRB1*09:01 carrying haplotypes were frequently found among non-Chibcha speaking tribes, while the DRB1*04:07 haplotype showed significant frequencies among Chibcha speaking tribes, and only marginal frequencies among non-Chibcha speaking tribes. Our results suggest that the differences in MHC class II haplotype frequency found among Chibcha and non-Chibcha speaking tribes could be due to genetic differentiation in Mesoamerica of the ancestral Amerindian population into Chibcha and non-Chibcha speaking populations before they entered into South America. PMID:23885196

  5. MHC II gene knockout in tissue engineering may prevent immune rejection of transplants.

    PubMed

    Yang, Miaomiao; Liu, Lei

    2008-01-01

    The repair and reconstruction of tissue defects and organ loss are severe problems, and many patients are eager to find avenues to these matters. Up until now, the number of methods used to repair tissue defects has increased, but all of these have their own advantages and inconveniences, and do not seem to have been optimized. The development of tissue engineering offers new hopes to patients with tissue defects. To regenerate tissues and organs, we first need a source of seed cells. However, the sources of autologous cells are restricted, cell number is small, and xenogenic cells result in immunological rejections. Major histocompatibility complex (MHC) polymorphism is a key factor in tissue grafts. MHC II, in particular, is associated with allogeneic transplantation. We hypothesize that if we knock-out the MHC II gene of mesenchymal stem cells (MSCs) in vitro, these cells would not express MHC II molecules, and rejection problems will be solved. Accordingly, the industrialization of tissue engineering will be feasible, and products of tissue engineering will be utilized widely for any clinical treatments. PMID:17904760

  6. MHC class II presentation is controlled by the lysosomal small GTPase, Arl8b.

    PubMed

    Michelet, Xavier; Garg, Salil; Wolf, Benjamin J; Tuli, Amit; Ricciardi-Castagnoli, Paola; Brenner, Michael B

    2015-03-01

    Dendritic cells (DCs) are specialized APCs with the ability to prime naive T cells. DCs first sample Ags from the environment and then orchestrate their processing and loading onto MHC class II (MHC II) Ag-presenting molecules in lysosomes. Once MHC II molecules have bound a peptide, the MHC II-peptide complex is delivered to the cell surface for presentation to CD4(+) T cells. Regulation of Ag uptake via macropinocytosis and phagocytosis has been extensively studied, as well as trafficking in early endocytic vesicles notably regulated by the small GTPase Rab5 and its effectors. However, little is known about the regulators of Ag delivery from early endosomes to lysosomal compartments where the proper pH, proteases, MHC II, invariant chain, and HLA-DM reside, awaiting exogenous Ags for loading. In this article, we report the crucial role of the small GTPase ADP-ribosylation factor-like 8b (Arl8b) in MHC II presentation in DCs. We show for the first time, to our knowledge, that Arl8b localizes to MHC II compartments in DCs and regulates formation of MHC II-peptide complexes. Arl8b-silenced DCs display a defect in MHC II-Ag complex formation and its delivery to the cell surface during infection resulting in a defect in T cell recognition. Our results highlight the role of Arl8b as a trafficking regulator of the late stage of complex formation and MHC II presentation in DCs. PMID:25637027

  7. The functional importance of sequence versus expression variability of MHC alleles in parasite resistance.

    PubMed

    Axtner, Jan; Sommer, Simone

    2012-12-01

    Understanding selection processes driving the pronounced allelic polymorphism of the major histocompatibility complex (MHC) genes and its functional associations to parasite load have been the focus of many recent wildlife studies. Two main selection scenarios are currently debated which explain the susceptibility or resistance to parasite infections either by the effects of (1) specific MHC alleles which are selected frequency-dependent in space and time or (2) a heterozygote or divergent allele advantage. So far, most studies have focused only on structural variance in co-evolutionary processes although this might not be the only trait subject to natural selection. In the present study, we analysed structural variance stretching from exon1 through exon3 of MHC class II DRB genes as well as genotypic expression variance in relation to the gastrointestinal helminth prevalence and infection intensity in wild yellow-necked mice (Apodemus flavicollis). We found support for the functional importance of specific alleles both on the sequence and expression level. By resampling a previously investigated study population we identified specific MHC alleles affected by temporal shifts in parasite pressure and recorded associated changes in allele frequencies. The allele Apfl-DRB*23 was associated with resistance to infections by the oxyurid nematode Syphacia stroma and at the same time with susceptibility to cestode infection intensity. In line with our expectation, MHC mRNA transcript levels tended to be higher in cestode-infected animals carrying the allele Apfl-DRB*23. However, no support for a heterozygote or divergent allele advantage on the sequence or expression level was detected. The individual amino acid distance of genotypes did not explain individual differences in parasite loads and the genetic distance had no effect on MHC genotype expression. For ongoing studies on the functional importance of expression variance in parasite resistance, allele

  8. CD40 promotes MHC class II expression on adipose tissue macrophages and regulates adipose tissue CD4+ T cells with obesity.

    PubMed

    Morris, David L; Oatmen, Kelsie E; Mergian, Taleen A; Cho, Kae Won; DelProposto, Jennifer L; Singer, Kanakadurga; Evans-Molina, Carmella; O'Rourke, Robert W; Lumeng, Carey N

    2016-06-01

    Obesity activates both innate and adaptive immune responses in adipose tissue, but the mechanisms critical for regulating these responses remain unknown. CD40/CD40L signaling provides bidirectional costimulatory signals between antigen-presenting cells and CD4(+) T cells, and CD40L expression is increased in obese humans. Therefore, we examined the contribution of CD40 to the progression of obesity-induced inflammation in mice. CD40 was highly expressed on adipose tissue macrophages in mice, and CD40/CD40L signaling promoted the expression of antigen-presenting cell markers in adipose tissue macrophages. When fed a high fat diet, Cd40-deficient mice had reduced accumulation of conventional CD4(+) T cells (Tconv: CD3(+)CD4(+)Foxp3(-)) in visceral fat compared with wild-type mice. By contrast, the number of regulatory CD4(+) T cells (Treg: CD3(+)CD4(+)Foxp3(+)) in lean and obese fat was similar between wild-type and knockout mice. Adipose tissue macrophage content and inflammatory gene expression in fat did not differ between obese wild-type and knockout mice; however, major histocompatibility complex class II and CD86 expression on adipose tissue macrophages was reduced in visceral fat from knockout mice. Similar results were observed in chimeric mice with hematopoietic Cd40-deficiency. Nonetheless, neither whole body nor hematopoietic disruption of CD40 ameliorated obesity-induced insulin resistance in mice. In human adipose tissue, CD40 expression was positively correlated with CD80 and CD86 expression in obese patients with type 2 diabetes. These findings indicate that CD40 signaling in adipose tissue macrophages regulates major histocompatibility complex class II and CD86 expression to control the expansion of CD4(+) T cells; however, this is largely dispensable for the development of obesity-induced inflammation and insulin resistance in mice. PMID:26658005

  9. Sibling rivalry: competition between MHC class II family members inhibits immunity.

    PubMed

    Denzin, Lisa K; Cresswell, Peter

    2013-01-01

    Peptide loading of major histocompatibility complex (MHC) class II molecules in the endosomes and lysosomes of antigen-presenting cells is catalyzed by human leukocyte antigen-DM (HLA-DM) and modulated by HLA-DO. In a structural study in this issue, Guce et al. show that HLA-DO is an MHC class II mimic and functions as a competitive and essentially irreversible inhibitor of HLA-DM activity, thereby inhibiting MHC class II antigen presentation. PMID:23288359

  10. Positive Regulatory Domain I-Binding Factor 1 mediates repression of the MHC Class II Transactivator (CIITA) type IV promoter

    PubMed Central

    Chen, Han; Gilbert, Carolyn A.; Hudson, John A.; Bolick, Sophia C.; Wright, Kenneth L.; Piskurich, Janet F.

    2006-01-01

    MHC class II transactivator (CIITA), a co-activator that controls MHC class II (MHC II) transcription, functions as the master regulator of MHC II expression. Persistent activity of the CIITA type III promoter (pIII), one of the four potential promoters of this gene, is responsible for constitutive expression of MHC II by B lymphocytes. In addition, IFN-γ induces expression of CIITA in these cells through the type IV promoter (pIV). Positive regulatory domain 1-binding factor 1 (PRDI-BF1), called B lymphocyte-induced maturation protein 1 (Blimp-1) in mice, represses the expression of CIITA pIII in plasma and multiple myeloma cells. To investigate regulation of CIITA pIV expression by PRDI-BF1 in the B lymphocyte lineage, protein/DNA binding studies, and functional promoter analyses were performed. PRDI-BF1 bound to the IRF-E site in CIITA pIV. Ectopic expression of either PRDI-BF1 or Blimp-1 repressed this promoter in B lymphocytes. In vitro binding and functional analyses of CIITA pIV demonstrated that the IFN regulatory factor-element (IRF-E) is the target of this repression. In vivo genomic footprint analysis demonstrated protein binding at the IRF-E site of CIITA pIV in U266 myeloma cells, which express PRDI-BF1. PRDI-BF1β, a truncated form of PRDI-BF1 that is co-expressed in myeloma cells, also bound to the IRF-E site and repressed CIITA pIV. These findings demonstrate for the first time that, in addition to silencing expression of CIITA pIII in B lymphocytes, PRDI-BF1 is capable of binding and suppressing CIITA pIV. PMID:16765445

  11. Diverse repertoire of the MHC class II-peptide complexes is required for presentation of viral superantigens.

    PubMed

    Golovkina, T; Agafonova, Y; Kazansky, D; Chervonsky, A

    2001-02-15

    Among other features, peptides affect MHC class II molecules, causing changes in the binding of bacterial superantigens (b-Sag). Whether peptides can alter binding of viral superantigens (v-Sag) to MHC class II was not known. Here we addressed the question of whether mutations limiting the diversity of peptides bound by the MHC class II molecules influenced the presentation of v-Sag and, subsequently, the life cycle of the mouse mammary tumor virus (MMTV). T cells reactive to v-Sag were found in mice lacking DM molecules as well as in A(b)Ep-transgenic mice in which MHC class II binding grooves were predominantly occupied by an invariant chain fragment or Ealpha(52-68) peptide, respectively. APCs from the mutant mice failed to present v-Sag, as determined by the lack of Sag-specific T cell activation, Sag-induced T cell deletion, and by the aborted MMTV infection. In contrast, mice that express I-A(b) with a variety of bound peptides presented v-Sag and were susceptible to MMTV infection. Comparison of v-Sag and b-Sag presentation by the same mutant cells suggested that presentation of v-Sag had requirements similar to that for presentation of toxic shock syndrome toxin-1. Thus, MHC class II peptide repertoire is critical for recognition of v-Sag by the T cells and affects the outcome of infection with a retrovirus. PMID:11160278

  12. Evaluating the role of HLA-DM in MHC II-peptide association reactions1

    PubMed Central

    Yin, Liusong; Maben, Zachary; Becerra, Aniuska; Stern, Lawrence J.

    2015-01-01

    Antigen presentation by major histocompatibility complex class II molecules (MHC II) to CD4+ T cells plays a key role in the regulation of the adaptive immune response. Loading of antigenic peptides onto MHC II is catalyzed by HLA-DM (DM), a non-classical MHC II molecule. The mechanism of DM-facilitated peptide loading is an outstanding problem in the field of antigen presentation. In this study we systemically explored possible kinetic mechanisms for DM-catalyzed peptide association, by measuring real time peptide association kinetics using fluorescence polarization assays and comparing the experimental data with numerically modeled peptide association reactions. We found that DM does not facilitate peptide association by stabilizing peptide-free MHC II against aggregation. Moreover, DM does not promote transition of an inactive peptide-averse conformation of MHC II to an active peptide-receptive conformation. Instead, DM forms an intermediate with MHC II that binds peptide with faster kinetics than MHC II in the absence of DM. In the absence of peptides, interaction of MHC II with DM leads to inactivation and formation of a peptide-averse form. This study provides novel insights into how DM efficiently catalyzes peptide loading during antigen presentation. PMID:26062997

  13. The relationship between MHC antigen expression and metastasis.

    PubMed

    Gopas, J; Rager-Zisman, B; Bar-Eli, M; Hämmerling, G J; Segal, S

    1989-01-01

    From the studies summarized here a complex picture of the role played by MHC products in determining tumorigenicity and metastasis is emerging. In order to be able to understand this relationship better, it is necessary to consider several factors. 1. Each tumor system or neoplastic tissue is unique, and its behavior reflects the influence of cell-specific characteristics, as well as its ability to modulate other cells and tissues--including cells belonging to the immune system--and also to be modulated by other cells and soluble factors. 2. Since metastasis formation is a multistep process in which only small subpopulations of tumor cells with complex and defined phenotypes are able to colonize secondary tissues, elimination of even one single phenotypic component of this structured process can easily reverse the metastatic capacity of the cells. Acquisition of metastatic ability, on the other hand, would be a more difficult task, since any new characteristic expressed by the cells or induced experimentally, such as gene transfection or results of IFN treatment, must be expressed in a temporal manner and in concert with other cellular characteristics. Therefore, an experimental protocol measuring a specific element in determining metastasis can easily produce conflicting results, depending on the type of cells and genetic background of the host studied. 3. The level of specific MHC products on tumor cells is one among many other cell characteristics that may determine the metastatic potential of cells. Moreover, each of the class 1 MHC products, and the relationship among them, including other than the classical K, L, or D products (Brickell et al., 1983), should be regarded as independent entities, with possible different regulatory roles in cell-cell recognition, in a general sense, which may be involved in determining invasiveness and homing as well as recognition by the immune system. 4. Both specific T-cell and nonspecific natural mediated immunity (which is

  14. Immunotoxin Against a Donor MHC Class II Molecule Induces Indefinite Survival of Murine Kidney Allografts.

    PubMed

    Brown, K; Nowocin, A K; Meader, L; Edwards, L A; Smith, R A; Wong, W

    2016-04-01

    Rejection of donor organs depends on the trafficking of donor passenger leukocytes to the secondary lymphoid organs of the recipient to elicit an immune response via the direct antigen presentation pathway. Therefore, the depletion of passenger leukocytes may be clinically applicable as a strategy to improve graft survival. Because major histocompatibility complex (MHC) class II(+) cells are most efficient at inducing immune responses, selective depletion of this population from donor grafts may dampen the alloimmune response and prolong graft survival. In a fully MHC mismatched mouse kidney allograft model, we describe the synthesis of an immunotoxin, consisting of the F(ab')2 fragment of a monoclonal antibody against the donor MHC class II molecule I-A(k) conjugated with the plant-derived ribosomal inactivating protein gelonin. This anti-I-A(k) gelonin immunotoxin depletes I-A(k) expressing cells specifically in vitro and in vivo. When given to recipients of kidney allografts, it resulted in indefinite graft survival with normal graft function, presence of Foxp3(+) cells within donor grafts, diminished donor-specific antibody formation, and delayed rejection of subsequent donor-type skin grafts. Strategies aimed at the donor arm of the immune system using agents such as immunotoxins may be a useful adjuvant to existing recipient-orientated immunosuppression. PMID:26799449

  15. Immunotoxin Against a Donor MHC Class II Molecule Induces Indefinite Survival of Murine Kidney Allografts

    PubMed Central

    Brown, K.; Nowocin, A. K.; Meader, L.; Edwards, L. A.; Smith, R. A.

    2016-01-01

    Rejection of donor organs depends on the trafficking of donor passenger leukocytes to the secondary lymphoid organs of the recipient to elicit an immune response via the direct antigen presentation pathway. Therefore, the depletion of passenger leukocytes may be clinically applicable as a strategy to improve graft survival. Because major histocompatibility complex (MHC) class II+ cells are most efficient at inducing immune responses, selective depletion of this population from donor grafts may dampen the alloimmune response and prolong graft survival. In a fully MHC mismatched mouse kidney allograft model, we describe the synthesis of an immunotoxin, consisting of the F(ab′)2 fragment of a monoclonal antibody against the donor MHC class II molecule I‐Ak conjugated with the plant‐derived ribosomal inactivating protein gelonin. This anti–I‐Ak gelonin immunotoxin depletes I‐Ak expressing cells specifically in vitro and in vivo. When given to recipients of kidney allografts, it resulted in indefinite graft survival with normal graft function, presence of Foxp3+ cells within donor grafts, diminished donor‐specific antibody formation, and delayed rejection of subsequent donor‐type skin grafts. Strategies aimed at the donor arm of the immune system using agents such as immunotoxins may be a useful adjuvant to existing recipient‐orientated immunosuppression. PMID:26799449

  16. Loss of MHC class-I expression in cervical carcinomas.

    PubMed

    Connor, M E; Stern, P L

    1990-12-15

    The expression of MHC class-I antigens was analysed in 67 cervical carcinoma biopsies; 16% of the biopsies showed complete or heterogeneous loss of HLA expression as judged by reactivity with antibodies recognizing monomorphic determinants of the class-I heavy chain bound to beta 2 microglobulin (beta 2m). In addition, other biopsies showed a loss in expression of particular allelic products: 23% for HLA-A2; 17% for HLA-A3; 23% for HLA-Bw4 and 19% for HLA-Bw6. Three biopsies showed changes at 2 alleles, 2 of which were at both HLA-A and -B loci. Down-regulation of class-I expression may be virally mediated and HPV DNA is frequently found in cervical carcinomas. However, there appeared to be no direct correlation between the detection of HPV 16 or 18 DNA in these tumours and changes in HLA expression. There was also no correlation with the expression of the oncofoetal antigen 5T4. Our results show that a significant proportion (at least 30%) of the cervical carcinomas showed some alteration in MHC class-I expression. Such changes may allow tumours to evade immune surveillance with more rapid progression. There was, however, no correlation with tumour type, degree of differentiation or stage of disease at presentation. PMID:2174412

  17. MHC class II allosteric site drugs: new immunotherapeutics for malignant, infectious and autoimmune diseases.

    PubMed

    Xu, M; Li, J; Gulfo, J V; Von Hofe, E; Humphreys, R E

    2001-01-01

    The discovery of the interactions of the 'Ii-Key' segment of the Ii protein with the major histocmpatibility complex (MHC) Class II allosteric site, which is adjacent to the antigenic peptide-binding site, creates therapeutic opportunities by regulating the antigenic peptide binding to MHC class II molecules. The binding of Ii-Key to the MHC class II allosteric site loosens the hold of the MHC Class II 'clamshell' on antigenic peptides and leads to highly efficient antigenic peptide charging to or releasing from the MHC class II antigenic peptide-binding groove. Ii-Key peptide-induced spilling of bound antigenic peptide, or replacement with inert blockers, leads to 'inert immunosuppression'. Highly efficient replacement of ambient with vaccine peptides by Ii-Key permits 'active immunosuppression' for antigen-specific control of autoimmune diseases in the absence of cytokines or adjuvants. On the other hand, active immunization against cancer or infectious disease can result from epitope replacement mediated by Ii-Key and accompanied by cytokines or other adjuvants. Finally, linking the Ii-Key peptide through a simple polymethylene bridge to an antigenic sequence vastly increases the potency of MHC Class II peptide vaccines. In summary, the discovery of the MHC class II allosteric site allows one to increase the efficiency of MHC class II-related, antigenic epitope-specific therapy for malignant, infectious, and autoimmune diseases. The focus of this review is on the mechanism and potential clinical use of such novel allosteric site-directed, Ii-key drugs. PMID:11439146

  18. Neuronal MHC Class I Expression Is Regulated by Activity Driven Calcium Signaling

    PubMed Central

    Peng, Yaqin; Liu, Jiane; Miao, Fengqin; Zhang, Jianqiong

    2015-01-01

    MHC class I (MHC-I) molecules are important components of the immune system. Recently MHC-I have been reported to also play important roles in brain development and synaptic plasticity. In this study, we examine the molecular mechanism(s) underlying activity-dependent MHC-I expression using hippocampal neurons. Here we report that neuronal expression level of MHC-I is dynamically regulated during hippocampal development after birth in vivo. Kainic acid (KA) treatment significantly increases the expression of MHC-I in cultured hippocampal neurons in vitro, suggesting that MHC-I expression is regulated by neuronal activity. In addition, KA stimulation decreased the expression of pre- and post-synaptic proteins. This down-regulation is prevented by addition of an MHC-I antibody to KA treated neurons. Further studies demonstrate that calcium-dependent protein kinase C (PKC) is important in relaying KA simulation activation signals to up-regulated MHC-I expression. This signaling cascade relies on activation of the MAPK pathway, which leads to increased phosphorylation of CREB and NF-κB p65 while also enhancing the expression of IRF-1. Together, these results suggest that expression of MHC-I in hippocampal neurons is driven by Ca2+ regulated activation of the MAPK signaling transduction cascade. PMID:26263390

  19. Towards universal structure-based prediction of class II MHC epitopes for diverse allotypes.

    PubMed

    Bordner, Andrew J

    2010-01-01

    The binding of peptide fragments of antigens to class II MHC proteins is a crucial step in initiating a helper T cell immune response. The discovery of these peptide epitopes is important for understanding the normal immune response and its misregulation in autoimmunity and allergies and also for vaccine design. In spite of their biomedical importance, the high diversity of class II MHC proteins combined with the large number of possible peptide sequences make comprehensive experimental determination of epitopes for all MHC allotypes infeasible. Computational methods can address this need by predicting epitopes for a particular MHC allotype. We present a structure-based method for predicting class II epitopes that combines molecular mechanics docking of a fully flexible peptide into the MHC binding cleft followed by binding affinity prediction using a machine learning classifier trained on interaction energy components calculated from the docking solution. Although the primary advantage of structure-based prediction methods over the commonly employed sequence-based methods is their applicability to essentially any MHC allotype, this has not yet been convincingly demonstrated. In order to test the transferability of the prediction method to different MHC proteins, we trained the scoring method on binding data for DRB1*0101 and used it to make predictions for multiple MHC allotypes with distinct peptide binding specificities including representatives from the other human class II MHC loci, HLA-DP and HLA-DQ, as well as for two murine allotypes. The results showed that the prediction method was able to achieve significant discrimination between epitope and non-epitope peptides for all MHC allotypes examined, based on AUC values in the range 0.632-0.821. We also discuss how accounting for peptide binding in multiple registers to class II MHC largely explains the systematically worse performance of prediction methods for class II MHC compared with those for class I MHC

  20. MHC2MIL: a novel multiple instance learning based method for MHC-II peptide binding prediction by considering peptide flanking region and residue positions

    PubMed Central

    2014-01-01

    Background Computational prediction of major histocompatibility complex class II (MHC-II) binding peptides can assist researchers in understanding the mechanism of immune systems and developing peptide based vaccines. Although many computational methods have been proposed, the performance of these methods are far from satisfactory. The difficulty of MHC-II peptide binding prediction comes mainly from the large length variation of binding peptides. Methods We develop a novel multiple instance learning based method called MHC2MIL, in order to predict MHC-II binding peptides. We deem each peptide in MHC2MIL as a bag, and some substrings of the peptide as the instances in the bag. Unlike previous multiple instance learning based methods that consider only instances of fixed length 9 (9 amino acids), MHC2MIL is able to deal with instances of both lengths of 9 and 11 (11 amino acids), simultaneously. As such, MHC2MIL incorporates important information in the peptide flanking region. For measuring the distances between different instances, furthermore, MHC2MIL explicitly highlights the amino acids in some important positions. Results Experimental results on a benchmark dataset have shown that, the performance of MHC2MIL is significantly improved by considering the instances of both 9 and 11 amino acids, as well as by emphasizing amino acids at key positions in the instance. The results are consistent with those reported in the literature on MHC-II peptide binding. In addition to five important positions (1, 4, 6, 7 and 9) for HLA(human leukocyte antigen, the name of MHC in Humans) DR peptide binding, we also find that position 2 may play some roles in the binding process. By using 5-fold cross validation on the benchmark dataset, MHC2MIL outperforms two state-of-the-art methods of MHC2SK and NN-align with being statistically significant, on 12 HLA DP and DQ molecules. In addition, it achieves comparable performance with MHC2SK and NN-align on 14 HLA DR molecules. MHC2MIL

  1. Bacterial superantigens promote acute nasopharyngeal infection by Streptococcus pyogenes in a human MHC Class II-dependent manner.

    PubMed

    Kasper, Katherine J; Zeppa, Joseph J; Wakabayashi, Adrienne T; Xu, Stacey X; Mazzuca, Delfina M; Welch, Ian; Baroja, Miren L; Kotb, Malak; Cairns, Ewa; Cleary, P Patrick; Haeryfar, S M Mansour; McCormick, John K

    2014-05-01

    Establishing the genetic determinants of niche adaptation by microbial pathogens to specific hosts is important for the management and control of infectious disease. Streptococcus pyogenes is a globally prominent human-specific bacterial pathogen that secretes superantigens (SAgs) as 'trademark' virulence factors. SAgs function to force the activation of T lymphocytes through direct binding to lateral surfaces of T cell receptors and class II major histocompatibility complex (MHC-II) molecules. S. pyogenes invariably encodes multiple SAgs, often within putative mobile genetic elements, and although SAgs are documented virulence factors for diseases such as scarlet fever and the streptococcal toxic shock syndrome (STSS), how these exotoxins contribute to the fitness and evolution of S. pyogenes is unknown. Here we show that acute infection in the nasopharynx is dependent upon both bacterial SAgs and host MHC-II molecules. S. pyogenes was rapidly cleared from the nasal cavity of wild-type C57BL/6 (B6) mice, whereas infection was enhanced up to ∼10,000-fold in B6 mice that express human MHC-II. This phenotype required the SpeA superantigen, and vaccination with an MHC -II binding mutant toxoid of SpeA dramatically inhibited infection. Our findings indicate that streptococcal SAgs are critical for the establishment of nasopharyngeal infection, thus providing an explanation as to why S. pyogenes produces these potent toxins. This work also highlights that SAg redundancy exists to avoid host anti-SAg humoral immune responses and to potentially overcome host MHC-II polymorphisms. PMID:24875883

  2. Dissection of the interferon gamma-MHC class II signal transduction pathway reveals that type I and type II interferon systems share common signalling component(s).

    PubMed Central

    Loh, J E; Chang, C H; Fodor, W L; Flavell, R A

    1992-01-01

    We have used a herpes virus thymidine kinase (HSV-TK) based metabolic selection system to isolate mutants defective in the interferon gamma mediated induction of the MHC class II promoter. All the mutations act in trans and result in no detectable induction of MHC and invariant chain (Ii) gene expression. Scatchard analysis indicates that the mutants have a normal number of surface IFN gamma receptors with the same affinity constant. The mutants fall into two broad categories. One class of mutants is still able to induce MHC class I, IRF-1, 9-27, 1-8 and GBP genes by IFN gamma. A second class of mutants is defective for the IFN gamma induction of all the genes tested; surprisingly, the IFN alpha/beta induction of MHC class I, 9-27, ISG54 and ISG15 genes is also defective in these mutants, although different members of this class can be discriminated by the response of the GBP and IRF-1 genes to type I interferons. These data demonstrate that the signalling pathways of both type I and type II interferon systems share common signal transduction component(s). These mutants will be useful for the study of IFN gamma regulation of class II genes and Ii chain, and to elucidate molecular components of type I and type II interferon signal transduction. Images PMID:1314162

  3. Vaccinia virus infection induces dendritic cell maturation but inhibits antigen presentation by MHC class II

    PubMed Central

    Yao, Yongxue; Li, Ping; Singh, Pratibha; Thiele, Allison T.; Wilkes, David S.; Renukaradhya, Gourapura J.; Brutkiewicz, Randy R.; Travers, Jeffrey B.; Luker, Gary D.; Hong, Soon-Cheol; Blum, Janice S.; Chang, Cheong-Hee

    2007-01-01

    Vaccinia virus (VV) infection is known to inhibit dendritic cells (DC) functions in vitro. Paradoxically, VV is also highly immunogenic and thus has been used as a vaccine. In the present study, we investigated the effects of an in vivo VV infection on DC function by focusing on early innate immunity. Our data indicated that DC are activated upon in vivo VV infection of mice. Splenic DC from VV-infected mice expressed elevated levels of MHC class I and co-stimulatory molecules on their cell surface and exhibited the enhanced potential to produce cytokines upon LPS stimulation. DC from VV-infected mice also expressed a high level of interferon-β. However, a VV infection resulted in the down-regulation of MHC class II expression and the impairment of antigen presentation to CD4 T cells by DC. Thus, during the early stage of a VV infection, although DC are impaired in some of the critical antigen presentation functions, they can promote innate immune defenses against viral infection. PMID:17678637

  4. Differences in MHC and TAP-1 expression in cervical cancer lymph node metastases as compared with the primary tumours.

    PubMed Central

    Cromme, F. V.; van Bommel, P. F.; Walboomers, J. M.; Gallee, M. P.; Stern, P. L.; Kenemans, P.; Helmerhorst, T. J.; Stukart, M. J.; Meijer, C. J.

    1994-01-01

    In previous studies we have shown down-regulation of class I major histocompatibility complex (MHC) expression in a significant proportion of primary cervical carcinomas, which was found to be strongly correlated with loss of expression of the transporter associated with antigen presentation (TAP). By contrast, class II MHC expression was frequently up-regulated on neoplastic keratinocytes in these malignancies. In order to investigate whether these changes are associated with biological behaviour of the tumours, 20 cervical carcinomas were analyzed for MHC (HLA-A, HLA-B/C, HLA-DR) and TAP-1 expression in the primary tumours and in lymph node metastases by immunohistochemistry. The results showed a significant increase in the prevalence of HLA-A and HLA-B/C down-regulation in metastasised neoplastic cells as compared with the primary tumour (P = 0.01). In all cases this was accompanied by loss of TAP-1 expression. Up-regulated HLA-DR expression was found exclusively in primary tumours and was absent in the corresponding metastases (P = 0.002). These data are consistent with the hypothesis that loss of TAP-1 and the consequent down-regulation of class I MHC expression provides a selective advantage for neoplastic cervical cells during metastasis. Furthermore, the lack of class II MHC expression in metastasised cells either reflects a different local lymphokine production or indicates that these cells may have escaped CD4+ cytotoxic T-lymphocyte (CTL)-mediated killing. Images Figure 1 Figure 2 Figure 3 PMID:8198988

  5. Use of MHC class II tetramers to investigate CD4+ T cell responses: problems and solutions.

    PubMed

    Cecconi, Virginia; Moro, Monica; Del Mare, Sara; Dellabona, Paolo; Casorati, Giulia

    2008-11-01

    MHC-class I tetramers technology enabled the characterization of peptide-specific T cells at the single cell level in a variety of studies. Several laboratories have also developed MHC-class II multimers to characterize Ag-specific CD4+ T cells. However, the generation and use of MHC-class II multimers seems more problematic than that of MHC-I multimers. We have generated HLA-DR*1101 tetramers in a versatile empty form, which can be loaded after purification with peptides of interest. We discuss the impact of critical biological and structural parameters for the optimal staining of Ag-specific CD4+ T cells using HLA-DR*1101 tetramers, such as: (i) activation state of CD4+ T cells; (ii) membrane trafficking in the target CD4+ T cells; (iii) binding characteristics of the loaded CD4 epitope. Our data indicate that reorganization of TCR on the plasma membrane upon CD4+ T cell activation, as well as an homogenous binding frame of the CD4 epitopes to the soluble HLA-DR monomer, are critical for a stable TCR/MHC-class II tetramer interaction. These factors, together with the low frequencies and affinities of specific CD4+ T cells, explain the need for in vitro expansion or ex vivo enrichment of specific T cells for the optimal visualization with MHC-class II tetramers. PMID:18612991

  6. Partial MHC class II constructs inhibit MIF/CD74 binding and downstream effects.

    PubMed

    Benedek, Gil; Meza-Romero, Roberto; Andrew, Shayne; Leng, Lin; Burrows, Gregory G; Bourdette, Dennis; Offner, Halina; Bucala, Richard; Vandenbark, Arthur A

    2013-05-01

    MIF and its receptor, CD74, are pivotal regulators of the immune system. Here, we demonstrate for the first time that partial MHC class II constructs comprised of linked β1α1 domains with covalently attached antigenic peptides (also referred to as recombinant T-cell receptor ligands - RTLs) can inhibit MIF activity by not only blocking the binding of rhMIF to immunopurified CD74, but also downregulating CD74 cell-surface expression. This bifunctional inhibition of MIF/CD74 interactions blocked downstream MIF effects, including enhanced secretion of proinflammatory cytokines, anti-apoptotic activity, and inhibition of random migration that all contribute to the reversal of clinical and histological signs of EAE. Moreover, we demonstrate that enhanced CD74 cell-surface expression on monocytes in mice with EAE and subjects with multiple sclerosis can be downregulated by humanized RTLs, resulting in reduced MIF binding to the cells. Thus, binding of partial MHC complexes to CD74 blocks both the accessibility and availability of CD74 for MIF binding and downstream inflammatory activity. PMID:23576302

  7. Partial MHC class II constructs inhibit MIF/CD74 binding and downstream effects

    PubMed Central

    Benedek, Gil; Meza-Romero, Roberto; Andrew, Shayne; Leng, Lin; Burrows, Gregory G.; Bourdette, Dennis; Offner, Halina; Bucala, Richard; Vandenbark, Arthur A.

    2013-01-01

    Macrophage migration inhibitory factor (MIF) and its receptor, CD74, are pivotal regulators of the immune system. Here we demonstrate for the first time that partial MHC class II constructs comprised of linked β1α1 domains with covalently attached antigenic peptides (also referred to as recombinant T-cell receptor ligands - RTLs) can inhibit MIF activity by not only blocking the binding of rhMIF to immunopurified CD74, but also down-regulating CD74 cell-surface expression. This bi-functional inhibition of MIF/CD74 interactions blocked downstream MIF effects, including enhanced secretion of proinflammatory cytokines, anti-apoptotic activity and inhibition of random migration that all contribute to the reversal of clinical and histological signs of experimental autoimmune encephalomyelitis (EAE). Moreover, we demonstrate that enhanced CD74 cell surface expression on monocytes in mice with EAE and subjects with multiple sclerosis (MS) can be down-regulated by humanized RTLs, resulting in reduced MIF binding to the cells. Thus, binding of partial MHC complexes to CD74 blocks both the accessibility and availability of CD74 for MIF binding and downstream inflammatory activity. PMID:23576302

  8. Quantitating MHC class II trafficking in primary dendritic cells using imaging flow cytometry

    PubMed Central

    Hennies, Cassandra M.; Lehn, Maria A.; Janssen, Edith M.

    2015-01-01

    Presentation of antigenic peptides in MHC class II (MHCII) on dendritic cells (DCs) is the first step in the activation of antigen-specific CD4+T cells. The expression of surface MHCII-peptide complexes is tightly regulated as the frequency of MHCII-peptide complexes can affect the magnitude, as well as the phenotype of the ensuing CD4+T cell response. The surface MHCII-peptide levels are determined by the balance between expression of newly generated complexes, complex internalization, and their subsequent re-emergence or degradation. However, the molecular mechanisms that underpin these processes are still poorly understood. Here we describe a multispectral imaging flow cytometry assay to visualize MHCII trafficking that can be used as a tool to dissect the molecular mechanisms that regulate MHCII homeostasis in primary mouse and human DCs. PMID:25967952

  9. High Levels of MeCP2 Depress MHC Class I Expression in Neuronal Cells

    PubMed Central

    Miralvès, Julie; Magdeleine, Eddy; Kaddoum, Lara; Brun, Hélène; Peries, Sophie; Joly, Etienne

    2007-01-01

    Background The expression of MHC class I genes is repressed in mature neurons. The molecular basis of this regulation is poorly understood, but the genes are particularly rich in CpG islands. MeCP2 is a transcriptional repressor that binds to methylated CpG dinucleotides; mutations in this protein also cause the neurodevelopmental disease called Rett syndrome. Because MHC class I molecules play a role in neuronal connectivity, we hypothesised that MeCP2 might repress MHC class I expression in the CNS and that this might play a role in the pathology of Rett syndrome. Methodology We show here that transiently transfected cells expressing high levels of MeCP2 specifically downregulate cell-surface expression of MHC class I molecules in the neuronal cell line N2A and they prevent the induction of MHC class I expression in response to interferon in these cells, supporting our first hypothesis. Surprisingly, however, overexpression of the mutated forms of MeCP2 that cause Rett syndrome had a similar effect on MHC class I expression as the wild-type protein. Immunohistological analyses of brain slices from MECP2 knockout mice (the MeCP2tm1.1Bird strain) demonstrated a small but reproducible increase in MHC class I when compared to their wild type littermates, but we found no difference in MHC class I expression in primary cultures of mixed glial cells (mainly neurons and astrocytes) from the knockout and wild-type mice. Conclusion These data suggest that high levels of MeCP2, such as those found in mature neurons, may contribute to the repression of MHC expression, but we find no evidence that MeCP2 regulation of MHC class I is important for the pathogenesis of Rett syndrome. PMID:18159237

  10. Coordinated changes of histone modifications and HDAC mobilization regulate the induction of MHC class II genes by Trichostatin A

    PubMed Central

    2006-01-01

    The deacetylase inhibitor Trichostatin A (TSA) induces the transcription of the Major Histocompatibility Class II (MHC II) DRA gene in a way independent of the master coactivator CIITA. To analyze the molecular mechanisms by which this epigenetic regulator stimulates MHC II expression, we used chromatin immunoprecipitation (ChIP) assays to monitor the alterations in histone modifications that correlate with DRA transcription after TSA treatment. We found that a dramatic increase in promoter linked histone acetylation is followed by an increase in Histone H3 lysine 4 methylation and a decrease of lysine 9 methylation. Fluorescence recovery after photobleaching (FRAP) experiments showed that TSA increases the mobility of HDAC while decreasing the mobility of the class II enhanceosome factor RFX5. These data, in combination with ChIP experiments, indicate that the TSA-mediated induction of DRA transcription involves HDAC relocation and enhanceosome stabilization. In order to gain a genome-wide view of the genes responding to inhibition of deacetylases, we compared the transcriptome of B cells before and after TSA treatment using Affymetrix microarrays. This analysis showed that in addition to the DRA gene, the entire MHC II family and the adjacent histone cluster that are located in chromosome 6p21-22 locus are strongly induced by TSA. A complex pattern of gene reprogramming by TSA involves immune recognition, antiviral, apoptotic and inflammatory pathways and extends the rationale for using Histone Deacetylase Inhibitors (HDACi) to modulate the immune response. PMID:16452299

  11. Downregulation of MHC-I expression is prevalent but reversible in Merkel cell carcinoma

    PubMed Central

    Paulson, Kelly G.; Tegeder, Andrew; Willmes, Christoph; Iyer, Jayasri G; Afanasiev, Olga K.; Schrama, David; Koba, Shinichi; Thibodeau, Renee; Nagase, Kotaro; Simonson, William T; Seo, Aaron; Koelle, David M.; Madeleine, Margaret; Bhatia, Shailender; Nakajima, Hideki; Sano, Shigetoshi; Hardwick, James S.; Disis, Mary L.; Cleary, Michele A; Becker, Jürgen C.; Nghiem, Paul

    2014-01-01

    Merkel cell carcinoma (MCC) is an aggressive, polyomavirus-associated skin cancer. Robust cellular immune responses are associated with excellent outcomes in MCC patients, but these responses are typically absent. We determined the prevalence and reversibility of class I MHC (MHC-I) downregulation in MCC, a potentially reversible immune evasion mechanism. Cell surface MHC-I expression was assessed on 5 MCC cell lines using flow cytometry as well as immunohistochemistry on tissue microarrays representing 114 patients. Three additional patients were included that had received intralesional interferon treatment and had evaluable specimens before and after treatment. mRNA expression analysis of antigen presentation pathway genes from 35 MCC tumors was used to examine mechanisms of downregulation. 84% of MCCs (total n=114) demonstrated reduced MHC-I expression as compared to surrounding tissues, and 51% had poor or undetectable MHC-1 expression. Expression of MHC-I was lower in polyomavirus-positive MCCs as compared to virus-negative MCCs (p<0.01). The MHC-I downregulation mechanism was multifactorial and did not depend solely on HLA gene expression. Treatment of MCC cell lines with ionizing radiation, etoposide, or interferon (IFN) resulted in MHC-I upregulation, with IFNs strongly upregulating MHC-I expression in vitro and in 3 of 3 patients treated with intralesional IFNs. MCC tumors may be amenable to immunotherapy, but downregulation of MHC-I is frequently present in these tumors, particularly those that are polyomavirus-positive. This downregulation is reversible with any of several clinically available treatments that may thus promote the effectiveness of immune stimulating therapies for MCC. PMID:25116754

  12. Trans-species polymorphism and selection in the MHC class II DRA genes of domestic sheep.

    PubMed

    Ballingall, Keith T; Rocchi, Mara S; McKeever, Declan J; Wright, Frank

    2010-01-01

    Highly polymorphic genes with central roles in lymphocyte mediated immune surveillance are grouped together in the major histocompatibility complex (MHC) in higher vertebrates. Generally, across vertebrate species the class II MHC DRA gene is highly conserved with only limited allelic variation. Here however, we provide evidence of trans-species polymorphism at the DRA locus in domestic sheep (Ovis aries). We describe variation at the Ovar-DRA locus that is far in excess of anything described in other vertebrate species. The divergent DRA allele (Ovar-DRA*0201) differs from the sheep reference sequences by 20 nucleotides, 12 of which appear non-synonymous. Furthermore, DRA*0201 is paired with an equally divergent DRB1 allele (Ovar-DRB1*0901), which is consistent with an independent evolutionary history for the DR sub-region within this MHC haplotype. No recombination was observed between the divergent DRA and B genes in a range of breeds and typical levels of MHC class II DR protein expression were detected at the surface of leukocyte populations obtained from animals homozygous for the DRA*0201, DRB1*0901 haplotype. Bayesian phylogenetic analysis groups Ovar-DRA*0201 with DRA sequences derived from species within the Oryx and Alcelaphus genera rather than clustering with other ovine and caprine DRA alleles. Tests for Darwinian selection identified 10 positively selected sites on the branch leading to Ovar-DRA*0201, three of which are predicted to be associated with the binding of peptide antigen. As the Ovis, Oryx and Alcelaphus genera have not shared a common ancestor for over 30 million years, the DRA*0201 and DRB1*0901 allelic pair is likely to be of ancient origin and present in the founding population from which all contemporary domestic sheep breeds are derived. The conservation of the integrity of this unusual DR allelic pair suggests some selective advantage which is likely to be associated with the presentation of pathogen antigen to T-cells and the

  13. Trans-Species Polymorphism and Selection in the MHC Class II DRA Genes of Domestic Sheep

    PubMed Central

    Ballingall, Keith T.; Rocchi, Mara S.; McKeever, Declan J.; Wright, Frank

    2010-01-01

    Highly polymorphic genes with central roles in lymphocyte mediated immune surveillance are grouped together in the major histocompatibility complex (MHC) in higher vertebrates. Generally, across vertebrate species the class II MHC DRA gene is highly conserved with only limited allelic variation. Here however, we provide evidence of trans-species polymorphism at the DRA locus in domestic sheep (Ovis aries). We describe variation at the Ovar-DRA locus that is far in excess of anything described in other vertebrate species. The divergent DRA allele (Ovar-DRA*0201) differs from the sheep reference sequences by 20 nucleotides, 12 of which appear non-synonymous. Furthermore, DRA*0201 is paired with an equally divergent DRB1 allele (Ovar-DRB1*0901), which is consistent with an independent evolutionary history for the DR sub-region within this MHC haplotype. No recombination was observed between the divergent DRA and B genes in a range of breeds and typical levels of MHC class II DR protein expression were detected at the surface of leukocyte populations obtained from animals homozygous for the DRA*0201, DRB1*0901 haplotype. Bayesian phylogenetic analysis groups Ovar-DRA*0201 with DRA sequences derived from species within the Oryx and Alcelaphus genera rather than clustering with other ovine and caprine DRA alleles. Tests for Darwinian selection identified 10 positively selected sites on the branch leading to Ovar-DRA*0201, three of which are predicted to be associated with the binding of peptide antigen. As the Ovis, Oryx and Alcelaphus genera have not shared a common ancestor for over 30 million years, the DRA*0201 and DRB1*0901 allelic pair is likely to be of ancient origin and present in the founding population from which all contemporary domestic sheep breeds are derived. The conservation of the integrity of this unusual DR allelic pair suggests some selective advantage which is likely to be associated with the presentation of pathogen antigen to T-cells and the

  14. Susceptibility of amphibians to chytridiomycosis is associated with MHC class II conformation

    PubMed Central

    Bataille, Arnaud; Cashins, Scott D.; Grogan, Laura; Skerratt, Lee F.; Hunter, David; McFadden, Michael; Scheele, Benjamin; Brannelly, Laura A.; Macris, Amy; Harlow, Peter S.; Bell, Sara; Berger, Lee; Waldman, Bruce

    2015-01-01

    The pathogenic chytrid fungus Batrachochytrium dendrobatidis (Bd) can cause precipitous population declines in its amphibian hosts. Responses of individuals to infection vary greatly with the capacity of their immune system to respond to the pathogen. We used a combination of comparative and experimental approaches to identify major histocompatibility complex class II (MHC-II) alleles encoding molecules that foster the survival of Bd-infected amphibians. We found that Bd-resistant amphibians across four continents share common amino acids in three binding pockets of the MHC-II antigen-binding groove. Moreover, strong signals of selection acting on these specific sites were evident among all species co-existing with the pathogen. In the laboratory, we experimentally inoculated Australian tree frogs with Bd to test how each binding pocket conformation influences disease resistance. Only the conformation of MHC-II pocket 9 of surviving subjects matched those of Bd-resistant species. This MHC-II conformation thus may determine amphibian resistance to Bd, although other MHC-II binding pockets also may contribute to resistance. Rescuing amphibian biodiversity will depend on our understanding of amphibian immune defence mechanisms against Bd. The identification of adaptive genetic markers for Bd resistance represents an important step forward towards that goal. PMID:25808889

  15. Susceptibility of amphibians to chytridiomycosis is associated with MHC class II conformation.

    PubMed

    Bataille, Arnaud; Cashins, Scott D; Grogan, Laura; Skerratt, Lee F; Hunter, David; McFadden, Michael; Scheele, Benjamin; Brannelly, Laura A; Macris, Amy; Harlow, Peter S; Bell, Sara; Berger, Lee; Waldman, Bruce

    2015-04-22

    The pathogenic chytrid fungus Batrachochytrium dendrobatidis (Bd) can cause precipitous population declines in its amphibian hosts. Responses of individuals to infection vary greatly with the capacity of their immune system to respond to the pathogen. We used a combination of comparative and experimental approaches to identify major histocompatibility complex class II (MHC-II) alleles encoding molecules that foster the survival of Bd-infected amphibians. We found that Bd-resistant amphibians across four continents share common amino acids in three binding pockets of the MHC-II antigen-binding groove. Moreover, strong signals of selection acting on these specific sites were evident among all species co-existing with the pathogen. In the laboratory, we experimentally inoculated Australian tree frogs with Bd to test how each binding pocket conformation influences disease resistance. Only the conformation of MHC-II pocket 9 of surviving subjects matched those of Bd-resistant species. This MHC-II conformation thus may determine amphibian resistance to Bd, although other MHC-II binding pockets also may contribute to resistance. Rescuing amphibian biodiversity will depend on our understanding of amphibian immune defence mechanisms against Bd. The identification of adaptive genetic markers for Bd resistance represents an important step forward towards that goal. PMID:25808889

  16. Intranasal antigen targeting to MHC class II molecules primes local IgA and serum IgG antibody responses in mice.

    PubMed

    Snider, D P; Underdown, B J; McDermott, M R

    1997-03-01

    Covalent conjugates of hen egg lysozyme (HEL) and anti-major histocompatibility complex (MHC) class II monoclonal antibodies (mAb) were used to immunize mice intranasally. Three weeks after intranasal (IN) priming, mice responded rapidly to IN challenge with a mixture of HEL and cholera toxin (CT), by producing large titres of anti-HEL IgA and IgG1 antibody in serum, and IgA antibody in nasal secretions. No secondary response to HEL plus CT occurred in mice that received no priming or mice primed with HEL alone. The secondary serum IgG antibody response was dominated by the IgG1 subclass. HEL combined with CT adjuvant worked much better than HEL alone in producing the secondary response. Control conjugates, containing an IgG isotype-matched mAb without specificity for mouse tissues, provided poor priming. Mice expressing MHC class II molecules, to which the anti-MHC II mAb could not bind, produced a weak antibody response compared with those that expressed the appropriate. MHC class II molecule. Our results demonstrate that immunotargeting to MHC class II molecules via the IN route allows priming of the local IgA and circulating IgG antibody, and indicate that this technique is a feasible approach for delivery of subunit vaccines in the upper respiratory tract. PMID:9155636

  17. Exploring genome-wide datasets of MHC class II antigen presentation.

    PubMed

    Wijdeven, Ruud H; Bakker, Jeroen M; Paul, Petra; Neefjes, Jacques

    2013-09-01

    MHC class II molecules (MHCII) are critical for presenting antigens to CD4(+) T-cells. They control ignition of CD4(+) T cells and are as such involved in most auto-immune diseases. To define proteins and pathways controlling MHCII antigen presentation and expression, we performed a genome-wide flow cytometry based RNAi screen. Hits were subsequently classified by two screens that monitored the intracellular distribution and transcription of MHCII. This multi-dimensional approach allowed subclassification of hits into functional groups as a first step to defining new pathways controlling MHCII antigen presentation. The datasets from this screen are used as a template for several follow-up studies. This overview focuses on how data from genome-wide screens can be used for target-lead finding, data mining, systems biology and systematic cell biology. PMID:23137594

  18. Evolution of MHC class II E beta diversity within the genus Peromyscus.

    PubMed Central

    Richman, Adam D; Herrera, L Gerardo; Nash, Deanna

    2003-01-01

    Progress in understanding the evolution of variation at the MHC has been slowed by an inability to assess the relative roles of mutation vs. intragenic recombination in contributing to observed polymorphism. Recent theoretical advances now permit a quantitative treatment of the problem, with the result that the amount of recombination is at least an order of magnitude greater than that of mutation in the history of class II genes. We suggest that this insight allows progress in evaluating the importance of other factors affecting the evolution of the MHC. We investigated the evolution of MHC class II E beta sequence diversity in the genus Peromyscus. We find evidence for extensive recombination in the history of these sequences. Nevertheless, it appears that intragenic recombination alone is insufficient to account for evolution of MHC diversity in Peromyscus. Significant differences in silent variation among subgenera arose over a relatively short period of time, with little subsequent change. We argue that these observations are consistent with the effects of historical population bottleneck(s). Population restrictions may explain general features of MHC evolution, including the large amount of recombination in the history of MHC genes, because intragenic recombination may efficiently regenerate allelic polymorphism following a population constriction. PMID:12750340

  19. Mutant MHC class II epitopes drive therapeutic immune responses to cancer

    PubMed Central

    Kreiter, Sebastian; Vormehr, Mathias; van de Roemer, Niels; Diken, Mustafa; Löwer, Martin; Diekmann, Jan; Boegel, Sebastian; Schrörs, Barbara; Vascotto, Fulvia; Castle, John C.; Tadmor, Arbel D.; Schoenberger, Stephen P.; Huber, Christoph; Türeci, Özlem; Sahin, Ugur

    2016-01-01

    Tumour-specific mutations are ideal targets for cancer immunotherapy as they lack expression in healthy tissues and can potentially be recognized as neo-antigens by the mature T-cell repertoire. Their systematic targeting by vaccine approaches, however, has been hampered by the fact that every patient’s tumour possesses a unique set of mutations (‘the mutanome’) that must first be identified. Recently, we proposed a personalized immunotherapy approach to target the full spectrum of a patient’s individual tumour-specific mutations1. Here we show in three independent murine tumour models that a considerable fraction of non-synonymous cancer mutations is immunogenic and that, unexpectedly, the majority of the immunogenic mutanome is recognized by CD4+ T cells. Vaccination with such CD4+ immunogenic mutations confers strong antitumour activity. Encouraged by these findings, we established a process by which mutations identified by exome sequencing could be selected as vaccine targets solely through bioinformatic prioritization on the basis of their expression levels and major histocompatibility complex (MHC) class II-binding capacity for rapid production as synthetic poly-neo-epitope messenger RNA vaccines. We show that vaccination with such polytope mRNA vaccines induces potent tumour control and complete rejection of established aggressively growing tumours in mice. Moreover, we demonstrate that CD4+ T cell neo-epitope vaccination reshapes the tumour microenvironment and induces cytotoxic T lymphocyte responses against an independent immunodominant antigen in mice, indicating orchestration of antigen spread. Finally, we demonstrate an abundance of mutations predicted to bind to MHC class II in human cancers as well by employing the same predictive algorithm on corresponding human cancer types. Thus, the tailored immunotherapy approach introduced here may be regarded as a universally applicable blueprint for comprehensive exploitation of the substantial neo

  20. Dual MHC class I and class II restriction of a single T cell receptor: distinct modes of tolerance induction by two classes of autoantigens.

    PubMed

    Arsov, I; Vukmanović, S

    1999-02-15

    In the final stages of thymic development, immature T cells undergo three distinct processes (positive selection, negative selection, and lineage commitment) that all depend on interactions of thymocyte TCRs with MHC molecules. It is currently thought that TCRs are preferentially restricted by either MHC class I or class II molecules. In this report, we present direct evidence that the TCR previously described as H-Y/H-2Db specific cross-reacts with H-2IAb if expressed in CD4+ cells. We also demonstrate an increase in thymocyte numbers in H-Y TCR-trangenic mice deficient in MHC class II, suggesting a relatively discrete form of negative selection by MHC class II compared with that induced by H-Y/H-2Db. We propose that inability to generate CD4+ T cells expressing H-Y TCR in different experimental settings may be due to tolerance to self-MHC class II. These results, therefore, support an intriguing possibility that tolerance to self may influence and/or interfere with the outcome of the lineage commitment. PMID:9973472

  1. Patterns of MHC-G-Like and MHC-B Diversification in New World Monkeys

    PubMed Central

    Lugo, Juan S.; Cadavid, Luis F.

    2015-01-01

    The MHC class I (MHC-I) region in New World monkeys (Platyrrhini) has remained relatively understudied. To evaluate the diversification patterns and transcription behavior of MHC-I in Platyrrhini, we first analyzed public genomic sequences from the MHC-G-like subregion in Saimiri boliviensis, Ateles geoffroyi and Callicebus moloch, and from the MHC-B subregion in Saimiri boliviensis. While S. boliviensis showed multiple copies of both MHC-G-like (10) and –B (15) loci, A. geoffroyi and C. moloch had only three and four MHC-G-like genes, respectively, indicating that not all Platyrrhini species have expanded their MHC-I loci. We then sequenced MHC-G-like and -B cDNAs from nine Platyrrhini species, recovering two to five unique cDNAs per individual for both loci classes. In two Saguinus species, however, no MHC-B cDNAs were found. In phylogenetic trees, MHC-G-like cDNAs formed genus-specific clusters whereas the MHC-B cDNAs grouped by Platyrrhini families, suggesting a more rapid diversification of the former. Furthermore, cDNA sequencing in 12 capuchin monkeys showed that they transcribe at least four MHC-G-like and five MHC-B polymorphic genes, showing haplotypic diversity for gene copy number and signatures of positive natural selection at the peptide binding region. Finally, a quantitative index for MHC:KIR affinity was proposed and tested to predict putative interacting pairs. Altogether, our data indicate that i) MHC-I genes has expanded differentially among Platyrrhini species, ii) Callitrichinae (tamarins and marmosets) MHC-B loci have limited or tissue-specific expression, iii) MHC-G-like genes have diversified more rapidly than MHC-B genes, and iv) the MHC-I diversity is generated mainly by genetic polymorphism and gene copy number variation, likely promoted by natural selection for ligand binding. PMID:26121030

  2. Characterization of MHC class I and II genes in a subantarctic seabird, the blue petrel, Halobaena caerulea (Procellariiformes).

    PubMed

    Strandh, Maria; Lannefors, Mimi; Bonadonna, Francesco; Westerdahl, Helena

    2011-10-01

    The great polymorphism observed in the major histocompatibility complex (MHC) genes is thought to be maintained by pathogen-mediated selection possibly combined with MHC-disassortative mating, guided by MHC-determined olfactory cues. Here, we partly characterize the MHC class I and II B of the blue petrel, Halobaena caerulea (Procellariiformes), a bird with significant olfactory abilities that lives under presumably low pathogen burdens in Subantarctica. Blue petrels are long-lived, monogamous birds which suggest the necessity of an accurate mate choice process. The species is ancestral to songbirds (Passeriformes; many MHC loci), although not to gamefowls (Galliformes; few MHC loci). Considering the phylogenetic relationships and the low subantarctic pathogen burden, we expected few rather than many MHC loci in the blue petrel. However, when we analysed partial MHC class I and class II B cDNA and gDNA sequences we found evidence for as many as at least eight MHC class I loci and at least two class II B loci. These class I and II B sequences showed classical MHC characteristics, e.g. high nucleotide diversity, especially in putative peptide-binding regions where signatures of positive selection was detected. Trans-species polymorphism was found between MHC class II B sequences of the blue petrel and those of thin-billed prion, Pachyptila belcheri, two species that diverged ∼25 MYA. The observed MHC allele richness in the blue petrel may well serve as a basis for mate choice, especially since olfactory discrimination of MHC types may be possible in this species. PMID:21607694

  3. Induction of class II major histocompatibility complex expression in human multiple myeloma cells by retinoid.

    PubMed

    Sanda, Takaomi; Iida, Shinsuke; Kayukawa, Satoshi; Ueda, Ryuzo

    2007-01-01

    Class II major histocompatibility complex (MHC II) is normally silenced in plasma/multiple myeloma (MM) cells at the transcriptional level through downregulation of class II transactivator (CIITA), allowing MM cells to escape from immunological responses. Here we demonstrate that a retinoic acid receptor-alpha/beta-selective retinoid Am80 (tamibarotene) could induce the expression of functional MHC II molecules in human MM cell lines. Am80 upregulated expression of the interferon regulatory factor-1 gene, followed by enhancement of CIITA expression. This is the first report demonstrating that retinoid can induce the expression of MHC II in terminally-differentiated plasma/MM cells. PMID:17229644

  4. Single-Molecule Motions of MHC Class II Rely on Bound Peptides

    PubMed Central

    Kozono, Haruo; Matsushita, Yufuku; Ogawa, Naoki; Kozono, Yuko; Miyabe, Toshihiro; Sekiguchi, Hiroshi; Ichiyanagi, Kouhei; Okimoto, Noriaki; Taiji, Makoto; Kanagawa, Osami; Sasaki, Yuji C.

    2015-01-01

    The major histocompatibility complex (MHC) class II protein can bind peptides of different lengths in the region outside the peptide-binding groove. Peptide-flanking residues (PFRs) contribute to the binding affinity of the peptide for MHC and change the immunogenicity of the peptide/MHC complex with regard to T cell receptor (TCR). The mechanisms underlying these phenomena are currently unknown. The molecular flexibility of the peptide/MHC complex may be an important determinant of the structures recognized by certain T cells. We used single-molecule x-ray analysis (diffracted x-ray tracking (DXT)) and fluorescence anisotropy to investigate these mechanisms. DXT enabled us to monitor the real-time Brownian motion of the peptide/MHC complex and revealed that peptides without PFRs undergo larger rotational motions than peptides with PFRs. Fluorescence anisotropy further revealed that peptides without PFRs exhibit slightly larger motions on the nanosecond timescale. These results demonstrate that peptides without PFRs undergo dynamic motions in the groove of MHC and consequently are able to assume diverse structures that can be recognized by T cells. PMID:25606683

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

    PubMed

    Reches, Adi; Nachmani, Daphna; Berhani, Orit; Duev-Cohen, Alexandra; Shreibman, Dorin; Ophir, Yael; Seliger, Barbara; Mandelboim, Ofer

    2016-06-15

    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. PMID:27194785

  6. Spectrum of MHC Class II Variability in Darwin’s Finches and Their Close Relatives

    PubMed Central

    Sato, Akie; Tichy, Herbert; Grant, Peter R.; Grant, B. Rosemary; Sato, Tetsuji; O’hUigin, Colm

    2011-01-01

    The study describes >400 major histocompatibility complex (MHC) class II B exon 2 and 114 intron 2 sequences of 36 passerine bird species, 13 of which belong to the group of Darwin’s finches (DFs) and the remaining 23 to close or more distant relatives of DFs in Central and South America. The data set is analyzed by a combination of judiciously selected statistical methods. The analysis reveals that reliable information concerning MHC organization, including the assignment of sequences to loci, and evolution, as well as the process of species divergence, can be obtained in the absence of genomic sequence data, if the analysis is taken several steps beyond the standard phylogenetic tree construction approach. The main findings of the present study are these: The MHC class II B region of the passerine birds is as elaborate in its organization, divergence, and genetic diversity as the MHC of the eutherian mammals, specifically the primates. Hence, the reported simplicity of the fowl MHC is an oddity. With the help of appropriate markers, the divergence of the MHC genes can be traced deep in the phylogeny of the bird taxa. Transspecies polymorphism is rampant at many of the bird MHC loci. In this respect, the DFs behave as if they were a single, genetically undifferentiated population. There is thus far no indication of alleles that could be considered species, genus, or even DF group specific. The implication of these findings is that DFs are in the midst of adaptive radiations, in which morphological differentiation into species is running ahead of genetic differentiation in genetic systems such as the MHC or the mitochondrial DNA. The radiations are so young that there has not been enough time to sort out polymorphisms at most of the loci among the morphologically differentiating species. These findings parallel those on Lake Victoria haplochromine fishes. Several of the DF MHC allelic lineages can be traced back to the MHC genes of the species Tiaris obscura

  7. Pulse-chase analysis for studies of MHC class II biosynthesis, maturation, and peptide loading

    PubMed Central

    Hou, Tieying; Rinderknecht, Cornelia H; Hadjinicolaou, Andreas V; Busch, Robert; Mellins, Elizabeth

    2014-01-01

    Pulse-chase analysis is a commonly used technique for studying the synthesis, processing and transport of proteins. Cultured cells expressing proteins of interest are allowed to take up radioactively labeled amino acids for a brief interval (“pulse”), during which all newly synthesized proteins incorporate the label. The cells are then returned to non-radioactive culture medium for various times (“chase”), during which proteins may undergo conformational changes, trafficking, or degradation. Proteins of interest are isolated (usually by immunoprecipitation) and resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and the fate of radiolabeled molecules is examined by autoradiography. This chapter describes a pulse-chase protocol suitable for studies of major histocompatibility complex (MHC) class II biosynthesis and maturation. We discuss how results are affected by the recognition by certain anti-class II antibodies of distinct class II conformations associated with particular biosynthetic states. Our protocol can be adapted to follow the fate of many other endogenously synthesized proteins, including viral or transfected gene products, in cultured cells. PMID:23329504

  8. In Situ Peptide-MHC-II Tetramer Staining of Antigen-Specific CD4+ T Cells in Tissues.

    PubMed

    Dileepan, Thamotharampillai; Kim, Hyeon O; Cleary, P Patrick; Skinner, Pamela J

    2015-01-01

    The invention of peptide-MHC-tetramer technology to label antigen-specific T cells has led to an enhanced understanding of T lymphocyte biology. Here we describe the development of an in situ pMHC-II tetramer staining method to visualize antigen-specific CD4+ T cells in tissues. This method complements other methods developed that similarly use MHC class II reagents to stain antigen-specific CD4+ T cells in situ. In this study, we used group A streptococcus (GAS) expressing a surrogate peptide (2W) to inoculate C57BL/6 mice, and used fresh nasal-associated lymphoid tissues (NALT) in optimizing the in situ staining of 2W:I-Ab specific CD4+ T cells. The results showed 2W:I-Ab tetramer-binding CD4+ T cells in GAS-2W but not GAS infected mice. This method holds promise to be broadly applicable to study the localization, abundance, and phenotype of antigen-specific CD4+ T cells in undisrupted tissues. PMID:26067103

  9. Characterization of anti-channel catfish MHC class II monoclonal antibodies

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study characterizes four monoclonal antibodies (mAb) developed against the major histocompatibility complex (MHC) class II beta chain of the channel catfish, Ictalurus punctatus. Immunoprecipitations using catfish clonal B cells revealed that each of these mAbs immunoselected proteins of appro...

  10. High-resolution analysis of the murine MHC class II immunopeptidome.

    PubMed

    Sofron, Adriana; Ritz, Danilo; Neri, Dario; Fugmann, Tim

    2016-02-01

    The reliable identification of peptides bound to major histocompatibility complex (MHC) class II is fundamental for the study of the host immune response against pathogens and the pathogenesis of autoimmune conditions. Here, we describe an improved methodology combining immuno-affinity enrichment of MHC class II complexes, optimized elution conditions and quadrupole Orbitrap mass spectrometry-based characterization of the immunopeptidome. The methodology allowed the identification of over 1000 peptides with 1% false discovery rate from 10(8) murine A20 lymphoma cells. The study revealed the I-A(d) -specific motif in high resolution after multisequence alignment. The methodology was generally applied to the purification of MHC class II from cell lines and murine spleens. We identified 2963 peptides from BALB/c and 2712 from C57BL/6 mouse spleens. The identification of peptides bound to MHC class II in vitro and in vivo will facilitate the characterization of T-cell specificities, as well as the development of biotherapeutics and vaccines. PMID:26495903

  11. Macroautophagy in Endogenous Processing of Self- and Pathogen-Derived Antigens for MHC Class II Presentation

    PubMed Central

    Duraes, Fernanda V.; Niven, Jennifer; Dubrot, Juan; Hugues, Stéphanie; Gannagé, Monique

    2015-01-01

    Although autophagy is a process that has been studied for several years its link with antigen presentation and T cell immunity has only recently emerged. Autophagy, which means “self-eating,” is important to maintain cell homeostasis and refers to a collection of mechanisms that delivers intracellular material for degradation into lysosomes. Among them, macroautophagy pathway has many implications in different biological processes, including innate and adaptive immunity. In particular, macroautophagy can provide a substantial source of intracellular antigens for loading onto MHC class II molecules using the alternative MHC class II pathway. Through autophagosomes, endogenous self-antigens as well as antigens derived from intracellular pathogens can be delivered to MHC class II compartment and presented to CD4+ T cells. The pathway will, therefore, impact both peripheral T cell tolerance and the pathogen specific immune response. This review will describe the contribution of autophagy to intracellular presentation of endogenous self- or pathogen-derived antigens via MHC class II and its consequences on CD4+ T cell responses. PMID:26441964

  12. Intestinal immunization of mice with antigen conjugated to anti-MHC class II antibodies.

    PubMed

    Estrada, A; McDermott, M R; Underdown, B J; Snider, D P

    1995-07-01

    We have explored a new technique for immunization of the intestinal tract of mice, using protein antigens bound to antibodies with specificity for murine MHC class II molecules (MHC-II). Either of two protein antigens, hen avidin (AV) or hen egg lysozyme (HEL) were covalently conjugated to anti-MHC-II antibodies and the purified conjugates were given orally (p.o.) or by direct intraduodenal (i.d.) injection into the intestinal lumen of mice. A secondary immunization p.o. with the same conjugate or with the non-conjugated antigen in the presence of cholera toxin (CTX) resulted in production of both intestinal secretory IgA and serum IgA antibody by those mice. In addition, serum IgG antibodies were produced. Conjugates with appropriate MHC-II specificity targeted the antigen because they induced more IgA and IgG antibody than conjugates with irrelevant antibody specificity or antigen alone, and because they induced antibody in mice that were genetic low responders to antigen. The results indicate the feasibility of oral subunit type vaccines with antibody targeting technology. PMID:7483762

  13. Neutral and selective processes shape MHC gene diversity and expression in stocked brook charr populations (Salvelinus fontinalis).

    PubMed

    Lamaze, Fabien C; Pavey, Scott A; Normandeau, Eric; Roy, Gabriel; Garant, Dany; Bernatchez, Louis

    2014-04-01

    The capacity of an individual to battle infection is an important fitness determinant in wild vertebrate populations. The major histocompatibility complex (MHC) genes are crucial for a host's adaptive immune system to detect pathogens. However, anthropogenic activities may disrupt natural cycles of co-evolution between hosts and pathogens. In this study, we investigated the dynamic sequence and expression variation of host parasite interactions in brook charr (Salvelinus fontinalis) in a context of past human disturbance via population supplementation from domestic individuals. To do so, we developed a new method to examine selection shaping MHC diversity within and between populations and found a complex interplay between neutral and selective processes that varied between lakes that were investigated. We provided evidence for a lower introgression rate of domestic alleles and found that parasite infection increased with domestic genomic background of individuals. We also documented an association between individual MHC alleles and parasite taxa. Finally, longer cis-regulatory minisatellites were positively correlated with MHC II down-regulation and domestic admixture, suggesting that inadvertent selection during domestication resulted in a lower immune response capacity, through a trade-off between growth and immunity, which explained the negative selection of domestic alleles at least under certain circumstances. PMID:24795997

  14. Influence of kinship and MHC class II genotype on visual traits in zebrafish larvae (Danio rerio).

    PubMed

    Hinz, Cornelia; Gebhardt, Katharina; Hartmann, Alexander K; Sigman, Lauren; Gerlach, Gabriele

    2012-01-01

    Kin recognition can drive kin selection and the evolution of social behaviour. In zebrafish (Danio rerio, Hamilton 1822), kin recognition is based on olfactory and visual imprinting processes. If larvae are exposed to visual and chemical cues of kin at day 5 and 6 post fertilization they will recognize kin throughout life, while exposure to non-kin fails to trigger any recognition. Chemical imprinting signals are transcribed by polymorphic genes of the major histocompatibility complex (MHC) code; however, the underlying mechanism for visual imprinting remains unclear. Here we provide evidence for the existence of family-specific differences in morphometry and pigmentation pattern of six day old zebrafish larvae. While rump, tail and body pigmentation were dependent on relatedness, iris pigmentation and morphometry were also influenced by MHC class II genotype. Our study revealed that the MHC not only influences the chemical signature of individuals, but also their visual appearance. PMID:23251449

  15. Alternative donor SCT for the treatment of MHC class II deficiency.

    PubMed

    Small, T N; Qasim, W; Friedrich, W; Chiesa, R; Bleesing, J J; Scurlock, A; Veys, P; Sparber-Sauer, M

    2013-02-01

    MHC Class II deficiency is a rare primary immunodeficiency disease characterized by absent HLA Class II expression resulting in CD4 lymphopenia, lack of Ag-specific responses and recurrent infection. Without successful allogeneic SCT, most children succumb to infection within the first decade of life. To date, alternative donor transplants for this disorder have been inferior to SCT for other forms of combined immunodeficiency disease due to an increased incidence of graft rejection, GVHD and death from infections generally acquired before haematopoietic cell transplantation. This study details the transplant outcome of 16 affected children consecutively transplanted at four centers since 1990, 8 of whom required mechanical ventilation pretransplant. Stem cells were derived from an HLA-mismatched family member (n=10), an HLA-matched unrelated adult donor (n=4), or an unrelated cord blood donor (n=2). Graft failure occurred in five children, all of whom underwent a second SCT. Six patients developed acute GVHD although no patient developed chronic GVHD after primary transplantation. CD4 T-cell reconstitution remained below the normal range for age, suggesting defective thymopoiesis after allo-SCT. Nonetheless, 69% of children survive without GVHD at a median follow-up of 5.7 years, indicating improved outcomes compared with previous studies. PMID:23000650

  16. Nonclassical antigen-processing pathways are required for MHC class II-restricted direct tumor recognition by NY-ESO-1-specific CD4(+) T cells.

    PubMed

    Matsuzaki, Junko; Tsuji, Takemasa; Luescher, Immanuel; Old, Lloyd J; Shrikant, Protul; Gnjatic, Sacha; Odunsi, Kunle

    2014-04-01

    Tumor antigen-specific CD4(+) T cells that directly recognize cancer cells are important for orchestrating antitumor immune responses at the local tumor sites. However, the mechanisms of direct MHC class II (MHC-II) presentation of intracellular tumor antigen by cancer cells are poorly understood. We found that two functionally distinct subsets of CD4(+) T cells were expanded after HLA-DPB1*04 (DP04)-binding NY-ESO-1157-170 peptide vaccination in patients with ovarian cancer. Although both subsets recognized exogenous NY-ESO-1 protein pulsed on DP04(+) target cells, only one type recognized target cells with intracellular expression of NY-ESO-1. The tumor-recognizing CD4(+) T cells more efficiently recognized the short 8-9-mer peptides than the non-tumor-recognizing CD4(+) T cells. In addition to endosomal/lysosomal proteases that are typically involved in MHC-II antigen presentation, several pathways in the MHC class I presentation pathways, such as the proteasomal degradation and transporter-associated with antigen-processing-mediated peptide transport, were also involved in the presentation of intracellular NY-ESO-1 on MHC-II. The presentation was inhibited significantly by primaquine, a small molecule that inhibits endosomal recycling, consistent with findings that pharmacologic inhibition of new protein synthesis enhances antigen presentation. Together, our data demonstrate that cancer cells selectively present peptides from intracellular tumor antigens on MHC-II by multiple nonclassical antigen-processing pathways. Harnessing the direct tumor-recognizing ability of CD4(+) T cells could be a promising strategy to enhance antitumor immune responses in the immunosuppressive tumor microenvironment. PMID:24764581

  17. Non-classical antigen processing pathways are required for MHC class II-restricted direct tumor recognition by NY-ESO-1-specific CD4+ T cells

    PubMed Central

    Matsuzaki, Junko; Tsuji, Takemasa; Luescher, Immanuel; Old, Lloyd J.; Shrikant, Protul; Gnjatic, Sacha; Odunsi, Kunle

    2014-01-01

    Tumor antigen-specific CD4+ T cells that directly recognize cancer cells are important for orchestrating antitumor immune responses at the local tumor sites. However, the mechanisms of direct MHC class II (MHC-II) presentation of intracellular tumor antigen by cancer cells are poorly understood. We found that two functionally distinct subsets of CD4+ T cells were expanded after HLA-DPB1*04 (DP04)-binding NY-ESO-1157–170 peptide vaccination in ovarian cancer patients. While both subsets similarly recognized exogenous NY-ESO-1 protein pulsed on DP04+ target cells, only one type recognized target cells with intracellular expression of NY-ESO-1. The tumor-recognizing CD4+ T cells more efficiently recognized the short 8–9-mer peptides than the non-tumor-recognizing CD4+ T cells. In addition to endosomal/lysosomal proteases that are typically involved in MHC-II antigen presentation, several pathways in the MHC class I presentation pathways such as the proteasomal degradation and transporter-associated with antigen-processing (TAP)-mediated peptide transport were also involved in the presentation of intracellular NY-ESO-1 on MHC-II. The presentation was inhibited significantly by primaquine, a small molecule that inhibits endosomal recycling, consistent with findings that pharmacological inhibition of new protein synthesis enhances antigen presentation. Together, our data demonstrated that cancer cells selectively present peptides from intracellular tumor antigens on MHC-II by multiple non-classical antigen-processing pathways. Harnessing direct tumor-recognizing ability of CD4+ T cells could be a promising strategy to enhance antitumor immune responses in the immunosuppressive tumor microenvironment. PMID:24764581

  18. Structural Basis for the Presentation of Tumor-Associated MHC Class II-Restricted Phosphopeptides to CD4+ T Cells

    SciTech Connect

    Li, Y.; Depontieu, F; Sidney, J; Salay, T; Engelhard, V; Hunt, D; Sette, A; Topalian, S; Mariuzza, R

    2010-01-01

    Dysregulated protein phosphorylation is a hallmark of malignant transformation. Transformation can generate major histocompatibility complex (MHC)-bound phosphopeptides that are differentially displayed on tumor cells for specific recognition by T cells. To understand how phosphorylation alters the antigenic identity of self-peptides and how MHC class II molecules present phosphopeptides for CD4{sup +} T-cell recognition, we determined the crystal structure of a phosphopeptide derived from melanoma antigen recognized by T cells-1 (pMART-1), selectively expressed by human melanomas, in complex with HLA-DR1. The structure revealed that the phosphate moiety attached to the serine residue at position P5 of pMART-1 is available for direct interactions with T-cell receptor (TCR) and that the peptide N-terminus adopts an unusual conformation orienting it toward TCR. This structure, combined with measurements of peptide affinity for HLA-DR1 and of peptide-MHC recognition by pMART-1-specific T cells, suggests that TCR recognition is focused on the N-terminal portion of pMART-1. This recognition mode appears to be distinct from that of foreign antigen complexes but is remarkably reminiscent of the way autoreactive TCRs engage self- or altered self-peptides, consistent with the tolerogenic nature of tumor-host immune interactions.

  19. Contrasting evolutionary histories of MHC class I and class II loci in grouse-effects of selection and gene conversion.

    PubMed

    Minias, P; Bateson, Z W; Whittingham, L A; Johnson, J A; Oyler-McCance, S; Dunn, P O

    2016-05-01

    Genes of the major histocompatibility complex (MHC) encode receptor molecules that are responsible for recognition of intracellular and extracellular pathogens (class I and class II genes, respectively) in vertebrates. Given the different roles of class I and II MHC genes, one might expect the strength of selection to differ between these two classes. Different selective pressures may also promote different rates of gene conversion at each class. Despite these predictions, surprisingly few studies have looked at differences between class I and II genes in terms of both selection and gene conversion. Here, we investigated the molecular evolution of MHC class I and II genes in five closely related species of prairie grouse (Centrocercus and Tympanuchus) that possess one class I and two class II loci. We found striking differences in the strength of balancing selection acting on MHC class I versus class II genes. More than half of the putative antigen-binding sites (ABS) of class II were under positive or episodic diversifying selection, compared with only 10% at class I. We also found that gene conversion had a stronger role in shaping the evolution of MHC class II than class I. Overall, the combination of strong positive (balancing) selection and frequent gene conversion has maintained higher diversity of MHC class II than class I in prairie grouse. This is one of the first studies clearly demonstrating that macroevolutionary mechanisms can act differently on genes involved in the immune response against intracellular and extracellular pathogens. PMID:26860199

  20. Contrasting evolutionary histories of MHC class I and class II loci in grouse—Effects of selection and gene conversion

    USGS Publications Warehouse

    Minias, Piotr; Bateson, Zachary W; Whittingham, Linda A; Johnson, Jeff A.; Oyler-McCance, Sara J.; Dunn, Peter O

    2016-01-01

    Genes of the major histocompatibility complex (MHC) encode receptor molecules that are responsible for recognition of intracellular and extracellular pathogens (class I and class II genes, respectively) in vertebrates. Given the different roles of class I and II MHC genes, one might expect the strength of selection to differ between these two classes. Different selective pressures may also promote different rates of gene conversion at each class. Despite these predictions, surprisingly few studies have looked at differences between class I and II genes in terms of both selection and gene conversion. Here, we investigated the molecular evolution of MHC class I and II genes in five closely related species of prairie grouse (Centrocercus and Tympanuchus) that possess one class I and two class II loci. We found striking differences in the strength of balancing selection acting on MHC class I versus class II genes. More than half of the putative antigen-binding sites (ABS) of class II were under positive or episodic diversifying selection, compared with only 10% at class I. We also found that gene conversion had a stronger role in shaping the evolution of MHC class II than class I. Overall, the combination of strong positive (balancing) selection and frequent gene conversion has maintained higher diversity of MHC class II than class I in prairie grouse. This is one of the first studies clearly demonstrating that macroevolutionary mechanisms can act differently on genes involved in the immune response against intracellular and extracellular pathogens.

  1. Multiple divergent haplotypes express completely distinct sets of class I MHC genes in zebrafish.

    PubMed

    McConnell, Sean C; Restaino, Anthony C; de Jong, Jill L O

    2014-03-01

    The zebrafish is an important animal model for stem cell biology, cancer, and immunology research. Histocompatibility represents a key intersection of these disciplines; however, histocompatibility in zebrafish remains poorly understood. We examined a set of diverse zebrafish class I major histocompatibility complex (MHC) genes that segregate with specific haplotypes at chromosome 19, and for which donor-recipient matching has been shown to improve engraftment after hematopoietic transplantation. Using flanking gene polymorphisms, we identified six distinct chromosome 19 haplotypes. We describe several novel class I U lineage genes and characterize their sequence properties, expression, and haplotype distribution. Altogether, ten full-length zebrafish class I genes were analyzed, mhc1uba through mhc1uka. Expression data and sequence properties indicate that most are candidate classical genes. Several substitutions in putative peptide anchor residues, often shared with deduced MHC molecules from additional teleost species, suggest flexibility in antigen binding. All ten zebrafish class I genes were uniquely assigned among the six haplotypes, with dominant or codominant expression of one to three genes per haplotype. Interestingly, while the divergent MHC haplotypes display variable gene copy number and content, the different genes appear to have ancient origin, with extremely high levels of sequence diversity. Furthermore, haplotype variability extends beyond the MHC genes to include divergent forms of psmb8. The many disparate haplotypes at this locus therefore represent a remarkable form of genomic region configuration polymorphism. Defining the functional MHC genes within these divergent class I haplotypes in zebrafish will provide an important foundation for future studies in immunology and transplantation. PMID:24291825

  2. Expression levels of MHC class I molecules are inversely correlated with promiscuity of peptide binding.

    PubMed

    Chappell, Paul; Meziane, El Kahina; Harrison, Michael; Magiera, Łukasz; Hermann, Clemens; Mears, Laura; Wrobel, Antony G; Durant, Charlotte; Nielsen, Lise Lotte; Buus, Søren; Ternette, Nicola; Mwangi, William; Butter, Colin; Nair, Venugopal; Ahyee, Trudy; Duggleby, Richard; Madrigal, Alejandro; Roversi, Pietro; Lea, Susan M; Kaufman, Jim

    2015-01-01

    Highly polymorphic major histocompatibility complex (MHC) molecules are at the heart of adaptive immune responses, playing crucial roles in many kinds of disease and in vaccination. We report that breadth of peptide presentation and level of cell surface expression of class I molecules are inversely correlated in both chickens and humans. This relationship correlates with protective responses against infectious pathogens including Marek's disease virus leading to lethal tumours in chickens and human immunodeficiency virus infection progressing to AIDS in humans. We propose that differences in peptide binding repertoire define two groups of MHC class I molecules strategically evolved as generalists and specialists for different modes of pathogen resistance. We suggest that differences in cell surface expression level ensure the development of optimal peripheral T cell responses. The inverse relationship of peptide repertoire and expression is evidently a fundamental property of MHC molecules, with ramifications extending beyond immunology and medicine to evolutionary biology and conservation. PMID:25860507

  3. The relative roles of MHC and non-MHC antigens in bone marrow transplantation in rats. Graft acceptance and antigenic expression on donor red blood cells.

    PubMed

    Pinto, M; Gill, T J; Kunz, H W; Dixon-McCarthy, B D

    1983-06-01

    In order to investigate the influence of MHC and non-MHC genes in bone marrow transplantation, various combinations of congenic and inbred strains of rats were used as donors and recipients. A standard regimen of busulfan and cyclophosphamide treatment was used to condition the recipients. The resultant survival patterns of the animals indicated that: (1) a difference across the entire RT1 (MHC) complex is sufficient for the induction of fatal graft-versus-host disease (GVHD) in 100% of the engrafted animals; and (2) the blood group antigens RT2 and RT3, which are controlled by non-MHC genes, do not cause bone marrow graft rejection or GVHD. There were sequential changes of expression in surface alloantigens on the red cells in different donor-recipient combinations without other hematologic changes in the busulfan-cyclophosphamide conditioned bone marrow chimeras. PMID:6346598

  4. Prediction of peptides binding to MHC class I and II alleles by temporal motif mining

    PubMed Central

    2013-01-01

    Background MHC (Major Histocompatibility Complex) is a key player in the immune response of most vertebrates. The computational prediction of whether a given antigenic peptide will bind to a specific MHC allele is important in the development of vaccines for emerging pathogens, the creation of possibilities for controlling immune response, and for the applications of immunotherapy. One of the problems that make this computational prediction difficult is the detection of the binding core region in peptides, coupled with the presence of bulges and loops causing variations in the total sequence length. Most machine learning methods require the sequences to be of the same length to successfully discover the binding motifs, ignoring the length variance in both motif mining and prediction steps. In order to overcome this limitation, we propose the use of time-based motif mining methods that work position-independently. Results The prediction method was tested on a benchmark set of 28 different alleles for MHC class I and 27 different alleles for MHC class II. The obtained results are comparable to the state of the art methods for both MHC classes, surpassing the published results for some alleles. The average prediction AUC values are 0.897 for class I, and 0.858 for class II. Conclusions Temporal motif mining using partial periodic patterns can capture information about the sequences well enough to predict the binding of the peptides and is comparable to state of the art methods in the literature. Unlike neural networks or matrix based predictors, our proposed method does not depend on peptide length and can work with both short and long fragments. This advantage allows better use of the available training data and the prediction of peptides of uncommon lengths. PMID:23368521

  5. Degradation, Promoter Recruitment and Transactivation Mediated by the Extreme N-Terminus of MHC Class II Transactivator CIITA Isoform III

    PubMed Central

    Ethier, Sylvain; Gaudreau, Luc; Steimle, Viktor

    2016-01-01

    Multiple relationships between ubiquitin-proteasome mediated protein turnover and transcriptional activation have been well documented, but the underlying mechanisms are still poorly understood. One way to induce degradation is via ubiquitination of the N-terminal α-amino group of proteins. The major histocompatibility complex (MHC) class II transactivator CIITA is the master regulator of MHC class II gene expression and we found earlier that CIITA is a short-lived protein. Using stable and transient transfections of different CIITA constructs into HEK-293 and HeLa cell lines, we show here that the extreme N-terminal end of CIITA isoform III induces both rapid degradation and transactivation. It is essential that this sequence resides at the N-terminal end of the protein since blocking of the N-terminal end with an epitope-tag stabilizes the protein and reduces transactivation potential. The first ten amino acids of CIITA isoform III act as a portable degron and transactivation sequence when transferred as N-terminal extension to truncated CIITA constructs and are also able to destabilize a heterologous protein. The same is observed with the N-terminal ends of several known N-terminal ubiquitination substrates, such as Id2, Cdt1 and MyoD. Arginine and proline residues within the N-terminal ends contribute to rapid turnover. The N-terminal end of CIITA isoform III is responsible for efficient in vivo recruitment to the HLA-DRA promoter and increased interaction with components of the transcription machinery, such as TBP, p300, p400/Domino, the 19S ATPase S8, and the MHC-II promoter binding complex RFX. These experiments reveal a novel function of free N-terminal ends of proteins in degradation-dependent transcriptional activation. PMID:26871568

  6. Degradation, Promoter Recruitment and Transactivation Mediated by the Extreme N-Terminus of MHC Class II Transactivator CIITA Isoform III.

    PubMed

    Beaulieu, Yves B; Leon Machado, Jorge A; Ethier, Sylvain; Gaudreau, Luc; Steimle, Viktor

    2016-01-01

    Multiple relationships between ubiquitin-proteasome mediated protein turnover and transcriptional activation have been well documented, but the underlying mechanisms are still poorly understood. One way to induce degradation is via ubiquitination of the N-terminal α-amino group of proteins. The major histocompatibility complex (MHC) class II transactivator CIITA is the master regulator of MHC class II gene expression and we found earlier that CIITA is a short-lived protein. Using stable and transient transfections of different CIITA constructs into HEK-293 and HeLa cell lines, we show here that the extreme N-terminal end of CIITA isoform III induces both rapid degradation and transactivation. It is essential that this sequence resides at the N-terminal end of the protein since blocking of the N-terminal end with an epitope-tag stabilizes the protein and reduces transactivation potential. The first ten amino acids of CIITA isoform III act as a portable degron and transactivation sequence when transferred as N-terminal extension to truncated CIITA constructs and are also able to destabilize a heterologous protein. The same is observed with the N-terminal ends of several known N-terminal ubiquitination substrates, such as Id2, Cdt1 and MyoD. Arginine and proline residues within the N-terminal ends contribute to rapid turnover. The N-terminal end of CIITA isoform III is responsible for efficient in vivo recruitment to the HLA-DRA promoter and increased interaction with components of the transcription machinery, such as TBP, p300, p400/Domino, the 19S ATPase S8, and the MHC-II promoter binding complex RFX. These experiments reveal a novel function of free N-terminal ends of proteins in degradation-dependent transcriptional activation. PMID:26871568

  7. Vaccinia Virus A35R Inhibits MHC Class II Antigen Presentation

    PubMed Central

    Rehm, Kristina E.; Connor, Ramsey F.; Jones, Gwendolyn J.B.; Yimbu, Kenneth; Roper, Rachel L.

    2009-01-01

    The Vaccinia virus gene A35R (Copenhagen designation) is highly conserved in mammalian-tropic poxviruses and is an important virulence factor, but its function was unknown. We show herein that A35 does not affect viral infectivity, apoptosis induction, or replication; however, we found that A35 significantly inhibited MHC class II-restricted antigen presentation, immune priming of T lymphocytes, and subsequent chemokine and cytokine synthesis. A35 localized to endosomes and reduced the amount of a model antigenic peptide displayed in the cleft of class II MHC. In addition, A35 decreased VV specific T cell responses in vivo. Thus, this is the first report identifying a function for the A35 protein in virulence as well as the first report identifying a VV gene that inhibits peptide antigen presentation. PMID:19954808

  8. MHC class II B diversity in blue tits: a preliminary study

    PubMed Central

    Aguilar, Juan Rivero-de; Schut, Elske; Merino, Santiago; Martínez, Javier; Komdeur, Jan; Westerdahl, Helena

    2013-01-01

    In this study, we partly characterize major histocompatibility complex (MHC) class II B in the blue tit (Cyanistes caeruleus). A total of 22 individuals from three different European locations: Spain, The Netherlands, and Sweden were screened for MHC allelic diversity. The MHC genes were investigated using both PCR-based methods and unamplified genomic DNA with restriction fragment length polymorphism (RFLP) and southern blots. A total of 13 different exon 2 sequences were obtained independently from DNA and/or RNA, thus confirming gene transcription and likely functionality of the genes. Nine out of 13 alleles were found in more than one country, and two alleles appeared in all countries. Positive selection was detected in the region coding for the peptide binding region (PBR). A maximum of three alleles per individual was detected by sequencing and the RFLP pattern consisted of 4–7 fragments, indicating a minimum number of 2–4 loci per individual. A phylogenetic analysis, demonstrated that the blue tit sequences are divergent compared to sequences from other passerines resembling a different MHC lineage than those possessed by most passerines studied to date. PMID:23919136

  9. DPA1*02012: A DPA1*0201-related Mhc class II allele in West Africa

    SciTech Connect

    Meyer, C.G.; May, J.; Spauke, D.; Schnittger, L.

    1994-12-31

    DNA techniques such as sequence-specific oligonucleotide probe (SSOP) hybridizations, restriction-fragment length polymorphism (RFLP) analyses, and DNA sequencing have greatly supported the characterization of Mhc class II allelic polymorphism. Here the authors describe a DPA 1 allele which has been identified in two male individuals from Liberia and Benin, West Africa, during a survey study on Mhc class II associations with the different manifestations after infection with Onchocerca volvulus. 4 refs., 1 fig.

  10. Force-Regulated In Situ TCR-Peptide-Bound MHC Class II Kinetics Determine Functions of CD4+ T Cells.

    PubMed

    Hong, Jinsung; Persaud, Stephen P; Horvath, Stephen; Allen, Paul M; Evavold, Brian D; Zhu, Cheng

    2015-10-15

    We have recently shown that two-dimensional (2D) and force-regulated kinetics of TCR-peptide-bound MHC class I (pMHC-I) interactions predict responses of CD8(+) T cells. To test whether these findings are applicable to CD4(+) T cells, we analyzed the in situ 3.L2 TCR-pMHC-II interactions for a well-characterized panel of altered peptide ligands on the T cell surface using the adhesion frequency assay with a micropipette and the thermal fluctuation and force-clamp assays with a biomembrane force probe. We found that the 2D effective TCR-pMHC-II affinity and off-rate correlate with, but better predict the T cell response than, the corresponding measurements with the surface plasmon resonance in three dimensions. The 2D affinity of the CD4 for MHC-II was very low, approaching the detection limit, making it one to two orders of magnitude lower than the affinity of CD8 for MHC-I. In addition, the signal-dependent cooperation between TCR and coreceptor for pMHC binding previously observed for CD8 was not observed for CD4. Interestingly, force elicited TCR-pMHC-II catch-slip bonds for agonists but slip-only bonds for antagonists, thereby amplifying the power of discrimination between altered peptide ligands. These results show that the force-regulated 2D binding kinetics of the 3.L2 TCR for pMHC-II determine functions of CD4(+) T cells. PMID:26336148

  11. DNA Vaccine that Targets Hemagglutinin to MHC Class II Molecules Rapidly Induces Antibody-Mediated Protection against Influenza

    PubMed Central

    Mjaaland, Siri; Roux, Kenneth H.; Fredriksen, Agnete Brunsvik

    2013-01-01

    New influenza A viruses with pandemic potential periodically emerge due to viral genomic reassortment. In the face of pandemic threats, production of conventional egg-based vaccines is time consuming and of limited capacity. We have developed in this study a novel DNA vaccine in which viral hemagglutinin (HA) is bivalently targeted to MHC class II (MHC II) molecules on APCs. Following DNA vaccination, transfected cells secreted vaccine proteins that bound MHC II on APCs and initiated adaptive immune responses. A single DNA immunization induced within 8 d protective levels of strain-specific Abs and also cross-reactive T cells. During the Mexican flu pandemic, a targeted DNA vaccine (HA from A/California/07/2009) was generated within 3 wk after the HA sequences were published online. These results suggest that MHC II–targeted DNA vaccines could play a role in situations of pandemic threats. The vaccine principle should be extendable to other infectious diseases. PMID:23956431

  12. MHC Class II Auto-Antigen Presentation is Unconventional

    PubMed Central

    Sadegh-Nasseri, Scheherazade; Kim, AeRyon

    2015-01-01

    Antigen presentation is highly critical in adoptive immunity. Only by interacting with antigens presented by major histocompatibility complex class II molecules, helper T cells can be stimulated to fight infections or diseases. The degradation of a full protein into small peptide fragments bound to class II molecules is a dynamic, lengthy process consisting of many steps and chaperons. Deregulation in any step of antigen processing could lead to the development of self-reactive T cells or defective immune response to pathogens. Indeed, human leukocyte antigens class II genes are the predominant contributors to susceptibility to autoimmune diseases. Conventional antigen-processing calls for internalization of extracellular antigens followed by processing and epitope selection within antigen-processing subcellular compartments, enriched with all necessary accessory molecules, processing enzymes, and proper pH and denaturing conditions. However, recent data examining the temporal relationship between antigen uptakes, processing, and epitope selection revealed unexpected characteristics for auto-antigenic epitopes, which were not shared with antigenic epitopes from pathogens. This review provides a discussion of the relevance of these findings to the mechanisms of autoimmunity. PMID:26257739

  13. Dendritic cell preactivation impairs MHC class II presentation of vaccines and endogenous viral antigens

    PubMed Central

    Young, Louise J.; Wilson, Nicholas S.; Schnorrer, Petra; Mount, Adele; Lundie, Rachel J.; La Gruta, Nicole L.; Crabb, Brendan S.; Belz, Gabrielle T.; Heath, William R.; Villadangos, Jose A.

    2007-01-01

    When dendritic cells (DCs) encounter signals associated with infection or inflammation, they become activated and undergo maturation. Mature DCs are very efficient at presenting antigens captured in association with their activating signal but fail to present subsequently encountered antigens, at least in vitro. Such impairment of MHC class II (MHC II) antigen presentation has generally been thought to be a consequence of down-regulation of endocytosis, so it might be expected that antigens synthesized by the DCs themselves (for instance, viral antigens) would still be presented by mature DCs. Here, we show that DCs matured in vivo could still capture and process soluble antigens, but were unable to present peptides derived from these antigens. Furthermore, presentation of viral antigens synthesized by the DCs themselves was also severely impaired. Indeed, i.v. injection of pathogen mimics, which caused systemic DC activation in vivo, impaired the induction of CD4 T cell responses against subsequently encountered protein antigens. This immunosuppressed state could be reversed by adoptive transfer of DCs loaded exogenously with antigens, demonstrating that impairment of CD4 T cell responses was due to lack of antigen presentation rather than to overt suppression of T cell activation. The biochemical mechanism underlying this phenomenon was the down-regulation of MHC II–peptide complex formation that accompanied DC maturation. These observations have important implications for the design of prophylactic and therapeutic DC vaccines and contribute to the understanding of the mechanisms causing immunosuppression during systemic blood infections. PMID:17978177

  14. CD4+ T cells from MHC II-dependent thymocyte–thymocyte interaction provide efficient help for B cells

    PubMed Central

    Kim, Eun Ji; Choi, Bomi; Moon, Hana; Lee, You Jeong; Jeon, Yoon Kyeong; Park, Seong Hoe; Kim, Tae Jin; Jung, Kyeong Cheon

    2011-01-01

    Recently, a novel CD4+ T-cell developmental pathway was reported that generates thymocyte–thymocyte (T–T) CD4+ T cells. We established a mouse system (CIITAtgCIITApIV−/−) where thymic positive selection occurred only by major histocompatibility complex (MHC) class II+ thymocytes. T–T CD4+ T cells selected via MHC class II-dependent T–T interaction are comprised of PLZF-negative and innate PLZF-positive populations. Until recently, the functional role of the PLZF-negative population was unclear. In this study, we demonstrate that naïve T–T CD4+ T cells provide B-cell help to a level comparable with that of naïve conventional CD4+ T cells. Considering the absence of PLZF expression in naïve T–T CD4+ T cells, these results suggest that PLZF-negative naïve T–T CD4+ T cells are functionally equivalent to conventional naïve CD4+ T cells in terms of B-cell help. PMID:21358747

  15. Selection, diversity and evolutionary patterns of the MHC class II DAB in free-ranging Neotropical marsupials

    PubMed Central

    Meyer-Lucht, Yvonne; Otten, Celine; Püttker, Thomas; Sommer, Simone

    2008-01-01

    Background Research on the genetic architecture and diversity of the MHC has focused mainly on eutherian mammals, birds and fish. So far, studies on model marsupials used in laboratory investigations indicated very little or even no variation in MHC class II genes. However, natural levels of diversity and selection are unknown in marsupials as studies on wild populations are virtually absent. We used two endemic South American mouse opossums, Gracilinanus microtarsus and Marmosops incanus, to investigate characteristic features of MHC selection. This study is the first investigation of MHC selection in free-ranging Neotropical marsupials. In addition, the evolutionary history of MHC lineages within the group of marsupials was examined. Results G. microtarsus showed extensive levels of MHC diversity within and among individuals as 47 MHC-DAB alleles and high levels of sequence divergence were detected at a minimum of four loci. Positively selected codon sites were identified, of which most were congruent with human antigen binding sites. The diversity in M. incanus was rather low with only eight observed alleles at presumably two loci. However, these alleles also revealed high sequence divergence. Again, positive selection was identified on specific codon sites, all congruent with human ABS and with positively selected sites observed in G. microtarsus. In a phylogenetic comparison alleles of M. incanus interspersed widely within alleles of G. microtarsus with four alleles being present in both species. Conclusion Our investigations revealed extensive MHC class II polymorphism in a natural marsupial population, contrary to previous assumptions. Furthermore, our study confirms for the first time in marsupials the presence of three characteristic features common at MHC loci of eutherian mammals, birds and fish: large allelic sequence divergence, positive selection on specific sites and trans-specific polymorphism. PMID:18534008

  16. Design of Peptide Immunotherapies for MHC Class-II-Associated Autoimmune Disorders

    PubMed Central

    2013-01-01

    Autoimmune disorders, that occur when autoreactive immune cells are induced to activate their responses against self-tissues, affect one percent of the world population and represent one of the top 10 leading causes of death. The major histocompatibility complex (MHC) is a principal susceptibility locus for many human autoimmune diseases, in which self-tissue antigens providing targets for pathogenic lymphocytes are bound to HLA molecules encoded by disease-associated alleles. In spite of the attempts to design strategies for inhibition of antigen presentation targeting the MHC-peptide/TCR complex via generation of blocking antibodies, altered peptide ligands (APL), or inhibitors of costimulatory molecules, potent therapies with minimal side effects have yet to be developed. Copaxone (glatiramer acetate, GA) is a random synthetic amino acid copolymer that reduces the relapse rate by about 30% in relapsing-remitting multiple sclerosis (MS) patients. Based on the elucidated binding motifs of Copaxone and of the anchor residues of the immunogenic myelin basic protein (MBP) peptide to HLA-DR molecules, novel copolymers have been designed and proved to be more effective in suppressing MS-like disease in mice. In this report, we describe the rationale for design of second-generation synthetic random copolymers as candidate drugs for a number of MHC class-II-associated autoimmune disorders. PMID:24324511

  17. Intracellular transport of MHC class II and associated invariant chain in antigen presenting cells from AP-3-deficient mocha mice.

    PubMed

    Sevilla, L M; Richter, S S; Miller, J

    2001-06-15

    MHC class II-restricted antigen presentation requires trafficking of newly synthesized class II-invariant chain complexes from the trans-Golgi network to endosomal, peptide-loading compartments. This transport is mediated by dileucine-like motifs within the cytosolic tail of the invariant chain. Although these signals have been well characterized, the cytosolic proteins that interact with these dileucine signals and mediate Golgi sorting and endosomal transport have not been identified. Recently, an adaptor complex, AP-3, has been identified that interacts with dileucine motifs and mediates endosomal/lysosomal transport in yeast, Drosophila, and mammals. In this report, we have assessed class II-invariant chain trafficking in a strain of mice (mocha) which lacks expression of AP-3. Our studies demonstrate that the lack of AP-3 does not affect the kinetics of invariant chain degradation, the route of class II-invariant chain transport, or the rate and extent of class II-peptide binding as assessed by the generation of SDS-stable dimers. The possible role of other known or unknown adaptor complexes in class II-invariant chain transport is discussed. PMID:11520080

  18. The diabetogenic mouse MHC class II molecule I-A[subscript g7] is endowed with a switch that modulates TCR affinity

    SciTech Connect

    Yoshida, Kenji; Corper, Adam L.; Herro, Rana; Jabri, Bana; Wilson, Ian A.; Teyton, Luc

    2011-11-16

    Genetic susceptibility to autoimmunity is frequently associated with specific MHC alleles. Diabetogenic MHC class II molecules, such as human HLA-DQ8 and mouse I-A{sub g7}, typically have a small, uncharged amino acid residue at position 57 of their {beta} chain ({beta}57); this results in the absence of a salt bridge between {beta}57 and Arg{alpha}76, which is adjacent to the P9 pocket of the peptide-binding groove. However, the influence of Arg{alpha}76 on the selection of the TCR repertoire remains unknown, particularly when the MHC molecule binds a peptide with a neutral amino acid residue at position P9. Here, we have shown that diabetogenic MHC class II molecules bound to a peptide with a neutral P9 residue primarily selected and expanded cells expressing TCRs bearing a negatively charged residue in the first segment of their complementarity determining region 3{beta}. The crystal structure of one such TCR in complex with I-A{sub g7} bound to a peptide containing a neutral P9 residue revealed that a network of favorable long-range (greater than 4 {angstrom}) electrostatic interactions existed among Arg{alpha}76, the neutral P9 residue, and TCR, which supported the substantially increased TCR/peptide-MHC affinity. This network could be modulated or switched to a lower affinity interaction by the introduction of a negative charge at position P9 of the peptide. Our results support the existence of a switch at residue {beta}57 of the I-Ag7 and HLA-DQ8 class II molecules and potentially link normal thymic TCR selection with abnormal peripheral behavior.

  19. The diabetogenic mouse MHC class II molecule I-A[superscript g7] is endowed with a switch that modulates TCR affinity

    SciTech Connect

    Yoshida, Kenji; Corper, Adam L.; Herro, Rana; Jabri, Bana; Wilson, Ian A.; Teyton, Luc

    2010-07-22

    Genetic susceptibility to autoimmunity is frequently associated with specific MHC alleles. Diabetogenic MHC class II molecules, such as human HLA-DQ8 and mouse I-A{sup g7}, typically have a small, uncharged amino acid residue at position 57 of their {beta} chain ({beta}57); this results in the absence of a salt bridge between {beta}57 and Arg{alpha}76, which is adjacent to the P9 pocket of the peptide-binding groove. However, the influence of Arg{alpha}76 on the selection of the TCR repertoire remains unknown, particularly when the MHC molecule binds a peptide with a neutral amino acid residue at position P9. Here, we have shown that diabetogenic MHC class II molecules bound to a peptide with a neutral P9 residue primarily selected and expanded cells expressing TCRs bearing a negatively charged residue in the first segment of their complementarity determining region 3{beta}. The crystal structure of one such TCR in complex with I-Ag7 bound to a peptide containing a neutral P9 residue revealed that a network of favorable long-range (greater than 4 {angstrom}) electrostatic interactions existed among Arg{alpha}76, the neutral P9 residue, and TCR, which supported the substantially increased TCR/peptide-MHC affinity. This network could be modulated or switched to a lower affinity interaction by the introduction of a negative charge at position P9 of the peptide. Our results support the existence of a switch at residue {beta}57 of the I-A{sup g7} and HLA-DQ8 class II molecules and potentially link normal thymic TCR selection with abnormal peripheral behavior.

  20. Use of MHC II structural features in the design of vaccines for organ-specific autoimmune diseases.

    PubMed

    Moustakas, Antonis K; Papadopoulos, George K

    2009-01-01

    The Major Histocompatibility Complex Class II locus is the primary genetic linkage to autoimmune diseases. Susceptibility to each such disease is linked to different alleles, with a few alleles showing also dominant protection. The design of vaccines for autoimmune diseases is a long sought-after goal. As knowledge about the pathogenesis of these diseases has increased, the tools for such an approach have of necessity been refined. We review below the structural essence of MHC II-linked autoimmune diseases which centers on the binding of antigenic peptides to the disease-linked MHC II proteins, and the consequent activation of cognate TCRs from pathogenic CD4+ T cells. The state of affairs in two organ-specific autoimmune diseases, type 1 diabetes, celiac disease are covered, including attempts to treat these via antigen-specific MHC II-guided measures. We offer a couple of testable suggestions as to how this approach could be improved. PMID:19860675

  1. Antigen-specific CD4(+) T cells regulate function of myeloid-derived suppressor cells in cancer via retrograde MHC class II signaling.

    PubMed

    Nagaraj, Srinivas; Nelson, Allison; Youn, Je-in; Cheng, Pingyan; Quiceno, David; Gabrilovich, Dmitry I

    2012-02-15

    Myeloid-derived suppressor cells (MDSC) play a major role in cancer-related immune suppression, yet the nature of this suppression remains controversial. In this study, we evaluated the ability of MDSCs to elicit CD4(+) T-cell tolerance in different mouse tumor models. In contrast to CD8(+) T-cell tolerance, which could be induced by MDSCs in all the tumor models tested, CD4(+) T-cell tolerance could be elicited in only one of the models (MC38) in which a substantial level of MHC class II was expressed on MDSCs compared with control myeloid cells. Mechanistic investigations revealed that MDSCs deficient in MHC class II could induce tolerance to CD8(+) T cells but not to CD4(+) T cells. Unexpectedly, antigen-specific CD4(+) T cells (but not CD8(+) T cells) could dramatically enhance the immune suppressive activity of MDSCs by converting them into powerful nonspecific suppressor cells. This striking effect was mediated by direct cell-cell contact through cross-linking of MHC class II on MDSCs. We also implicated an Ets-1 transcription factor-regulated increase in expression of Cox-2 and prostaglandin E2 in MDSCs in mediating this effect. Together, our findings suggest that activated CD4(+) T cells that are antigen specific may enhance the immune suppressive activity of MDSCs, a mechanism that might serve normally as a negative feedback loop to control immune responses that becomes dysregulated in cancer. PMID:22237629

  2. Low MHC class II diversity in the Tasmanian devil (Sarcophilus harrisii).

    PubMed

    Cheng, Yuanyuan; Sanderson, Claire; Jones, Menna; Belov, Katherine

    2012-07-01

    The largest remaining carnivorous marsupial, the Tasmanian devil (Sarcophilus harrisii), is currently under threat of extinction due to a fatal contagious cancer-devil facial tumour disease. Low major histocompatibility complex (MHC) class I diversity is believed to have contributed to the transmission of the tumour allograft through devil populations. Here, we report low MHC class II variability in this species, with DA β chain genes (Saha-DAB1, 2 and 3) exhibiting very limited diversity and the sole α chain gene (Saha-DAA) monomorphic. Three, six and three alleles were found at Saha-DAB1, 2 and 3, respectively, with a predominant allele found at each locus. Heterozygosity at these three loci is low in the eastern population and modestly higher in northwestern individuals. The results are indicative of a selective sweep likely due to an infectious disease resulting in the fixation of selectively favoured alleles and depletion of genetic diversity at devil class II loci. Several attempts were made to isolate the other marsupial classical class II gene family, namely, DB, resulting in only one DBB pseudogene being found. These findings further support the view that this species has a compromised capacity to respond to pathogen evolution, emerging infectious diseases and environmental changes. PMID:22460528

  3. Is alopecia areata an autoimmune-response against melanogenesis-related proteins, exposed by abnormal MHC class I expression in the anagen hair bulb?

    PubMed Central

    Paus, R.; Slominski, A.; Czarnetzki, B. M.

    1993-01-01

    The etiology of alopecia areata (AA), a putative autoimmune disease characterized by sudden hair loss, has remained obscure. It is not understood, how the characteristic inflammatory infiltrate that selectively attacks anagen hair follicles in AA is generated. We hypothesize that this reflects an unexplored form of autoimmunity, a cytotoxic T cell attack on rhythmically synthesized autoantigens normally sequestered by a lack or very low level of MHC class I (MHC I)-expression, and suggest the following mechanism of AA pathogenesis: Microtrauma, neurogenic inflammation, or microbial antigens cause a localized breakdown of MHC I-"negativity" in the proximal anagen hair bulb via proinflammatory cytokines. This exposes autoantigens derived from melanogenesis-related proteins (MRP-DP), which are only generated during anagen, and triggers two successive waves of autoimmune responses: CD8+ cytotoxic T cells initiate AA after recognizing MRP-DP abnormally presented by MHC I molecules on hair matrix melanocytes and/or keratinocytes; a secondary attack, carried by CD4+ T cells and antigen presenting cells, is then mounted against MHC class II--presented additional autoantigens exposed by damaged melanocytes and keratinocytes. The latter causes most of the follicular damage, and extrafollicular disease, and depends greatly on the immunogenetic background of affected individuals. This unifying hypothesis explains the clinical heterogeneity and all salient features of AA, and argues that only the unlikely coincidence of multiple predisposing events triggers AA. The suppression of MHC I--expression and synthesis of MRP in the hair bulb, and the "tolerization" of MRP-DP autoreactive CD8+ T cells may be promising strategies for treating AA. PMID:7716973

  4. Molecular cloning, expression pattern, and 3D structural analysis of the MHC class IIB gene in the Chinese longsnout catfish (Leiocassis longirostris).

    PubMed

    Shen, Tong; Xu, Shixia; Yang, Mei; Pang, Shuying; Yang, Guang

    2011-05-15

    Major histocompatibility complex (MHC) class I and class II molecules encode glycoproteins which mediate the specificity of the vertebrate adaptive immune response. In this study, MHC class IIB gene from the Chinese longsnout catfish (Leiocassis longirostris) was cloned and sequenced, which encoded a predicted protein of 248 amino acids (28.06 kDa) containing a signal peptide, a beta 1 domain, a beta 2 domain, a connecting peptide, a transmembrane region, and a cytoplasmic tail. Using PCR with primers designed from known fish MHC class IIB sequences followed by elongation of the 5' and 3' ends using rapid amplification of cDNA ends (RACE), the full-length cDNA of longsnout catfish MHC class IIB was identified to be 1293 bp, consisting of a 26 bp 5'-terminal untranslated region (UTR), a 520 bp 3'-UTR, and a 747 bp open reading frame (ORF) bearing characteristics of the immunoglobulin C-type 1 (IGc1) family. The deduced amino acid sequences of the Chinese longsnout catfish MHC class IIB gene had 58-75% identity with those of other fishes. Six class IIB alleles were identified from five individuals. At most two different alleles observed in each individual may infer the existence of a single locus of class IIB gene in the Chinese longsnout catfish genome. An extensive study of polymorphism was examined in 60 individuals. A total of 11 haplotypes of exon 2 were detected in the sampled Chinese longsnout catfish. The rates of nonsynonymous substitutions (d(N)) occurred at a higher frequency than that of synonymous substitutions (d(S)), suggesting the polymorphism of exon 2 seemed to be maintained by the balancing selection. By using long PCR technique, the genomic sequence was further identified to be 2345 bp in length, which contained six exons and five introns. Interestingly, a 98 bp intron 5 cut the 3'-UTR into two parts. Real-time quantitative RT-PCR demonstrated high expression of MHC IIB in gills, spleen, head kidney, and intestine, moderate expression in liver and

  5. Drift Rather than Selection Dominates MHC Class II Allelic Diversity Patterns at the Biogeographical Range Scale in Natterjack Toads Bufo calamita

    PubMed Central

    Zeisset, Inga; Beebee, Trevor J. C.

    2014-01-01

    Study of major histocompatibility complex (MHC) loci has gained great popularity in recent years, partly due to their function in protecting vertebrates from infections. This is of particular interest in amphibians on account of major threats many species face from emergent diseases such as chytridiomycosis. In this study we compare levels of diversity in an expressed MHC class II locus with neutral genetic diversity at microsatellite loci in natterjack toad (Bufo (Epidalea) calamita) populations across the whole of the species’ biogeographical range. Variation at both classes of loci was high in the glacial refugium areas (REF) and much lower in postglacial expansion areas (PGE), especially in range edge populations. Although there was clear evidence that the MHC locus was influenced by positive selection in the past, congruence with the neutral markers suggested that historical demographic events were the main force shaping MHC variation in the PGE area. Both neutral and adaptive genetic variation declined with distance from glacial refugia. Nevertheless, there were also some indications from differential isolation by distance and allele abundance patterns that weak effects of selection have been superimposed on the main drift effect in the PGE zone. PMID:24937211

  6. Giving CD4+ T cells the slip: viral interference with MHC class II-restricted antigen processing and presentation.

    PubMed

    Forsyth, Katherine S; Eisenlohr, Laurence C

    2016-06-01

    Activation of CD4+ T cells through interactions with peptides bound to Major Histocompatibility Complex Class II (MHC-II) molecules is a crucial step in clearance of most pathogens. Consequently, many viruses have evolved ways of blocking this aspect of adaptive immunity, from specific targeting of processing and presentation components to modulation of signaling pathways that regulate peptide presentation in addition to many other host defense mechanisms. Such cases of interference are far less common compared to what has been elucidated in MHC-I processing and presentation. This may be attributable in part to the complexity of MHC-II antigen processing, the scope of which is only now coming to light. PMID:27115617

  7. Heparan Sulfate Proteoglycans Promote Telomerase Internalization and MHC Class II Presentation on Dendritic Cells.

    PubMed

    Galaine, Jeanne; Kellermann, Guillaume; Guillaume, Yves; Boidot, Romain; Picard, Emilie; Loyon, Romain; Queiroz, Lise; Boullerot, Laura; Beziaud, Laurent; Jary, Marine; Mansi, Laura; André, Claire; Lethier, Lydie; Ségal-Bendirdjian, Evelyne; Borg, Christophe; Godet, Yann; Adotévi, Olivier

    2016-09-01

    Telomerase is a prototype-shared tumor Ag and represents an attractive target for anticancer immunotherapy. We have previously described promiscuous and immunogenic HLA-DR-restricted peptides derived from human telomerase reverse transcriptase (hTERT) and referred as universal cancer peptide (UCP). In nonsmall cell lung cancer, the presence of spontaneous UCP-specific CD4 T cell responses increases the survival of chemotherapy-responding patients. However, the precise mechanisms of hTERT's uptake, processing, and presentation on MHC-II molecules to stimulate CD4 T cells are poorly understood. In this work, by using well-characterized UCP-specific CD4 T cell clones, we showed that hTERT processing and presentation on MHC-II involve both classical endolysosomal and nonclassical cytosolic pathways. Furthermore, to our knowledge, we demonstrated for the first time that hTERT's internalization by dendritic cells requires its interaction with surface heparan sulfate proteoglycans. Altogether, our findings provide a novel mechanism of tumor-specific CD4 T cell activation and will be useful for the development of novel cancer immunotherapies that harness CD4 T cells. PMID:27481844

  8. High levels of MHC class II allelic diversity in lake trout from Lake Superior

    USGS Publications Warehouse

    Dorschner, M.O.; Duris, T.; Bronte, C.R.; Burnham-Curtis, M. K.; Phillips, R.B.

    2000-01-01

    Sequence variation in a 216 bp portion of the major histocompatibility complex (MHC) II B1 domain was examined in 74 individual lake trout (Salvelinus namaycush) from different locations in Lake Superior. Forty-three alleles were obtained which encoded 71-72 amino acids of the mature protein. These sequences were compared with previous data obtained from five Pacific salmon species and Atlantic salmon using the same primers. Although all of the lake trout alleles clustered together in the neighbor-joining analysis of amino acid sequences, one amino acid allelic lineage was shared with Atlantic salmon (Salmo salar), a species in another genus which probably diverged from Salvelinus more than 10-20 million years ago. As shown previously in other salmonids, the level of nonsynonymous nucleotide substitution (d(N)) exceeded the level of synonymous substitution (d(S)). The level of nucleotide diversity at the MHC class II B1 locus was considerably higher in lake trout than in the Pacific salmon (genus Oncorhynchus). These results are consistent with the hypothesis that lake trout colonized Lake Superior from more than one refuge following the Wisconsin glaciation. Recent population bottlenecks may have reduced nucleotide diversity in Pacific salmon populations.

  9. Expression levels of MHC class I molecules are inversely correlated with promiscuity of peptide binding

    PubMed Central

    Chappell, Paul E; Meziane, El Kahina; Harrison, Michael; Magiera, Łukasz; Hermann, Clemens; Mears, Laura; Wrobel, Antoni G; Durant, Charlotte; Nielsen, Lise Lotte; Buus, Søren; Ternette, Nicola; Mwangi, William; Butter, Colin; Nair, Venugopal; Ahyee, Trudy; Duggleby, Richard; Madrigal, Alejandro; Roversi, Pietro; Lea, Susan M; Kaufman, Jim

    2015-01-01

    Highly polymorphic major histocompatibility complex (MHC) molecules are at the heart of adaptive immune responses, playing crucial roles in many kinds of disease and in vaccination. We report that breadth of peptide presentation and level of cell surface expression of class I molecules are inversely correlated in both chickens and humans. This relationship correlates with protective responses against infectious pathogens including Marek's disease virus leading to lethal tumours in chickens and human immunodeficiency virus infection progressing to AIDS in humans. We propose that differences in peptide binding repertoire define two groups of MHC class I molecules strategically evolved as generalists and specialists for different modes of pathogen resistance. We suggest that differences in cell surface expression level ensure the development of optimal peripheral T cell responses. The inverse relationship of peptide repertoire and expression is evidently a fundamental property of MHC molecules, with ramifications extending beyond immunology and medicine to evolutionary biology and conservation. DOI: http://dx.doi.org/10.7554/eLife.05345.001 PMID:25860507

  10. MHC I Expression Regulates Co-clustering and Mobility of Interleukin-2 and -15 Receptors in T Cells.

    PubMed

    Mocsár, Gábor; Volkó, Julianna; Rönnlund, Daniel; Widengren, Jerker; Nagy, Péter; Szöllősi, János; Tóth, Katalin; Goldman, Carolyn K; Damjanovich, Sándor; Waldmann, Thomas A; Bodnár, Andrea; Vámosi, György

    2016-07-12

    MHC glycoproteins form supramolecular clusters with interleukin-2 and -15 receptors in lipid rafts of T cells. The role of highly expressed MHC I in maintaining these clusters is unknown. We knocked down MHC I in FT7.10 human T cells, and studied protein clustering at two hierarchic levels: molecular aggregations and mobility by Förster resonance energy transfer and fluorescence correlation spectroscopy; and segregation into larger domains or superclusters by superresolution stimulated emission depletion microscopy. Fluorescence correlation spectroscopy-based molecular brightness analysis revealed that the studied molecules diffused as tight aggregates of several proteins of a kind. Knockdown reduced the number of MHC I containing molecular aggregates and their average MHC I content, and decreased the heteroassociation of MHC I with IL-2Rα/IL-15Rα. The mobility of not only MHC I but also that of IL-2Rα/IL-15Rα increased, corroborating the general size decrease of tight aggregates. A multifaceted analysis of stimulated emission depletion images revealed that the diameter of MHC I superclusters diminished from 400-600 to 200-300 nm, whereas those of IL-2Rα/IL-15Rα hardly changed. MHC I and IL-2Rα/IL-15Rα colocalized with GM1 ganglioside-rich lipid rafts, but MHC I clusters retracted to smaller subsets of GM1- and IL-2Rα/IL-15Rα-rich areas upon knockdown. Our results prove that changes in expression level may significantly alter the organization and mobility of interacting membrane proteins. PMID:27410738

  11. Enhanced Detection of Antigen-Specific CD4+ T Cells Using Altered Peptide Flanking Residue Peptide–MHC Class II Multimers

    PubMed Central

    Holland, Christopher J.; Dolton, Garry; Scurr, Martin; Ladell, Kristin; Schauenburg, Andrea J.; Miners, Kelly; Madura, Florian; Sewell, Andrew K.; Price, David A.

    2015-01-01

    Fluorochrome-conjugated peptide–MHC (pMHC) class I multimers are staple components of the immunologist’s toolbox, enabling reliable quantification and analysis of Ag-specific CD8+ T cells irrespective of functional outputs. In contrast, widespread use of the equivalent pMHC class II (pMHC-II) reagents has been hindered by intrinsically weaker TCR affinities for pMHC-II, a lack of cooperative binding between the TCR and CD4 coreceptor, and a low frequency of Ag-specific CD4+ T cell populations in the peripheral blood. In this study, we show that peptide flanking regions, extending beyond the central nonamer core of MHC-II–bound peptides, can enhance TCR–pMHC-II binding and T cell activation without loss of specificity. Consistent with these findings, pMHC-II multimers incorporating peptide flanking residue modifications proved superior for the ex vivo detection, characterization, and manipulation of Ag-specific CD4+ T cells, highlighting an unappreciated feature of TCR–pMHC-II interactions. PMID:26553072

  12. The Forgotten: Identification and Functional Characterization of MHC Class II Molecules H2-Eb2 and RT1-Db2.

    PubMed

    Monzón-Casanova, Elisa; Rudolf, Ronald; Starick, Lisa; Müller, Ingrid; Söllner, Christian; Müller, Nora; Westphal, Nico; Miyoshi-Akiyama, Tohru; Uchiyama, Takehiko; Berberich, Ingolf; Walter, Lutz; Herrmann, Thomas

    2016-02-01

    In this article, we report the complete coding sequence and to our knowledge, the first functional analysis of two homologous nonclassical MHC class II genes: RT1-Db2 of rat and H2-Eb2 of mouse. They differ in important aspects compared with the classical class II β1 molecules: their mRNA expression by APCs is much lower, they show minimal polymorphism in the Ag-binding domain, and they lack N-glycosylation and the highly conserved histidine 81. Also, their cytoplasmic region is completely different and longer. To study and compare them with their classical counterparts, we transduced them in different cell lines. These studies show that they can pair with the classical α-chains (RT1-Da and H2-Ea) and are expressed at the cell surface where they can present superantigens. Interestingly, compared with the classical molecules, they have an extraordinary capacity to present the superantigen Yersinia pseudotuberculosis mitogen. Taken together, our findings suggest that the b2 genes, together with the respective α-chain genes, encode for H2-E2 or RT1-D2 molecules, which could function as Ag-presenting molecules for a particular class of Ags, as modulators of Ag presentation like nonclassical nonpolymorphic class II molecules DM and DO do, or even as players outside the immune system. PMID:26740108

  13. DNA Vaccines: MHC II-Targeted Vaccine Protein Produced by Transfected Muscle Fibres Induces a Local Inflammatory Cell Infiltrate in Mice

    PubMed Central

    Løvås, Tom-Ole; Gundersen, Kristian; Bogen, Bjarne

    2014-01-01

    Vaccination with naked DNA holds great promise but immunogenicity needs to be improved. DNA constructs encoding bivalent proteins that bind antigen-presenting cells (APC) for delivery of antigen have been shown to enhance T and B cell responses and protection in tumour challenge experiments. However, the mechanism for the increased potency remains to be determined. Here we have constructed DNA vaccines that express the fluorescent protein mCherry, a strategy which allowed tracking of vaccine proteins. Transfected muscle fibres in mice were visualized, and their relationship to infiltrating mononuclear cells could be determined. Interestingly, muscle fibers that produced MHC class II-specific dimeric vaccine proteins with mCherry were for weeks surrounded by a localized intense cellular infiltrate composed of CD45+, MHC class II+ and CD11b+ cells. Increasing numbers of eosinophils were observed among the infiltrating cells from day 7 after immunization. The local infiltrate surrounding mCherry+ muscle fibers was dependent on the MHC II-specificity of the vaccine proteins since the control, a non-targeted vaccine protein, failed to induce similar infiltrates. Chemokines measured on day 3 in immunized muscle indicate both a DNA effect and an electroporation effect. No influence of targeting was observed. These results contribute to our understanding for why targeted DNA vaccines have an improved immunogenicity. PMID:25299691

  14. Extraordinary MHC class II B diversity in a non-passerine, wild bird: the Eurasian Coot Fulica atra (Aves: Rallidae)

    PubMed Central

    Alcaide, Miguel; Muñoz, Joaquin; Martínez-de la Puente, Josué; Soriguer, Ramón; Figuerola, Jordi

    2014-01-01

    The major histocompatibility complex (MHC) hosts the most polymorphic genes ever described in vertebrates. The MHC triggers the adaptive branch of the immune response, and its extraordinary variability is considered an evolutionary consequence of pathogen pressure. The last few years have witnessed the characterization of the MHC multigene family in a large diversity of bird species, unraveling important differences in its polymorphism, complexity, and evolution. Here, we characterize the first MHC class II B sequences isolated from a Rallidae species, the Eurasian Coot Fulica atra. A next-generation sequencing approach revealed up to 265 alleles that translated into 251 different amino acid sequences (β chain, exon 2) in 902 individuals. Bayesian inference identified up to 19 codons within the presumptive peptide-binding region showing pervasive evidence of positive, diversifying selection. Our analyses also detected a significant excess of high-frequency segregating sites (average Tajima's D = 2.36, P < 0.05), indicative of balancing selection. We found one to six different alleles per individual, consistent with the occurrence of at least three MHC class II B gene duplicates. However, the genotypes comprised of three alleles were by far the most abundant in the population investigated (49.4%), followed by those with two (29.6%) and four (17.5%) alleles. We suggest that these proportions are in agreement with the segregation of MHC haplotypes differing in gene copy number. The most widespread segregating haplotypes, according to our findings, would contain one single gene or two genes. The MHC class II of the Eurasian Coot is a valuable system to investigate the evolutionary implications of gene copy variation and extensive variability, the greatest ever found, to the best of our knowledge, in a wild population of a non-passerine bird. PMID:24683452

  15. Detection of Foreign Antigen-specific CD4+Foxp3+ Regulatory T Cells by MHC Class II Tetramer and Intracellular CD154 Staining

    PubMed Central

    Choi, Jin Young

    2013-01-01

    The unrestricted population of CD4+Foxp3+ regulatory T (Treg) cells, which have been known to control the expression of autoimmune diseases and protective immunity to inflammatory reactions, has led to greater appreciation of functional plasticity. Detecting and/or isolating Ag-specific CD4+Foxp3+ Tregs at the single cell level are required to study their function and plasticity. In this study, we established and compared both MHC class II tetramer and intracellular CD154 staining, in order to detect CD4+Foxp3+ Treg specific for foreign Ag in acute and chronic infections with lymphocytic choriomeningitis virus (LCMV). Our results revealed that MHC class II tetramer staining showed a lower detection rate of LCMV GP66-77-specific CD4+ T cells because most of MHC class II tetramers were unbound and unstable when combined staining was performed with intracellular cytokines. In contrast, intracellular CD154 staining was revealed to be easier and simple for detecting LCMV GP66-77-specific CD4+ T cells, compared to MHC class II tetramer staining. Subsequently, we employed intracellular CD154 staining to detect LCMV GP66-77-specific CD4+Foxp3+ Tregs using Foxp3GFP knock-in mouse, and found that LCMV GP66-77-specific CD4+Foxp3+ Tregs and polyclonal CD4+Foxp3+ Tregs showed differential expansion in mice infected with LCMV Arms or Cl13 at acute (8 and 13 days pi) and chronic phases (35 days pi). Therefore, our results provide insight into the valuable use of intracellular CD154 staining to detect and characterize foreign Ag-specific CD4+Foxp3+ Treg in various models. PMID:24385945

  16. Induction of tolerance against the arthritogenic antigen with type-II collagen peptide-linked soluble MHC class II molecules.

    PubMed

    Park, Yoon-Kyung; Jung, Sundo; Park, Se-Ho

    2016-06-01

    In murine collagen-induced arthritis (CIA), self-reactive T cells can recognize peptide antigens derived from type-II collagen (CII). Activation of T cells is an important mediator of autoimmune diseases. Thus, T cells have become a focal point of study to treat autoimmune diseases. In this study, we evaluated the efficacy of recombinant MHC class II molecules in the regulation of antigen-specific T cells by using a self peptide derived from CII (CII260-274; IAGFKGEQGPKGEPG) linked to mouse I-A(q) in a murine CIA model. We found that recombinant I-A(q)/CII260-274 molecules could be recognized by CII-specific T cells and inhibit the same T cells in vitro. Furthermore, the development of CIA in mice was successfully prevented by in vivo injection of recombinant I-A(q)/CII260-274 molecules. Thus, treatment with recombinant soluble MHC class II molecules in complex with an immunodominant self-peptide might offer a potential therapeutic for chronic inflammation in autoimmune disease such as rheumatoid arthritis. [BMB Reports 2016; 49(6): 331-336]. PMID:26779996

  17. Immunotherapy augments the effect of 5-azacytidine on HPV16-associated tumours with different MHC class I-expression status

    PubMed Central

    Šímová, J; Polláková, V; Indrová, M; Mikyšková, R; Bieblová, J; Štěpánek, I; Bubeník, J; Reiniš, M

    2011-01-01

    Background: Epigenetic mechanisms have important roles in the tumour escape from immune responses, such as in MHC class I downregulation or altered expression of other components involved in antigen presentation. Chemotherapy with DNA methyltransferase inhibitors (DNMTi) can thus influence the tumour cell interactions with the immune system and their sensitivity to immunotherapy. Methods: We evaluated the therapeutic effects of the DNMTi 5-azacytidine (5AC) against experimental MHC class I-deficient and -positive tumours. The 5AC therapy was combined with immunotherapy, using a murine model for HPV16-associated tumours. Results: We have demonstrated 5AC additive effects against MHC class I-positive and -deficient tumours when combined with unmethylated CpG oligodeoxynucleotides or with IL-12-producing cellular vaccine. The efficacy of the combined chemoimmunotherapy against originally MHC class I-deficient tumours was partially dependent on the CD8+-mediated immune responses. Increased cell surface expression of MHC class I cell molecules, associated with upregulation of the antigen-presenting machinery-related genes, as well as of genes encoding selected components of the IFNγ-signalling pathway in tumours explanted from 5AC-treated animals, were observed. Conclusion: Our data suggest that chemotherapy of MHC class I-deficient tumours with 5AC combined with immunotherapy is an attractive setting in the treatment of MHC class I-deficient tumours. PMID:22015556

  18. Expression Regulation of Major Histocompatibility Complex Class I and Class II Encoding Genes

    PubMed Central

    van den Elsen, Peter J.

    2011-01-01

    Major histocompatibility complex (MHC)-I and MHC-II molecules play an essential role in the immune response to pathogens by virtue of their ability to present peptides to CD8+ and CD4+ T cells, respectively. Given this critical role, MHC-I and MHC-II genes are regulated in a tight fashion at the transcriptional level by a variety of transcription factors that interact with conserved cis-acting regulatory promoter elements. In addition to the activities of these regulatory factors, modification of chromatin also plays an essential role in the efficient transcription of these genes to meet with local requirement for an effective immune response. The focus of this review is on the transcription factors that interact with conserved cis-acting promoter elements and the epigenetic mechanisms that modulate induced and constitutive expression of these MHC genes. PMID:22566838

  19. LAMP-2C Inhibits MHC Class II Presentation of Cytoplasmic Antigens by Disrupting Chaperone-Mediated Autophagy.

    PubMed

    Pérez, Liliana; McLetchie, Shawna; Gardiner, Gail J; Deffit, Sarah N; Zhou, Delu; Blum, Janice S

    2016-03-15

    Cells use multiple autophagy pathways to sequester macromolecules, senescent organelles, and pathogens. Several conserved isoforms of the lysosome-associated membrane protein-2 (LAMP-2) regulate these pathways influencing immune recognition and responses. LAMP-2A is required for chaperone-mediated autophagy (CMA), which promotes Ag capture and MHC class II (MHCII) presentation in B cells and signaling in T cells. LAMP-2B regulates lysosome maturation to impact macroautophagy and phagocytosis. Yet, far less is known about LAMP-2C function. Whereas LAMP2A and LAMP2B mRNA were broadly detected in human tissues, LAMP2C expression was more limited. Transcripts for the three LAMP2 isoforms increased with B cell activation, although specific gene induction varied depending on TLR versus BCR engagement. To examine LAMP-2C function in human B cells and specifically its role in Ag presentation, we used ectopic gene expression. Increased LAMP-2C expression in B cells did not alter MHCII expression or invariant chain processing, but did perturb cytoplasmic Ag presentation via CMA. MHCII presentation of epitopes from exogenous and membrane Ags was not affected by LAMP-2C expression in B cells. Similarly, changes in B cell LAMP-2C expression did not impact macroautophagy. The gene expression of other LAMP2 isoforms and proteasome and lysosomal proteases activities were unperturbed by LAMP-2C ectopic expression. LAMP-2C levels modulated the steady-state expression of several cytoplasmic proteins that are targeted for degradation by CMA and diminished peptide translocation via this pathway. Thus, LAMP-2C serves as a natural inhibitor of CMA that can selectively skew MHCII presentation of cytoplasmic Ags. PMID:26856698

  20. A predominant role for the HLA class II region in the association of the MHC region with multiple sclerosis.

    PubMed

    Lincoln, Matthew R; Montpetit, Alexandre; Cader, M Zameel; Saarela, Janna; Dyment, David A; Tiislar, Milvi; Ferretti, Vincent; Tienari, Pentti J; Sadovnick, A Dessa; Peltonen, Leena; Ebers, George C; Hudson, Thomas J

    2005-10-01

    Genetic susceptibility to multiple sclerosis is associated with genes of the major histocompatibility complex (MHC), particularly HLA-DRB1 and HLA-DQB1 (ref. 1). Both locus and allelic heterogeneity have been reported in this genomic region. To clarify whether HLA-DRB1 itself, nearby genes in the region encoding the MHC or combinations of these loci underlie susceptibility to multiple sclerosis, we genotyped 1,185 Canadian and Finnish families with multiple sclerosis (n = 4,203 individuals) with a high-density SNP panel spanning the genes encoding the MHC and flanking genomic regions. Strong associations in Canadian and Finnish samples were observed with blocks in the HLA class II genomic region (P < 4.9 x 10(-13) and P < 2.0 x 10(-16), respectively), but the strongest association was with HLA-DRB1 (P < 4.4 x 10(-17)). Conditioning on either HLA-DRB1 or the most significant HLA class II haplotype block found no additional block or SNP association independent of the HLA class II genomic region. This study therefore indicates that MHC-associated susceptibility to multiple sclerosis is determined by HLA class II alleles, their interactions and closely neighboring variants. PMID:16186814

  1. Genetic characterization of MHC class II DQB exon 2 variants in gayal (Bos frontalis)

    PubMed Central

    Sun, Yongke; Xi, Dongmei; Li, Guozhi; Hao, Tiantian; Chen, Yuhan; Yang, Yuai

    2014-01-01

    In the present study, exon 2 of major histocompatibility complex (MHC) class II DQB gene from 39 gayals (Bos frontalis) was isolated, characterized and compared with previously reported patterns for other bovidae. It was revealed by sequence analyses that there are 36 DQB exon 2 variants among 39 gayals. These variants exhibited a high degree of nucleotide and amino acid substitutions with most amino acid variations occurring at positions forming the peptide-binding sites (PBS). The DQB loci were analysed for patterns of synonymous (d S) and non-synonymous (d N) substitution. The gayals were observed to be under strong balancing selection in the DQB exon 2 PBS (d N = 0.094, P = 0.001). It appears that this variability among gayals could confer the ability to mount immune responses to a wide variety of peptides or pathogens. PMID:26019566

  2. Absence of MHC class II on cDCs results in microbial-dependent intestinal inflammation.

    PubMed

    Loschko, Jakob; Schreiber, Heidi A; Rieke, Gereon J; Esterházy, Daria; Meredith, Matthew M; Pedicord, Virginia A; Yao, Kai-Hui; Caballero, Silvia; Pamer, Eric G; Mucida, Daniel; Nussenzweig, Michel C

    2016-04-01

    Conventional dendritic cells (cDCs) play an essential role in host immunity by initiating adaptive T cell responses and by serving as innate immune sensors. Although both innate and adaptive functions of cDCs are well documented, their relative importance in maintaining immune homeostasis is poorly understood. To examine the significance of cDC-initiated adaptive immunity in maintaining homeostasis, independent of their innate activities, we generated a cDC-specific Cre mouse and crossed it to a floxed MHC class II (MHCII) mouse. Absence of MHCII on cDCs resulted in chronic intestinal inflammation that was alleviated by antibiotic treatment and entirely averted under germ-free conditions. Uncoupling innate and adaptive functions of cDCs revealed that innate immune functions of cDCs are insufficient to maintain homeostasis and antigen presentation by cDCs is essential for a mutualistic relationship between the host and intestinal bacteria. PMID:27001748

  3. Analysis of T-cell hybridomas with an unusual MHC class II-dependent ligand specificity.

    PubMed Central

    Mendiratta, S K; Singh, N; Bal, V; Rath, S

    1996-01-01

    We have characterized two unusual T-cell hybridomas, 1E3 and 3B8, from H-2k mice immunized with I-Ab-transfected L cells (H-2k), that are stimulated by L cells transfected with I-Ab, I-Ak or I-Eb, but not by non-transfected L cells. These hybridomas could not be stimulated by spleen cells from H-2i3, H-2k, H-2b or H-2d mice. Monoclonal anti-I-A antibodies did not block their responses, suggesting that mouse major histocompatibility complex (MHC) class II molecules may be peptide donors rather than restriction elements for them. The stimulation of these hybridomas by fibroblast targets was not blocked by an anti-H-2kk, Dk-specific monoclonal antibody. Lipopolysaccharide (LPS)-activated splenic and peritoneal exudate cells from H-2k, H-2d, H-2i3, H-2b as well as beta 2-microglobulin-deficient, TAP-1-deficient and I-A alpha-deficient H-2b mice stimulated these hybridomas. LPS could also activate a macrophage cell line, but not a B-cell line, to become stimulatory for 1E3. A rat antiserum against untransfected L cells specifically and significantly blocked the response of 1E3. Thus, 1E3 may recognize a conserved murine MHC class II peptide loaded in a TAP-1-independent fashion on a non-classical, monomorphic, beta 2-microglobulin-independent restriction element. PMID:8943720

  4. Outer membrane proteins preferentially load MHC class II peptides: Implications for as a Chlamydia trachomatis T cell vaccine

    PubMed Central

    Karunakaran, Karuna P.; Yu, Hong; Jiang, Xiaozhou; Chan, Queenie; Moon, Kyung-Mee; Foster, Leonard J.; Brunham, Robert C.

    2015-01-01

    CD4 T cell immune responses such as interferon-γ and tumor necrosis factor-α secretion are necessary for Chlamydia immunity. We used an immunoproteomic approach in which Chlamydia trachomatis and Chlamydia muridarum-derived peptides presented by MHC class II molecules on the surface of infected dendritic cells (DCs) were identified by tandem mass spectrometry using bone marrow derived DCs (BMDCs) from mice of different MHC background. We first compared the C. muridarum immunoproteome in C3H mice to that previously identified in C57BL/6 mice. Fourteen MHC class II binding peptides from 11 Chlamydia proteins were identified from C3H infected BMDCs. Two C. muridarum proteins overlapped between C3H and C57B/6 mice and both were polymorphic membrane proteins (Pmps) which presented distinct class II binding peptides. Next we studied DCs from C57BL/6 mice infected with the human strain, C. trachomatis serovar D. Sixty MHC class II binding peptides derived from 27 C. trachomatis proteins were identified. Nine proteins were orthologous T cell antigens between C. trachomatis and C. muridarum and 2 of the nine were Pmps which generated MHC class II binding epitopes at distinct sequences within the proteins. As determined by antigen specific splenocyte responses outer membrane proteins PmpF, -G and -H and the major outer membrane protein (MOMP) were antigenic in mice previously infected with C. muridarum or C. trachomatis. Furthermore a recombinant protein vaccine consisting of the four Pmps (PmpEFGH) with MOMP formulated with a Th1 polarizing adjuvant significantly accelerated (p < 0.001) clearance in the C57BL/6 mice C. trachomatis transcervical infection model. We conclude that Chlamydia outer membrane proteins are important T cell antigens useful in the development of a C. trachomatis subunit vaccine. PMID:25738816

  5. Cholesterol Corrects Altered Conformation of MHC-II Protein in Leishmania donovani Infected Macrophages: Implication in Therapy

    PubMed Central

    Chakrabarti, Saikat; Roy, Syamal

    2016-01-01

    Background Previously we reported that Kala-azar patients show progressive decrease in serum cholesterol as a function of splenic parasite burden. Splenic macrophages (MΦ) of Leishmania donovani (LD) infected mice show decrease in membrane cholesterol, while LD infected macrophages (I-MΦ) show defective T cell stimulating ability that could be corrected by liposomal delivery of cholesterol. T helper cells recognize peptide antigen in the context of class II MHC molecule. It is known that the conformation of a large number of membrane proteins is dependent on membrane cholesterol. In this investigation we tried to understand the influence of decreased membrane cholesterol in I-MΦ on the conformation of MHC-II protein and peptide-MHC-II stability, and its bearing on the antigen specific T-cell activation. Methodology/Principal Findings MΦ of CBA/j mice were infected with Leishmania donovani (I-MΦ). Two different anti-Aκ mAbs were used to monitor the status of MHC-II protein under parasitized condition. One of them (11.5–2) was conformation specific, whereas the other one (10.2.16) was not. Under parasitized condition, the binding of 11.5–2 decreased significantly with respect to the normal counterpart, whereas that of 10.2.16 remained unaltered. The binding of 11.5–2 was restored to normal upon liposomal delivery of cholesterol in I-MΦ. By molecular dynamics (MD) simulation studies we found that there was considerable conformational fluctuation in the transmembrane domain of the MHC-II protein in the presence of membrane cholesterol than in its absence, which possibly influenced the distal peptide binding groove. This was evident from the faster dissociation of the cognate peptide from peptide-MHC complex under parasitized condition, which could be corrected by liposomal delivery of cholesterol in I-MΦ. Conclusion The decrease in membrane cholesterol in I-MΦ may lead to altered conformation of MHC II, and this may contribute to a faster dissociation of

  6. A single nomenclature and associated database for alleles at the MHC class II DRB1 locus of sheep: IPD-MHC-OLA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The development of standardised nomenclatures with associated databases containing reference sequences for alleles at polymorphic loci within the Major Histocompatibility Complex (MHC) has been facilitated by the development of the Immuno Polymorphism Database (IPD-MHC). Recently, included within I...

  7. Construction of bioactive chimeric MHC class I tetramer by expression and purification of human-murine chimeric MHC heavy chain and beta(2)m as a fusion protein in Escherichia coli.

    PubMed

    Ren, Ding; Wang, Fang; He, Xiaowen; Jiang, Lei; Li, Dean; Ying, He; Sun, Shuhan

    2006-12-01

    Major histocompatibility (MHC) class I tetramers are used in the quantitative analysis of epitope peptide-specific CD8+ T-cells. An MHC class I tetramer was composed of 4 MHC class I complexes and a fluorescently labeled streptavidin (SA) molecule. Each MHC class I complex consists of an MHC heavy chain, a beta(2)-microglobulin (beta(2)m) molecule and a synthetic epitope peptide. In most previous studies, an MHC class I complex was formed in the refolding buffer with an expressed MHC heavy chain molecule and beta(2)m, respectively. This procedure inevitably resulted in the disadvantages of forming unwanted multimers and self-refolding products, and the purification of each kind of monomer was time-consuming. In the present study, the genes of a human/murine chimeric MHC heavy chain (HLA-A2 alpha1, HLA-A2 alpha2 and MHC-H2D alpha3) and beta(2)m were tandem-cloned into plasmid pET17b and expressed as a fusion protein. The recombinant fusion protein was refolded with each of the three HLA-A2 restricted peptides (HBc18-27 FLPSDFFPSI, HBx52-60 HLSLRGLPV, and HBx92-100 VLHKRTLGL) and thus three chimeric MHC class I complexes were obtained. Biotinylation was performed, and its level of efficiency was observed via a band-shift assay in non-reducing polyacrylamide gel electrophoresis (PAGE). Such chimeric MHC class I tetramers showed a sensitive binding activity in monitoring HLA/A2 restrictive cytotoxic T lymphocytes (CTLs) in immunized HLA/A*0201 transgenic mice. PMID:17046278

  8. Adrenomedullin 2 Improves Early Obesity-Induced Adipose Insulin Resistance by Inhibiting the Class II MHC in Adipocytes.

    PubMed

    Zhang, Song-Yang; Lv, Ying; Zhang, Heng; Gao, Song; Wang, Ting; Feng, Juan; Wang, Yuhui; Liu, George; Xu, Ming-Jiang; Wang, Xian; Jiang, Changtao

    2016-08-01

    MHC class II (MHCII) antigen presentation in adipocytes was reported to trigger early adipose inflammation and insulin resistance. However, the benefits of MHCII inhibition in adipocytes remain largely unknown. Here, we showed that human plasma polypeptide adrenomedullin 2 (ADM2) levels were negatively correlated with HOMA of insulin resistance in obese human. Adipose-specific human ADM2 transgenic (aADM2-tg) mice were generated. The aADM2-tg mice displayed improvements in high-fat diet-induced early adipose insulin resistance. This was associated with increased insulin signaling and decreased systemic inflammation. ADM2 dose-dependently inhibited CIITA-induced MHCII expression by increasing Blimp1 expression in a CRLR/RAMP1-cAMP-dependent manner in cultured adipocytes. Furthermore, ADM2 treatment restored the high-fat diet-induced early insulin resistance in adipose tissue, mainly via inhibition of adipocyte MHCII antigen presentation and CD4(+) T-cell activation. This study demonstrates that ADM2 is a promising candidate for the treatment of early obesity-induced insulin resistance. PMID:27207558

  9. Dissection of the role of MHC class II A and E genes in autoimmune susceptibility in murine lupus models with intragenic recombination.

    PubMed

    Zhang, Danqing; Fujio, Keishi; Jiang, Yi; Zhao, Jingyuan; Tada, Norihiro; Sudo, Katsuko; Tsurui, Hiromichi; Nakamura, Kazuhiro; Yamamoto, Kazuhiko; Nishimura, Hiroyuki; Shira, Toshikazu; Hirose, Sachiko

    2004-09-21

    Systemic lupus erythematosus (SLE) is a multigenic autoimmune disease, and the major histocompatibility complex (MHC) class II polymorphism serves as a key genetic element. In SLE-prone (NZB x NZW)F(1) mice, the MHC H-2(d/z) heterozygosity (H-2(d) of NZB and H-2(z) of NZW) has a strong impact on disease; thus, congenic H-2(d/d) homozygous F(1) mice do not develop severe disease. In this study, we used Ea-deficient intra-H-2 recombination to establish A(d/d)-congenic (NZB x NZW)F(1) mice, with or without E molecule expression, and dissected the role of class II A and E molecules. Here we found that A(d/d) homozygous F(1) mice lacking E molecules developed severe SLE similar to that seen in wild-type F1 mice, including lupus nephritis, autoantibody production, and spontaneously occurring T cell activation. Additional evidence revealed that E molecules prevent the disease in a dose-dependent manner; however, the effect is greatly influenced by the haplotype of A molecules, because wild-type H-2(d/z) F(1) mice develop SLE, despite E molecule expression. Studies on the potential of dendritic cells to present a self-antigen chromatin indicated that dendritic cells from wild-type F(1) mice induced a greater response of chromatin-specific T cells than did those from A(d/d) F(1) mice, irrespective of the presence or absence of E molecules, suggesting that the self-antigen presentation is mediated by A, but not by E, molecules. Our mouse models are useful for analyzing the molecular mechanisms by which MHC class II regions regulate the process of autoimmune responses. PMID:15361580

  10. Genomic organization of the crested ibis MHC provides new insight into ancestral avian MHC structure

    PubMed Central

    Chen, Li-Cheng; Lan, Hong; Sun, Li; Deng, Yan-Li; Tang, Ke-Yi; Wan, Qiu-Hong

    2015-01-01

    The major histocompatibility complex (MHC) plays an important role in immune response. Avian MHCs are not well characterized, only reporting highly compact Galliformes MHCs and extensively fragmented zebra finch MHC. We report the first genomic structure of an endangered Pelecaniformes (crested ibis) MHC containing 54 genes in three regions spanning ~500 kb. In contrast to the loose BG (26 loci within 265 kb) and Class I (11 within 150) genomic structures, the Core Region is condensed (17 within 85). Furthermore, this Region exhibits a COL11A2 gene, followed by four tandem MHC class II αβ dyads retaining two suites of anciently duplicated “αβ” lineages. Thus, the crested ibis MHC structure is entirely different from the known avian MHC architectures but similar to that of mammalian MHCs, suggesting that the fundamental structure of ancestral avian class II MHCs should be “COL11A2-IIαβ1-IIαβ2.” The gene structures, residue characteristics, and expression levels of the five class I genes reveal inter-locus functional divergence. However, phylogenetic analysis indicates that these five genes generate a well-supported intra-species clade, showing evidence for recent duplications. Our analyses suggest dramatic structural variation among avian MHC lineages, help elucidate avian MHC evolution, and provide a foundation for future conservation studies. PMID:25608659

  11. MHC class II alleles and haplotypes in patients with pemphigus vulgaris from India.

    PubMed

    Delgado, J C; Yunis, D E; Bozón, M V; Salazar, M; Deulofeut, R; Turbay, D; Mehra, N K; Pasricha, J S; Raval, R S; Patel, H; Shah, B K; Bhol, K; Alper, C A; Ahmed, A R; Yunis, E J

    1996-12-01

    Pemphigus vulgaris (PV) is a blistering disease of the skin and mucous membranes characterized by an autoantibody response against a keratinocyte adhesion molecule, desmoglein 3, causing acantholysis and blister formation. We compared high resolution MHC class II alleles and haplotype frequencies (HLA-DRB, DQA1 and DQB1) in 37 patients with PV to 89 haplotypes of normal relatives from New Delhi and Ahmedabad. We found that PV patients had significantly increased frequencies of DRB1*1404 (P < 0.0001), DQA1*0101 (P = 0.001), and DQB1*0503 (P < 0.0001). These associations were due to the increased frequencies of the haplotype HLA-DRB1*1404, DRB3*0202, DQA1*0101, DQB1*0503 in patients compared to control haplotypes (p < 0.0001). Also, patients from Ahmedabad had a significant increase in HLA-DQB1*0302 (p = 0.03). An identical amino acid sequence (Leu-Leu-Glu-Arg-Arg-Arg-Ala-Glu), in positions 67-74 of the beta domain of DRB alleles is restricted to some DR14 alleles. Therefore, there are three possible explanations for class II allele involvement in autoantibody in PV patients with class II haplotypes marked by HLA-DR14. First, the class II alleles could be markers for an unidentified susceptibility gene in linkage disequilibrium with them. Second, the primary association could be with DQB1*0503 and the association with HLA-DR14 alleles would be the result of linkage disequilibrium. Third, the HLA-DRB1 locus susceptibility could involve a specific amino acid sequence in the third hypervariable region shared by several HLA-DR14 alleles. PMID:9008309

  12. Schwann cell differentiation inhibits interferon-gamma induction of expression of major histocompatibility complex class II and intercellular adhesion molecule-1.

    PubMed

    Lisak, Robert P; Bealmear, Beverly; Benjamins, Joyce A

    2016-06-15

    Interferon-gamma (IFN-γ) upregulates major histocompatibility complex class II (MHC class II) antigens and intercellular adhesion molecule-1 (ICAM-1) on Schwann cells (SC) in vitro, but in nerves of animals and patients MHC class II is primarily expressed on inflammatory cells. We investigated whether SC maturation influences their expression. IFN-γ induced MHC class II and upregulated ICAM-1; the axolemma-like signal 8-bromo cyclic adenosine monophosphate (8 Br cAMP) with IFN-γ inhibited expression. Delaying addition of 8 Br cAMP to SC already exposed to IFN-γ inhibited ongoing expression; addition of IFN-γ to SC already exposed to 8 Br cAMP resulted in minimal expression. Variability of cytokine-induced MHC class II and ICAM-1 expression by SC in vivo may represent the variability of signals from axolemma. PMID:27235355

  13. TCR-like antibodies distinguish conformational and functional differences in two- versus four-domain auto reactive MHC class II-peptide complexes.

    PubMed

    Dahan, Rony; Tabul, Moran; Chou, Yuan K; Meza-Romero, Roberto; Andrew, Shayne; Ferro, Adolph J; Burrows, Gregory G; Offner, Halina; Vandenbark, Arthur A; Reiter, Yoram

    2011-05-01

    Antigen-presenting cell-associated four-domain MHC class II (MHC-II) molecules play a central role in activating autoreactive CD4(+) T cells involved in multiple sclerosis (MS) and type 1 diabetes (T1D). In contrast, two-domain MHC-II structures with the same covalently attached self-peptide (recombinant T-cell receptor ligands (RTLs)) can regulate pathogenic CD4(+) T cells and reverse clinical signs of experimental autoimmune diseases. RTL1000, which is composed of the β1α1 domains of human leukocyte antigen (HLA)-DR2 linked to the encephalitogenic human myelin oligodendrocyte glycoprotein (MOG)-35-55 peptide, was recently shown to be safe and well tolerated in a phase I clinical trial in MS. To evaluate the opposing biological effects of four- versus two-domain MHC-II structures, we screened phage Fab antibodies (Abs) for the neutralizing activity of RTL1000. Five different TCR-like Abs were identified that could distinguish between the two- versus four-domain MHC-peptide complexes while the cognate TCR was unable to make such a distinction. Moreover, Fab detection of native two-domain HLA-DR structures in human plasma implies that there are naturally occurring regulatory MHC-peptide complexes. These results demonstrate for the first time distinct conformational determinants characteristic of activating versus tolerogenic MHC-peptide complexes involved in human autoimmunity. PMID:21469129

  14. RNA-seq liver transcriptome analysis reveals an activated MHC-I pathway and an inhibited MHC-II pathway at the early stage of vaccine immunization in zebrafish

    PubMed Central

    2012-01-01

    Background Zebrafish (Danio rerio) is a prominent vertebrate model of human development and pathogenic disease and has recently been utilized to study teleost immune responses to infectious agents threatening the aquaculture industry. In this work, to clarify the host immune mechanisms underlying the protective effects of a putative vaccine and improve its immunogenicity in the future efforts, high-throughput RNA sequencing technology was used to investigate the immunization-related gene expression patterns of zebrafish immunized with Edwardsiella tarda live attenuated vaccine. Results Average reads of 18.13 million and 14.27 million were obtained from livers of zebrafish immunized with phosphate buffered saline (mock) and E. tarda vaccine (WED), respectively. The reads were annotated with the Ensembl zebrafish database before differential expressed genes sequencing (DESeq) comparative analysis, which identified 4565 significantly differentially expressed genes (2186 up-regulated and 2379 down-regulated in WED; p<0.05). Among those, functional classifications were found in the Gene Ontology database for 3891 and in the Kyoto Encyclopedia of Genes and Genomes database for 3467. Several pathways involved in acute phase response, complement activation, immune/defense response, and antigen processing and presentation were remarkably affected at the early stage of WED immunization. Further qPCR analysis confirmed that the genes encoding the factors involved in major histocompatibility complex (MHC)-I processing pathway were up-regulated, while those involved in MHC-II pathway were down-regulated. Conclusion These data provided insights into the molecular mechanisms underlying zebrafish immune response to WED immunization and might aid future studies to develop a highly immunogenic vaccine against gram-negative bacteria in teleosts. PMID:22805612

  15. NetMHCIIpan-3.0, a common pan-specific MHC class II prediction method including all three human MHC class II isotypes, HLA-DR, HLA-DP and HLA-DQ

    PubMed Central

    Karosiene, Edita; Rasmussen, Michael; Blicher, Thomas; Lund, Ole; Buus, Søren; Nielsen, Morten

    2013-01-01

    Major histocompatibility complex class II (MHCII) molecules play an important role in cell-mediated immunity. They present specific peptides derived from endosomal proteins for recognition by T helper cells. The identification of peptides that bind to MHCII molecules is therefore of great importance for understanding the nature of immune responses and identifying T cell epitopes for the design of new vaccines and immunotherapies. Given the large number of MHC variants, and the costly experimental procedures needed to evaluate individual peptide–MHC interactions, computational predictions have become particularly attractive as first-line methods in epitope discovery. However, only a few so-called pan-specific prediction methods capable of predicting binding to any MHC molecule with known protein sequence are currently available, and all of them are limited to HLA-DR. Here, we present the first pan-specific method capable of predicting peptide binding to any HLA class II molecule with a defined protein sequence. The method employs a strategy common for HLA-DR, HLA-DP and HLA-DQ molecules to define the peptide-binding MHC environment in terms of a pseudo sequence. This strategy allows the inclusion of new molecules even from other species. The method was evaluated in several benchmarks and demonstrates a significant improvement over molecule-specific methods as well as the ability to predict peptide binding of previously uncharacterised MHCII molecules. To the best of our knowledge, the NetMHCIIpan-3.0 method is the first pan-specific predictor covering all HLA class II molecules with known sequences including HLA-DR, HLA-DP, and HLA-DQ. The NetMHCpan-3.0 method is available at http://www.cbs.dtu.dk/services/NetMHCIIpan-3.0. PMID:23900783

  16. Translational diffusion of individual class II MHC membrane proteins in cells.

    PubMed Central

    Vrljic, Marija; Nishimura, Stefanie Y; Brasselet, Sophie; Moerner, W E; McConnell, Harden M

    2002-01-01

    Single-molecule epifluorescence microscopy was used to observe the translational motion of GPI-linked and native I-E(k) class II MHC membrane proteins in the plasma membrane of CHO cells. The purpose of the study was to look for deviations from Brownian diffusion that might arise from barriers to this motion. Detergent extraction had suggested that these proteins may be confined to lipid microdomains in the plasma membrane. The individual I-E(k) proteins were visualized with a Cy5-labeled peptide that binds to a specific extracytoplasmic site common to both proteins. Single-molecule trajectories were used to compute a radial distribution of displacements, yielding average diffusion coefficients equal to 0.22 (GPI-linked I-E(k)) and 0.18 microm(2)/s (native I-E(k)). The relative diffusion of pairs of proteins was also studied for intermolecular separations in the range 0.3-1.0 microm, to distinguish between free diffusion of a protein molecule and diffusion of proteins restricted to a rapidly diffusing small domain. Both analyses show that motion is predominantly Brownian. This study finds no strong evidence for significant confinement of either GPI-linked or native I-E(k) in the plasma membrane of CHO cells. PMID:12414700

  17. Porcine MHC classical class I genes are coordinately expressed in superantigen-activated mononuclear cells.

    PubMed

    Kametani, Yoshie; Ohshima, Shino; Kita, Yuki F; Shimada, Shin; Kamiguchi, Hiroshi; Shiina, Takashi; Inoko, Hidetoshi; Kulski, Jerzy K; Ando, Asako

    2012-08-15

    The expression of the major histocompatibility complex (MHC) classical class I genes is important for the adaptive immune response to target virus-infected cells and cancer cells. The up-regulation of the MHC is achieved by hormonal/cytokine signals including IFN-γ-inducible elements. The swine leukocyte antigen (SLA), the MHC class I region of pigs, consists of the duplicated classical class I genes, SLA-1, SLA-2 and SLA-3, but the molecular mechanisms involved in their up-regulation after T cell stimulation have not been fully elucidated. In order to better understand some of the putative regulatory mechanisms of SLA class I gene expression in activated T cells, we examined the coordinated expression of the SLA classical class I, IFN-γ and interferon regulatory factor-1 (IRF-1) genes in the peripheral blood mononuclear cells (PBMCs) of SLA homozygous Clawn miniature swine stimulated for 72 h with either IFN-γ or an enterotoxin produced by Staphylococcus aureus. This enterotoxin, toxic shock syndrome-1 (TSST-1), is known to act as a superantigen (sAG) to activate the T cells in various vertebrate species. We showed by using mAbs and flow cytometry that the CD4(+)CD25(+) cell number of swine PBMCs was also increased by TSST-1 and to a lesser degree by IFN-γ. Time course analyses of the expression of the IFN-γ, IRF-1 and the three classical class I genes, SLA-1, SLA-2, and SLA-3, in PBMCs by quantitative real-time PCR revealed a transitory response to TSST-1 or IFN-γ stimulation. The IFN-γ mRNA levels in the PBMCs were continuously up-regulated over the first 48 h by TSST-1 or IFN-γ. In contrast, SLA class I expression moderately increased at 24h and then decreased to a baseline level or less at 72 h of IFN-γ or TSST-1 stimulation. The three classical SLA class I genes showed similar expression kinetics, although SLA-3 mRNA level was consistently lower than those of SLA-1 and -2. The expression of IRF-1, a modulator of SLA expression, showed similar

  18. Positive regulatory domain I binding factor 1 silences class II transactivator expression in multiple myeloma cells.

    PubMed

    Ghosh, N; Gyory, I; Wright, G; Wood, J; Wright, K L

    2001-05-01

    The major histocompatibility complex (MHC) class II transactivator (CIITA) acts as a master switch to activate expression of the genes required for MHC-II antigen presentation. During B-cell to plasma cell differentiation, MHC-II expression is actively silenced, but the mechanism has been unknown. In plasma cell tumors such as multiple myeloma the repression of MHC-II is associated with the loss of CIITA. We have identified that positive regulatory domain I binding factor 1 (PRDI-BF1), a transcriptional repressor, inhibits CIITA expression in multiple myeloma cell lines. Repression of CIITA depends on the DNA binding activity of PRDI-BF1 and its specific binding site in the CIITA promoter. Deletion of a histone deacetylase recruitment domain in PRDI-BF1 does not inhibit repression of CIITA nor does blocking histone deacetylase activity. This is in contrast to PRDI-BF1 repression of the c-myc promoter. Repression of CIITA requires either the N-terminal acidic and conserved PR motif or the proline-rich domain. PRDI-BF1 has been shown to be a key regulator of B-cell and macrophage differentiation. These findings now indicate that PRDI-BF1 has at least two mechanisms of repression whose function is dependent on the nature of the target promoter. Importantly, PRDI-BF1 is defined as the key molecule in silencing CIITA and thus MHC-II in multiple myeloma cells. PMID:11279146

  19. Molecular characterization of MHC class II in the Australian invasive cane toad reveals multiple splice variants.

    PubMed

    Lillie, Mette; Cui, Jian; Shine, Richard; Belov, Katherine

    2016-07-01

    The cane toad has gained notoriety for its invasion across the Australian landscape, with significant impacts on the native Australian fauna. The invasion has accelerated over time, with invading cane toads adapted for highly dispersive traits. This, however, has come at the cost of the immune system, with lower investment in some immune functions. To investigate the cane toad's immunogenetics, we characterized four major histocompatibility complex (MHC) class IIA and three MHC class IIB loci. Preliminary observations suggest very low allelic diversity at all loci. We also observed various splice isoforms. One isoform seen at one class IIA and two class IIB loci was missing exon 2, which is essential to peptide binding and presentation. The other isoform, observed at a class IIA locus, is likely to be a soluble MHC product. These results may suggest a significant role of alternative splicing of MHC loci in the Australian cane toad. PMID:27233954

  20. Presentation of antagonist peptides to naive CD4+ T cells abrogates spatial reorganization of class II MHC peptide complexes on the surface of dendritic cells

    PubMed Central

    Chmielowski, Bartosz; Pacholczyk, Rafal; Kraj, Piotr; Kisielow, Pawel; Ignatowicz, Leszek

    2002-01-01

    By using dendritic cells (DCs) transduced with retroviruses encoding covalent Abβ/peptide fusion proteins tagged with fluorescent proteins, we followed the relocation of class II MHC molecules loaded with agonist or null peptides during the onset of activation of naive and effector CD4+ T cells. Clusters of T cell receptor (TCR)/CD3 complex formed in parallel with clusters of agonist class II MHC/peptide complexes on the surface of DCs. However, activation of naive but not effector T cells was accompanied by expulsion of the null class II MHC/peptide complexes from the T cell–DC interface. These effects were perturbed in the presence of exogenously supplied antagonist peptide. These results suggest that interference with selective relocation of agonist and null MHC/peptide complexes in the immunological synapse contributes to the inhibitory effect of antagonist peptides on the response of naive CD4+ T cells to agonist ligands. PMID:12411579

  1. MMTV superantigens coerce an unconventional topology between the TCR and MHC class II.

    PubMed

    Fortin, Jean-Simon; Genève, Laetitia; Gauthier, Catherine; Shoukry, Naglaa H; Azar, Georges A; Younes, Souheil; Yassine-Diab, Bader; Sékaly, Rafick-Pierre; Fremont, Daved H; Thibodeau, Jacques

    2014-02-15

    Mouse mammary tumor virus superantigens (vSAGs) are notorious for defying structural characterization, and a consensus has yet to be reached regarding their ability to bridge the TCR to MHC class II (MHCII). In this study, we determined the topology of the T cell signaling complex by examining the respective relation of vSAG7 with the MHCII molecule, MHCII-associated peptide, and TCR. We used covalently linked peptide/MHCII complexes to demonstrate that vSAG presentation is tolerant to variation in the protruding side chains of the peptide, but can be sensitive to the nature of the protruding N-terminal extension. An original approach in which vSAG was covalently linked to either MHCII chain confirmed that vSAG binds outside the peptide binding groove. Also, whereas the C-terminal vSAG segment binds to the MHCII α-chain in a conformation-sensitive manner, the membrane-proximal N-terminal domain binds the β-chain. Because both moieties of the mature vSAG remain noncovalently associated after processing, our results suggest that vSAG crosslinks MHCII molecules. Comparing different T cell hybridomas, we identified key residues on the MHCII α-chain that are differentially recognized by the CDR3β when engaged by vSAG. Finally, we show that the highly conserved tyrosine residue found in the vSAg TGXY motif is required for T cell activation. Our results reveal a novel SAG/MHCII/TCR architecture in which vSAGs coerce a near-canonical docking between MHCII and TCR that allows eschewing of traditional CDR3 binding with the associated peptide in favor of MHCII α-chain binding. Our findings highlight the plasticity of the TCR CDRs. PMID:24453254

  2. Genetic variation of the major histocompatibility complex (MHC class II B gene) in the threatened Hume's pheasant, Syrmaticus humiae.

    PubMed

    Chen, Weicai; Bei, Yongjian; Li, Hanhua

    2015-01-01

    Major histocompatibility complex (MHC) genes are the most polymorphic genes in vertebrates and encode molecules that play a crucial role in pathogen resistance. As a result of their diversity, they have received much attention in the fields of evolutionary and conservation biology. Here, we described the genetic variation of MHC class II B (MHCIIB) exon 2 in a wild population of Hume's pheasant (Syrmaticus humiae), which has suffered a dramatic decline in population over the last three decades across its ranges in the face of heavy exploitation and habitat loss. Twenty-four distinct alleles were found in 73 S. humiae specimens. We found seven shared alleles among four geographical groups as well as six rare MHCIIB alleles. Most individuals displayed between one to five alleles, suggesting that there are at least three MHCIIB loci of the Hume's pheasant. The dN ⁄ dS ratio at putative antigen-binding sites (ABS) was significantly greater than one, indicating balancing selection is acting on MHCIIB exon 2. Additionally, recombination and gene conversion contributed to generating MHCIIB diversity in the Hume's pheasant. One to three recombination events and seventy-five significant gene conversion events were observed within the Hume's pheasant MHCIIB loci. The phylogenetic tree and network analysis revealed that the Hume's pheasant alleles do not cluster together, but are scattered through the tree or network indicating a trans-species evolutionary mode. These findings revealed the evolution of the Hume's pheasant MHC after suffering extreme habitat fragmentation. PMID:25629763

  3. MHC class II derived recombinant T cell receptor ligands protect DBA/1LacJ mice from collagen-induced arthritis.

    PubMed

    Huan, Jianya; Kaler, Laurie J; Mooney, Jeffery L; Subramanian, Sandhya; Hopke, Corwyn; Vandenbark, Arthur A; Rosloniec, Edward F; Burrows, Gregory G; Offner, Halina

    2008-01-15

    We previously demonstrated the therapeutic effects of MHC class II derived recombinant T cell receptor ligands (RTL), single-chain two domain complexes of the alpha1 and beta1 domains of MHC class II molecules genetically linked with an immunodominant peptide, in experimental autoimmune encephalomyelitis. In the current study, we produced a monomeric murine I-Aq-derived RTL construct covalently linked with bovine collagen type II peptide (bCII257-270) suitable for use in DBA/1LacJ mice that develop collagen-induced arthritis (CIA), an animal model of human rheumatoid arthritis, after immunization with bCII protein in CFA. In this study, we demonstrate that the I-Aq-derived RTLs reduced the incidence of the disease, suppressed the clinical and histological signs of CIA and induced long-term modulation of T cells specific for arthritogenic Ags. Our results showed that the I-Aq/bCII257-270 molecule could systemically reduce proinflammatory IL-17 and IFN-gamma production and significantly increase anti-inflammatory IL-10, IL-13, and FoxP3 gene expression in splenocytes. Moreover, I-Aq/bCII257-270 molecule could also selectively inhibit IL-1beta, IL-6, and IL-23 expression in local joint tissue. This is the first report demonstrating effective prevention of joint inflammation and clinical signs of CIA with an I-Aq-derived RTL, thus supporting the possible clinical use of this approach for treating rheumatoid arthritis in humans. PMID:18178865

  4. MHC Class II Derived Recombinant T Cell Receptor Ligands Protect DBA/1LacJ Mice from Collagen-Induced Arthritis1

    PubMed Central

    Huan, Jianya; Kaler, Laurie J.; Mooney, Jeffery L.; Subramanian, Sandhya; Hopke, Corwyn; Vandenbark, Arthur A.; Rosloniec, Edward F.; Burrows, Gregory G.; Offner, Halina

    2012-01-01

    We previously demonstrated the therapeutic effects of MHC class II derived recombinant T cell receptor ligands (RTL), single-chain two domain complexes of the α1 and β1 domains of MHC class II molecules genetically linked with an immunodominant peptide, in experimental autoimmune encephalomyelitis. In the current study, we produced a monomeric murine I-Aq-derived RTL construct covalently linked with bovine collagen type II peptide (bCII257–270) suitable for use in DBA/1LacJ mice that develop collagen-induced arthritis (CIA), an animal model of human rheumatoid arthritis, after immunization with bCII protein in CFA. In this study, we demonstrate that the I-Aq-derived RTLs reduced the incidence of the disease, suppressed the clinical and histological signs of CIA and induced long-term modulation of T cells specific for arthritogenic Ags. Our results showed that the I-Aq/bCII257–270 molecule could systemically reduce proinflammatory IL-17 and IFN-γ production and significantly increase anti-inflammatory IL-10, IL-13, and FoxP3 gene expression in splenocytes. Moreover, I-Aq/bCII257–270 molecule could also selectively inhibit IL-1β, IL-6, and IL-23 expression in local joint tissue. This is the first report demonstrating effective prevention of joint inflammation and clinical signs of CIA with an I-Aq-derived RTL, thus supporting the possible clinical use of this approach for treating rheumatoid arthritis in humans. PMID:18178865

  5. Asparagine endopeptidase is not essential for class II MHC antigen presentation but is required for processing of cathepsin L in mice.

    PubMed

    Maehr, René; Hang, Howard C; Mintern, Justine D; Kim, You-Me; Cuvillier, Armelle; Nishimura, Mikio; Yamada, Kenji; Shirahama-Noda, Kanae; Hara-Nishimura, Ikuko; Ploegh, Hidde L

    2005-06-01

    Class II MHC molecules survey the endocytic compartments of APCs and present antigenic peptides to CD4 T cells. In this context, lysosomal proteases are essential not only for the generation of antigenic peptides but also for proteolysis of the invariant chain to allow the maturation of class II MHC molecules. Recent studies with protease inhibitors have implicated the asparagine endopeptidase (AEP) in class II MHC-restricted Ag presentation. We now report that AEP-deficient mice show no differences in processing of the invariant chain or maturation of class II MHC products compared with wild-type mice. In the absence of AEP, presentation to primary T cells of OVA and myelin oligodendrocyte glycoprotein, two Ags that contain asparagine residues within or in proximity to the relevant epitopes was unimpaired. Cathepsin (Cat) L, a lysosomal cysteine protease essential for the development to CD4 and NK T cells, fails to be processed into its mature two-chain form in AEP-deficient cells. Despite this, the numbers of CD4 and NK T cells are normal, showing that the single-chain form of Cat L is sufficient for its function in vivo. We conclude that AEP is essential for processing of Cat L but not for class II MHC-restricted Ag presentation. PMID:15905550

  6. Identification and quantitation of MHC class II-bound peptides from mouse spleen dendritic cells by immunoprecipitation and mass spectrometry analysis

    PubMed Central

    Bozzacco, Leonia; Yu, Haiqiang

    2014-01-01

    Summary Advances in immunology and immune therapies require knowledge of antigenic peptide sequences that are presented on MHC class II and class I molecules of antigen presenting cells. The most specialized antigen presenting cells are dendritic cells (DCs). In the past, the small number of DCs that can be isolated from mouse spleen prevented direct analysis of the MHC II peptide repertoire presented by DCs. Here we describe a protocol that integrates immunological methods (in vivo enrichment of mouse spleen DCs by Flt3L treatment and immunoprecipation of MHC II-peptide complexes), mass spectrometry analysis and peptide synthesis (LC-MS/MS and quantitation analysis for non tryptic peptides) to identify and quantitate the endogenous peptides that are bound to MHC II molecules on DCs. The described method produces quantitative data that are reproducible and reliable enough to cover a wide range of peptide copy numbers. We propose the application of this method in future studies to quantitatively investigate the MHC II repertoire on DCs presented during viral infections or different immunizations in vaccine development research. PMID:23963941

  7. CD8 T cell memory recall is enhanced by novel direct interactions with CD4 T cells enabled by MHC class II transferred from APCs.

    PubMed

    Romagnoli, Pablo A; Premenko-Lanier, Mary F; Loria, Gilbert D; Altman, John D

    2013-01-01

    Protection against many intracellular pathogens is provided by CD8 T cells, which are thought to need CD4 T cell help to develop into effective memory CD8 T cells. Because murine CD8 T cells do not transcribe MHC class II (MHC-II) genes, several models have proposed antigen presenting cells (APCs) as intermediaries required for CD4 T cells to deliver their help to CD8 T cells. Here, we demonstrate the presence of MHC-II molecules on activated murine CD8 T cells in vitro as well as in vivo. These MHC-II molecules are acquired via trogocytosis by CD8 T cells from their activating APCs, particularly CD11c positive dendritic cells (DCs). Transferred MHC-II molecules on activated murine CD8 T cells were functionally competent in stimulating specific indicator CD4 T cells. CD8 T cells that were "helped" in vitro and subsequently allowed to rest in vivo showed enhanced recall responses upon challenge compared to "helpless" CD8 T cells; in contrast, no differences were seen upon immediate challenge. These data indicate that direct CD8:CD4 T cell interactions may significantly contribute to help for CD8 T cells. Furthermore, this mechanism may enable CD8 T cells to communicate with different subsets of interacting CD4 T cells that could modulate immune responses. PMID:23441229

  8. Relationship between target antigens and major histocompatibility complex (MHC) class II genes in producing two pathogenic antibodies simultaneously

    PubMed Central

    Zakka, L R; Keskin, D B; Reche, P; Ahmed, A R

    2010-01-01

    In this report, we present 15 patients with histological and immunopathologically proven pemphigus vulgaris (PV). After a mean of 80 months since the onset of disease, when evaluated serologically, they had antibodies typical of PV and pemphigoid (Pg). Similarly, 18 patients with bullous pemphigoid (BP) and mucous membrane pemphigoid (MMP) were diagnosed on the basis of histology and immunopathology. After a mean of 60 months since the onset of disease, when their sera were evaluated they were found to have Pg and PV autoantibodies. In both groups of patients the diseases were characterized by a chronic course, which included several relapses and recurrences and were non-responsive to conventional therapy. The major histocompatibility complex class II (MHC II) genes were studied in both groups of patients and phenotypes associated typically with them were observed. Hence, in 33 patients, two different pathogenic autoantibodies were detected simultaneously. The authors provide a computer model to show that each MHC II gene has relevant epitopes that recognize the antigens associated with both diseases. Using the databases in these computer models, the authors present the hypothesis that these two autoantibodies are produced simultaneously due to the phenomena of epitope spreading. PMID:21069937

  9. Atheroprotective Vaccination with MHC-II Restricted Peptides from ApoB-100

    PubMed Central

    Tse, Kevin; Gonen, Ayelet; Sidney, John; Ouyang, Hui; Witztum, Joseph L.; Sette, Alessandro; Tse, Harley; Ley, Klaus

    2013-01-01

    Background: Subsets of CD4+ T-cells have been proposed to serve differential roles in the development of atherosclerosis. Some T-cell types are atherogenic (T-helper type 1), while others are thought to be protective (regulatory T-cells). Lineage commitment toward one type of helper T-cell versus another is strongly influenced by the inflammatory context in which antigens are recognized. Immunization of atherosclerosis-prone mice with low-density lipoprotein (LDL) or its oxidized derivative (ox-LDL) is known to be atheroprotective. However, the antigen specificity of the T-cells induced by vaccination and the mechanism of protection are not known. Methods: Identification of two peptide fragments (ApoB3501–3516 and ApoB978–993) from murine ApoB-100 was facilitated using I-Ab prediction models, and their binding to I-Ab determined. Utilizing a vaccination scheme based on complete and incomplete Freund’s adjuvant (CFA and IFA) [1 × CFA + 4 × IFA], we immunized Apoe−/−mice with ApoB3501–3516 or ApoB978–993 emulsified in CFA once and subsequently boosted in IFA four times over 15 weeks. Spleens, lymph nodes, and aortas were harvested and evaluated by flow cytometry and real time RT-PCR. Total atherosclerotic plaque burden was determined by aortic pinning and by aortic root histology. Results: Mice immunized with ApoB3501–3516 or ApoB978–993 demonstrated 40% reduction in overall plaque burden when compared to adjuvant-only control mice. Aortic root frozen sections from ApoB3501–3516 immunized mice showed a >60% reduction in aortic sinus plaque development. Aortas from both ApoB3501–3516 and ApoB978–993 immunized mice contained significantly more mRNA for IL-10. Both antigen-specific IgG1 and IgG2c titers were elevated in ApoB3501–3516 or ApoB978–993 immunized mice, suggesting helper T-cell immune activity after immunization. Conclusion: Our data show that MHC Class II restricted ApoB-100 peptides can be atheroprotective

  10. Structural Basis for the Recognition of Mutant Self by a Tumor-Specific, MHC Class II-Restricted T Cell Receptor

    SciTech Connect

    Deng,L.; Langley, R.; Brown, P.; Xu, G.; Teng, L.; Wang, Q.; Gonzales, M.; Callender, G.; Nishimura, M.; et al.

    2007-01-01

    Structural studies of complexes of T cell receptor (TCR) and peptide-major histocompatibility complex (MHC) have focused on TCRs specific for foreign antigens or native self. An unexplored category of TCRs includes those specific for self determinants bearing alterations resulting from disease, notably cancer. We determined here the structure of a human melanoma-specific TCR (E8) bound to the MHC molecule HLA-DR1 and an epitope from mutant triosephosphate isomerase. The structure had features intermediate between 'anti-foreign' and autoimmune TCR-peptide-MHC class II complexes that may reflect the hybrid nature of altered self. E8 manifested very low affinity for mutant triosephosphate isomerase-HLA-DR1 despite the highly tumor-reactive properties of E8 cells. A second TCR (G4) had even lower affinity but underwent peptide-specific formation of dimers, suggesting this as a mechanism for enhancing low-affinity TCR-peptide-MHC interactions for T cell activation.

  11. A Novel HURRAH Protocol Reveals High Numbers of Monomorphic MHC Class II Loci and Two Asymmetric Multi-Locus Haplotypes in the Père David's Deer

    PubMed Central

    Wan, Qiu-Hong; Zhang, Pei; Ni, Xiao-Wei; Wu, Hai-Long; Chen, Yi-Yan; Kuang, Ye-Ye; Ge, Yun-Fa; Fang, Sheng-Guo

    2011-01-01

    The Père David's deer is a highly inbred, but recovered, species, making it interesting to consider their adaptive molecular evolution from an immunological perspective. Prior to this study, genomic sequencing was the only method for isolating all functional MHC genes within a certain species. Here, we report a novel protocol for isolating MHC class II loci from a species, and its use to investigate the adaptive evolution of this endangered deer at the level of multi-locus haplotypes. This protocol was designated “HURRAH” based on its various steps and used to estimate the total number of MHC class II loci. We confirmed the validity of this novel protocol in the giant panda and then used it to examine the Père David's deer. Our results revealed that the Père David's deer possesses nine MHC class II loci and therefore has more functional MHC class II loci than the eight genome-sequenced mammals for which full MHC data are currently available. This could potentially account at least in part for the strong survival ability of this species in the face of severe bottlenecking. The results from the HURRAH protocol also revealed that: (1) All of the identified MHC class II loci were monomorphic at their antigen-binding regions, although DRA was dimorphic at its cytoplasmic tail; and (2) these genes constituted two asymmetric functional MHC class II multi-locus haplotypes: DRA1*01 ∼ DRB1 ∼ DRB3 ∼ DQA1 ∼ DQB2 (H1) and DRA1*02 ∼ DRB2 ∼ DRB4 ∼ DQA2 ∼ DQB1 (H2). The latter finding indicates that the current members of the deer species have lost the powerful ancestral MHC class II haplotypes of nine or more loci, and have instead fixed two relatively weak haplotypes containing five genes. As a result, the Père David's deer are currently at risk for increased susceptibility to infectious pathogens. PMID:21267075

  12. The Thermodynamic Mechanism of Peptide–MHC Class II Complex Formation Is a Determinant of Susceptibility to HLA-DM

    PubMed Central

    Templeton, Megan; Hoffman, Megan; Castellini, Margaret J.

    2015-01-01

    Peptides bind MHC class II molecules through a thermodynamically nonadditive process consequent to the flexibility of the reactants. Currently, how the specific outcome of this binding process affects the ensuing epitope selection needs resolution. Calorimetric assessment of binding thermodynamics for hemagglutinin 306–319 peptide variants to the human MHC class II HLA-DR1 (DR1) and a mutant DR1 reveals that peptide/DR1 complexes can be formed with different enthalpic and entropic contributions. Complexes formed with a smaller entropic penalty feature circular dichroism spectra consistent with a non–compact form, and molecular dynamics simulation shows a more flexible structure. The opposite binding mode, compact and less flexible, is associated with greater entropic penalty. These structural variations are associated with rearrangements of residues known to be involved in HLA-DR (DM) binding, affinity of DM for the complex, and complex susceptibility to DM-mediated peptide exchange. Thus, the thermodynamic mechanism of peptide binding to DR1 correlates with the structural rigidity of the complex, and DM mediates peptide exchange by “sensing” flexible complexes in which the aforementioned residues are rearranged at a higher frequency than in more rigid ones. PMID:26116504

  13. The Atlantic Salmon MHC class II alpha and beta promoters are active in mammalian cell lines.

    PubMed

    Vestrheim, O; Lundin, M; Syed, M

    2007-01-01

    The major histocompatibility complex class II (MHCII) genes are only constitutively expressed in certain immune response cells such as B cells, macrophages, dendritic cells and other antigen presenting cells. This cell specific expression pattern and the presence of conserved regions such as the X-, X2-, Y-, and W-boxes make the MHCII promoters especially interesting as vector constructs. We tested whether the Atlantic salmon (Salmo salar L.) MHCII promoters can function in cell lines from other organisms. We found that the salmon MHCII alpha and MHCII beta promoters could drive expression of a LacZ reporter gene in adherent lymphoblast cell lines from dog (DH82) and rabbit (HybL-L). This paper shows that the promoters of Atlantic salmon MHCII alpha and beta genes can function in mammalian cell lines. PMID:17934904

  14. TCR-like antibodies distinguish conformational and functional differences in two vs. four-domain auto-reactive MHC II-peptide complexes

    PubMed Central

    Dahan, Rony; Tabul, Moran; Chou, Yuan K.; Meza-Romero, Roberto; Andrew, Shayne; Ferro, Adolph J.; Burrows, Gregory G.; Offner, Halina; Vandenbark, Arthur A.; Reiter, Yoram

    2011-01-01

    SUMMARY Antigen presenting cell-associated four-domain MHC class-II molecules play a central role in activating autoreactive CD4+ T-cells involved in Multiple Sclerosis (MS) and Type 1 Diabetes (T1D). In contrast, two-domain MHC-II structures with the same covalently-attached self peptide (Recombinant T-cell receptor Ligands=RTLs) can regulate pathogenic CD4+ T-cells and reverse clinical signs of experimental autoimmune diseases. RTL1000, comprised of the β1α1 domains of HLA-DR2 linked to the encephalitogenic human MOG-35-55 peptide, was recently shown to be safe and well-tolerated in a Phase I clinical trial in MS. To evaluate the opposing biological effects of four- vs. two-domain class-II structures, we screened phage Fab antibodies (Abs) for neutralizing activity of RTL1000. . Five different TCR-like Abs were identified that could distinguish between the two- vs. four-domain MHC peptide complexes, while the cognate TCR was unable to make such a distinction. Moreover, Fab detection of native two-domain HLA-DR structures in human plasma implies that there are naturally-occurring regulatory MHC-peptide complexes. These results demonstrate for the first time distinct conformational determinants characteristic of activating vs. tolerogenic MHC-peptide complexes involved in human autoimmunity. PMID:21469129

  15. MHC class II variation in a rare and ecological specialist mouse lemur reveals lower allelic richness and contrasting selection patterns compared to a generalist and widespread sympatric congener.

    PubMed

    Pechouskova, Eva; Dammhahn, Melanie; Brameier, Markus; Fichtel, Claudia; Kappeler, Peter M; Huchard, Elise

    2015-04-01

    The polymorphism of immunogenes of the major histocompatibility complex (MHC) is thought to influence the functional plasticity of immune responses and, consequently, the fitness of populations facing heterogeneous pathogenic pressures. Here, we evaluated MHC variation (allelic richness and divergence) and patterns of selection acting on the two highly polymorphic MHC class II loci (DRB and DQB) in the endangered primate Madame Berthe's mouse lemur (Microcebus berthae). Using 454 pyrosequencing, we examined MHC variation in a total of 100 individuals sampled over 9 years in Kirindy Forest, Western Madagascar, and compared our findings with data obtained previously for its sympatric congener, the grey mouse lemur (Microcebus murinus). These species exhibit a contrasting ecology and demography that were expected to affect MHC variation and molecular signatures of selection. We found a lower allelic richness concordant with its low population density, but a similar level of allelic divergence and signals of historical selection in the rare feeding specialist M. berthae compared to the widespread generalist M. murinus. These findings suggest that demographic factors may exert a stronger influence than pathogen-driven selection on current levels of allelic richness in M. berthae. Despite a high sequence similarity between the two congeners, contrasting selection patterns detected at DQB suggest its potential functional divergence. This study represents a first step toward unravelling factors influencing the adaptive divergence of MHC genes between closely related but ecologically differentiated sympatric lemurs and opens new questions regarding potential functional discrepancy that would explain contrasting selection patterns detected at DQB. PMID:25687337

  16. MHC class II restricted innate-like double negative T cells contribute to optimal primary and secondary immunity to Leishmania major.

    PubMed

    Mou, Zhirong; Liu, Dong; Okwor, Ifeoma; Jia, Ping; Orihara, Kanami; Uzonna, Jude Ezeh

    2014-09-01

    Although it is generally believed that CD4(+) T cells play important roles in anti-Leishmania immunity, some studies suggest that they may be dispensable, and that MHC II-restricted CD3(+)CD4(-)CD8(-) (double negative, DN) T cells may be more important in regulating primary anti-Leishmania immunity. In addition, while there are reports of increased numbers of DN T cells in Leishmania-infected patients, dogs and mice, concrete evidence implicating these cells in secondary anti-Leishmania immunity has not yet been documented. Here, we report that DN T cells extensively proliferate and produce effector cytokines (IFN-γ, TNF and IL-17) and granzyme B (GrzB) in the draining lymph nodes and spleens of mice following primary and secondary L. major infections. DN T cells from healed mice display functional characteristics of protective anti-Leishmania memory-like cells: rapid and extensive proliferation and effector cytokines production following L. major challenge in vitro and in vivo. DN T cells express predominantly (> 95%) alpha-beta T cell receptor (αβ TCR), are Leishmania-specific, restricted mostly by MHC class II molecules and display transcriptional profile of innate-like genes. Using in vivo depletion and adoptive transfer studies, we show that DN T cells contribute to optimal primary and secondary anti-Leishmania immunity in mice. These results directly identify DN T cells as important players in effective and protective primary and secondary anti-L. major immunity in experimental cutaneous leishmaniasis. PMID:25233487

  17. Structure of the Epstein-Barr virus gp42 protein bound to the MHC class II recepter HLA-DR1

    SciTech Connect

    Mullen, M.; Haan, K.M.; Longnecker, R.; Jardetzky, T.

    2010-03-08

    Epstein-Barr virus (EBV) causes infectious mononucleosis, establishes long-term latent infections, and is associated with a variety of human tumors. The EBV gp42 glycoprotein binds MHC class II molecules, playing a critical role in infection of B lymphocytes. EBV gp42 belongs to the C-type lectin superfamily, with homology to NK receptors of the immune system. We report the crystal structure of gp42 bound to the human MHC class II molecule HLA-DR1. The gp42 binds HLA-DR1 using a surface site that is distinct from the canonical lectin and NK receptor ligand binding sites. At the canonical ligand binding site, gp42 forms a large hydrophobic groove, which could interact with other ligands necessary for EBV entry, providing a mechanism for coupling MHC recognition and membrane fusion.

  18. Predicting Hemagglutinin MHC-II Ligand Analogues in Anti-TNFα Biologics: Implications for Immunogenicity of Pharmaceutical Proteins

    PubMed Central

    Cauley, Brianna; O’Donnell, Lauren A.; Meng, Wilson S.

    2015-01-01

    The purpose of this study was to evaluate the extent of overlapping immunogenic peptides between three pharmaceutical biologics and influenza viruses. Clinical studies have shown that subsets of patients with rheumatoid arthritis (RA) develop anti-drug antibodies towards anti-TNFα biologics. We postulate that common infectious pathogens, including influenza viruses, may sensitize RA patients toward recombinant proteins. We hypothesize that embedded within infliximab (IFX), adalimumab (ADA), and etanercept (ETN) are ligands of class II major histocompatibility complex (MHC-II) that mimic T cell epitopes derived from influenza hemagglutinin (HA). The rationale is that repeated administration of the biologics would reactivate HA-primed CD4 T cells, stimulating B cells to produce cross-reactive antibodies. Custom scripts were constructed using MATLAB to compare MHC-II ligands of HA and the biologics; all ligands were predicted using tools in Immune Epitope Database and Resources (IEDB). We analyzed three HLA-DR1 alleles (0101, 0401 and 1001) that are prominent in RA patients, and two alleles (0103 and 1502) that are not associated with RA. The results indicate that 0401 would present more analogues of HA ligands in the three anti-TNFα biologics compared to the other alleles. The approach led to identification of potential ligands in IFX and ADA that shares sequence homology with a known HA-specific CD4 T cell epitope. We also discovered a peptide in the complementarity-determining region 3 (CDR-3) of ADA that encompasses both a potential CD4 T cell epitope and a known B cell epitope in HA. The results may help generate new hypotheses for interrogating patient variability of immunogenicity of the anti-TNFα drugs. The approach would aid development of new recombinant biologics by identifying analogues of CD4 T cell epitopes of common pathogens at the preclinical stage. PMID:26270649

  19. MHC class I expression in HPV 16 positive cervical carcinomas is post-transcriptionally controlled and independent from c-myc overexpression.

    PubMed

    Cromme, F V; Snijders, P J; van den Brule, A J; Kenemans, P; Meijer, C J; Walboomers, J M

    1993-11-01

    Squamous cell carcinomas of the uterine cervix (n = 23) were selected for the presence of human papillomavirus type 16 (HPV 16) using the polymerase chain reaction (PCR). Localization of transcripts coding for the E7 protein was demonstrated in neoplastic cells with RNA in situ hybridization. Consecutive tissue sections were investigated for expression of the major histocompatibility complex class I (MHC-I) and c-myc using immunohistochemical double staining procedures, since a role has been suggested for the c-myc protein in MHC-I down-regulation and c-myc overexpression has been described in cervical carcinomas. Reduced expression of class I heavy chains was observed in neoplastic cells from 18 out of 23 carcinomas (78%). Varying levels of c-myc overexpression were observed in 12 carcinomas (52%), from which four showed positive MHC-I expression in c-myc overexpressing cells. In the remaining eight c-myc overexpressing carcinomas MHC-I down-regulation was observed. Additional RNA in situ hybridization with class I heavy chain locus-specific RNA-probes revealed presence of class I mRNAs in those neoplastic cells that show negative staining for MHC-I protein. These data strongly indicate that MHC-I down-regulation in cervical carcinomas involves post-transcriptional mechanisms, not directly related to E7 transcription and overexpression of c-myc. PMID:8414499

  20. In vitro analysis of a primary, major histocompatibility complex (MHC)-restricted, cytotoxic T-lymphocyte response to avian leukosis virus (ALV), using target cells expressing MHC class I cDNA inserted into a recombinant ALV vector.

    PubMed

    Thacker, E L; Fulton, J E; Hunt, H D

    1995-10-01

    The interaction between the major histocompatibility complex (MHC) and cytotoxic T lymphocytes (CTLs) is an important component of the host's resistance to viral infections and tumor formation. In this study, an avian leukosis virus (ALV) vector system, RCASBP, expressing MHC chicken class I (B-F) cDNA was used to develop target cells expressing the chicken class I glycoproteins complexed with ALV antigens on the cell surface. Peripheral blood from chickens inoculated with ALV was shown to contain antigen-specific, MHC-restricted, CD8+ effector CTLs, using a 51Cr release assay utilizing the RCASBP B-F target cells. The stimulated effector cells were also predominantly alpha beta T-cell receptor-positive (TCR2) T cells. The CTL response varied between two haplotypes of chickens which differed in their response to Rous sarcoma virus (RSV)-induced tumors. Chickens with the B21 haplotype which regress RSV-induced tumors showed maximal cytolytic activity, while chickens with the B13 haplotype which do not regress RSV-induced tumors had minimal to no cytolytic activity. In addition to assessing the CTL response to ALV, the creation of MHC-specific immortal target cell lines will be extremely useful in evaluating CTL responses to other viral disease in chickens. PMID:7666545

  1. MHC and Evolution in Teleosts.

    PubMed

    Grimholt, Unni

    2016-01-01

    Major histocompatibility complex (MHC) molecules are key players in initiating immune responses towards invading pathogens. Both MHC class I and class II genes are present in teleosts, and, using phylogenetic clustering, sequences from both classes have been classified into various lineages. The polymorphic and classical MHC class I and class II gene sequences belong to the U and A lineages, respectively. The remaining class I and class II lineages contain nonclassical gene sequences that, despite their non-orthologous nature, may still hold functions similar to their mammalian nonclassical counterparts. However, the fact that several of these nonclassical lineages are only present in some teleost species is puzzling and questions their functional importance. The number of genes within each lineage greatly varies between teleost species. At least some gene expansions seem reasonable, such as the huge MHC class I expansion in Atlantic cod that most likely compensates for the lack of MHC class II and CD4. The evolutionary trigger for similar MHC class I expansions in tilapia, for example, which has a functional MHC class II, is not so apparent. Future studies will provide us with a more detailed understanding in particular of nonclassical MHC gene functions. PMID:26797646

  2. MHC and Evolution in Teleosts

    PubMed Central

    Grimholt, Unni

    2016-01-01

    Major histocompatibility complex (MHC) molecules are key players in initiating immune responses towards invading pathogens. Both MHC class I and class II genes are present in teleosts, and, using phylogenetic clustering, sequences from both classes have been classified into various lineages. The polymorphic and classical MHC class I and class II gene sequences belong to the U and A lineages, respectively. The remaining class I and class II lineages contain nonclassical gene sequences that, despite their non-orthologous nature, may still hold functions similar to their mammalian nonclassical counterparts. However, the fact that several of these nonclassical lineages are only present in some teleost species is puzzling and questions their functional importance. The number of genes within each lineage greatly varies between teleost species. At least some gene expansions seem reasonable, such as the huge MHC class I expansion in Atlantic cod that most likely compensates for the lack of MHC class II and CD4. The evolutionary trigger for similar MHC class I expansions in tilapia, for example, which has a functional MHC class II, is not so apparent. Future studies will provide us with a more detailed understanding in particular of nonclassical MHC gene functions. PMID:26797646

  3. MHC Class II Antigen Presentation by Dendritic Cells Regulated through Endosomal Sorting

    PubMed Central

    ten Broeke, Toine; Wubbolts, Richard; Stoorvogel, Willem

    2013-01-01

    For the initiation of adaptive immune responses, dendritic cells present antigenic peptides in association with major histocompatibility complex class II (MHCII) to naïve CD4+ T lymphocytes. In this review, we discuss how antigen presentation is regulated through intracellular processing and trafficking of MHCII. Newly synthesized MHCII is chaperoned by the invariant chain to endosomes, where peptides from endocytosed pathogens can bind. In nonactivated dendritic cells, peptide-loaded MHCII is ubiquitinated and consequently sorted by the ESCRT machinery to intraluminal vesicles of multivesicular bodies, ultimately leading to lysosomal degradation. Ubiquitination of newly synthesized MHCII is blocked when dendritic cells are activated, now allowing its transfer to the cell surface. This mode of regulation for MHCII is a prime example of how molecular processing and sorting at multivesicular bodies can determine the expression of signaling receptors at the plasma membrane. PMID:24296169

  4. Structurally Defined αMHC-II Nanobody-Drug Conjugates: A Therapeutic and Imaging System for B-Cell Lymphoma.

    PubMed

    Fang, Tao; Duarte, Joao N; Ling, Jingjing; Li, Zeyang; Guzman, Jonathan S; Ploegh, Hidde L

    2016-02-12

    Antibody-drug conjugates (ADCs) of defined structure hold great promise for cancer therapies, but further advances are constrained by the complex structures of full-sized antibodies. Camelid-derived single-domain antibody fragments (VHHs or nanobodies) offer a possible solution to this challenge by providing expedited target screening and validation through switching between imaging and therapeutic activities. We used a nanobody (VHH7) specific for murine MHC-II and rendered "sortase-ready" for the introduction of oligoglycine-modified cytotoxic payloads or NIR fluorophores. The VHH7 conjugates outcompeted commercial monoclonal antibodies (mAbs) for internalization and exhibited high specificity and cytotoxicity against A20 murine B-cell lymphoma. Non-invasive NIR imaging with a VHH7-fluorophore conjugate showed rapid tumor targeting on both localized and metastatic lymphoma models. Subsequent treatment with the nanobody-drug conjugate efficiently controlled tumor growth and metastasis without obvious systemic toxicity. PMID:26840214

  5. Btn2a2, a T cell immunomodulatory molecule coregulated with MHC class II genes

    PubMed Central

    Sarter, Kerstin; Leimgruber, Elisa; Gobet, Florian; Agrawal, Vishal; Dunand-Sauthier, Isabelle; Barras, Emmanuèle; Mastelic-Gavillet, Béatris; Kamath, Arun; Fontannaz, Paola; Guéry, Leslie; Duraes, Fernanda do Valle; Lippens, Carla; Ravn, Ulla; Santiago-Raber, Marie-Laure; Magistrelli, Giovanni; Fischer, Nicolas; Siegrist, Claire-Anne; Hugues, Stéphanie

    2016-01-01

    Evidence has recently emerged that butyrophilins, which are members of the extended B7 family of co-stimulatory molecules, have diverse functions in the immune system. We found that the human and mouse genes encoding butyrophilin-2A2 (BTN2A2) are regulated by the class II trans-activator and regulatory factor X, two transcription factors dedicated to major histocompatibility complex class II expression, suggesting a role in T cell immunity. To address this, we generated Btn2a2-deficient mice. Btn2a2−/− mice exhibited enhanced effector CD4+ and CD8+ T cell responses, impaired CD4+ regulatory T cell induction, potentiated antitumor responses, and exacerbated experimental autoimmune encephalomyelitis. Altered immune responses were attributed to Btn2a2 deficiency in antigen-presenting cells rather than T cells or nonhematopoietic cells. These results provide the first genetic evidence that BTN2A2 is a co-inhibitory molecule that modulates T cell–mediated immunity. PMID:26809444

  6. Low immunogenicity of in vitro-expanded human neural cells despite high MHC expression.

    PubMed

    Odeberg, Jenny; Piao, Jing-Hua; Samuelsson, Eva-Britt; Falci, Scott; Akesson, Elisabet

    2005-04-01

    The ability to expand human neural precursor cells in vitro offers new possibilities for future cell therapies. However, concern over immunologically based rejection of in vitro-expanded human neural cells confounds their use as donor cells. Here, we demonstrate that the expression of human leukocyte antigen (HLA) class I and II molecules, but not the co-stimulatory proteins CD40, CD80 and CD86, substantially increase during expansion of neurospheres. Furthermore, peripheral lymphocytes were unresponsive when co-cultured with in vitro-expanded neural cells. Taken together, these results suggest a low immunogenicity of these cultured human neural cells despite HLA incompatibility and high HLA expression. PMID:15748938

  7. The role of polymorphic amino acids of the MHC molecule in the selection of the T cell repertoire

    SciTech Connect

    Bhayani, H.R.; Hedrick, S.M. )

    1991-02-15

    Allelic variants of MHC molecules expressed on cells of the thymus affect the selection and the specificity of the T cell repertoire. The selection is based on either the direct recognition by the TCR of the MHC molecules, or the recognition of a complex determinant formed by self-peptides bound to MHC molecules. In an analysis of the T cell repertoire in bone marrow chimeras that express allelic forms of MHC class II molecules in the thymus epithelium, we find that amino acid substitutions that are predicted to affect peptide binding influence the selection of the T cell repertoire during thymic selection.

  8. Trans-species polymorphism of the Mhc class II DRB-like gene in banded penguins (genus Spheniscus).

    PubMed

    Kikkawa, Eri F; Tsuda, Tomi T; Sumiyama, Daisuke; Naruse, Taeko K; Fukuda, Michio; Kurita, Masanori; Wilson, Rory P; LeMaho, Yvon; Miller, Gary D; Tsuda, Michio; Murata, Koichi; Kulski, Jerzy K; Inoko, Hidetoshi

    2009-05-01

    The Major Histocompatibility Complex (Mhc) class II DRB locus of vertebrates is highly polymorphic and some alleles may be shared between closely related species as a result of balancing selection in association with resistance to parasites. In this study, we developed a new set of PCR primers to amplify, clone, and sequence overlapping portions of the Mhc class II DRB-like gene from the 5'UTR end to intron 3, including exons 1, 2, and 3 and introns 1 and 2 in four species (20 Humboldt, six African, five Magellanic, and three Galapagos penguins) of penguin from the genus Spheniscus (Sphe). Analysis of gene sequence variation by the neighbor-joining method of 21 Sphe sequences and 20 previously published sequences from four other penguin species revealed overlapping clades within the Sphe species, but species-specific clades for the other penguin species. The overlap of the DRB-like gene sequence variants between the four Sphe species suggests that, despite their allopatric distribution, the Sphe species are closely related and that some shared DRB1 alleles may have undergone a trans-species inheritance because of balancing selection and/or recent rapid speciation. The new primers and PCR assays that we have developed for the identification of the DRB1 DNA and protein sequence variations appear to be useful for the characterization of the molecular evolution of the gene in closely related Penguin species and might be helpful for the assessment of the genetic health and the management of the conservation and captivity of these endangered species. PMID:19319519

  9. Cheetah paradigm revisited: MHC diversity in the world's largest free-ranging population.

    PubMed

    Castro-Prieto, Aines; Wachter, Bettina; Sommer, Simone

    2011-04-01

    For more than two decades, the cheetah (Acinonyx jubatus) has been considered a paradigm of disease vulnerability associated with low genetic diversity, particularly at the immune genes of the major histocompatibility complex (MHC). Cheetahs have been used as a classic example in numerous conservation genetics textbooks as well as in many related scientific publications. However, earlier studies used methods with low resolution to quantify MHC diversity and/or small sample sizes. Furthermore, high disease susceptibility was reported only for captive cheetahs, whereas free-ranging cheetahs show no signs of infectious diseases and a good general health status. We examined whether the diversity at MHC class I and class II-DRB loci in 149 Namibian cheetahs was higher than previously reported using single-strand conformation polymorphism analysis, cloning, and sequencing. MHC genes were examined at the genomic and transcriptomic levels. We detected ten MHC class I and four class II-DRB alleles, of which nine MHC class I and all class II-DRB alleles were expressed. Phylogenetic analyses and individual genotypes suggested that the alleles belong to four MHC class I and three class II-DRB putative loci. Evidence of positive selection was detected in both MHC loci. Our study indicated that the low number of MHC class I alleles previously observed in cheetahs was due to a smaller sample size examined. On the other hand, the low number of MHC class II-DRB alleles previously observed in cheetahs was further confirmed. Compared with other mammalian species including felids, cheetahs showed low levels of MHC diversity, but this does not seem to influence the immunocompetence of free-ranging cheetahs in Namibia and contradicts the previous conclusion that the cheetah is a paradigm species of disease vulnerability. PMID:21183613

  10. MHC class II transactivator represses human IL-4 gene transcription by interruption of promoter binding with CBP/p300, STAT6 and NFAT1 via histone hypoacetylation

    PubMed Central

    Zhou, Xiaorong; Jiang, Yang; Lu, Liming; Ding, Qing; Jiao, Zhijun; Zhou, Yun; Xin, Lijun; Chou, Kuang-Yen

    2007-01-01

    In addition to its property of enhancing major histocompatibility complex (MHC) class II expression, the class II transactivator (CIITA) was recently demonstrated to be involved in T helper type 1/type 2 (Th1/Th2) differentiation by regulating interleukin-4 (IL-4) gene transcription. There was however, controversy regarding whether CIITA promotes or suppresses IL-4 expression in the experiments with transgenic mice. To clarify the discrepancy by using simpler experimental systems, human Jurkat T cells that express IL-4 but not interferon-γ, even if stimulated with phorbol 12-myristate 13-acetate plus ionomycin, were used for CIITA transfection. Significant suppression of IL-4 gene expression was demonstrated. Simultaneously, histones H3 and H4 in the IL-4 promoter were hypoacetylated. The suppression could be totally reversed by the histone deacetylatase inhibitor trichostatin A. Furthermore, the IL-4 expression was determined in primarily established human Th1/Th2 cells to which CIITA small interference RNA (siRNA) had been introduced. A substantially increased level of IL-4 was recorded in the CIITA siRNA-transfected Th1 cells, which was in parallel with significantly enhanced acetylation in histone H3 of the IL-4 promoter. Chromatin immunoprecipitation analysis indicated that CIITA abrogated the binding of coactivator CBP/p300 and transcription factors STAT6/NFAT1 to IL-4 promoter in the CIITA-transfected cells. In conclusion, CIITA was active in the repression of transcription activation of human IL-4 gene in both the T-cell line and the primary human CD4 T cells by preventing transcription factors from binding to IL-4 promoter through histone hypoacetylation. Our data confirm a potential significant role of CIITA in controlling Th1/Th2 differentiation via modulation of IL-4 gene activation. PMID:17645498

  11. HIV-Infected Dendritic Cells Present Endogenous MHC Class II-Restricted Antigens to HIV-Specific CD4+ T Cells.

    PubMed

    Coulon, Pierre-Grégoire; Richetta, Clémence; Rouers, Angéline; Blanchet, Fabien P; Urrutia, Alejandra; Guerbois, Mathilde; Piguet, Vincent; Theodorou, Ioannis; Bet, Anne; Schwartz, Olivier; Tangy, Frédéric; Graff-Dubois, Stéphanie; Cardinaud, Sylvain; Moris, Arnaud

    2016-07-15

    It is widely assumed that CD4(+) T cells recognize antigenic peptides (epitopes) derived solely from incoming, exogenous, viral particles or proteins. However, alternative sources of MHC class II (MHC-II)-restricted Ags have been described, in particular epitopes derived from newly synthesized proteins (so-called endogenous). In this study, we show that HIV-infected dendritic cells (DC) present MHC-II-restricted endogenous viral Ags to HIV-specific (HS) CD4(+) T cells. This endogenous pathway functions independently of the exogenous route for HIV Ag presentation and offers a distinct possibility for the immune system to activate HS CD4(+) T cells. We examined the implication of autophagy, which plays a crucial role in endogenous viral Ag presentation and thymic selection of CD4(+) T cells, in HIV endogenous presentation. We show that infected DC do not use autophagy to process MHC-II-restricted HIV Ags. This is unlikely to correspond to a viral escape from autophagic degradation, as infecting DC with Nef- or Env-deficient HIV strains did not impact HS T cell activation. However, we demonstrate that, in DC, specific targeting of HIV Ags to autophagosomes using a microtubule-associated protein L chain 3 (LC3) fusion protein effectively enhances and broadens HS CD4(+) T cell responses, thus favoring an endogenous MHC-II-restricted presentation. In summary, in DC, multiple endogenous presentation pathways lead to the activation of HS CD4(+) T cell responses. These findings will help in designing novel strategies to activate HS CD4(+) T cells that are required for CTL activation/maintenance and B cell maturation. PMID:27288536

  12. Fish oil disrupts MHC class II lateral organization on the B-cell side of the immunological synapse independent of B-T cell adhesion.

    PubMed

    Rockett, Benjamin Drew; Melton, Mark; Harris, Mitchel; Bridges, Lance C; Shaikh, Saame Raza

    2013-11-01

    Fish oil-enriched long chain n-3 polyunsaturated fatty acids disrupt the molecular organization of T-cell proteins in the immunological synapse. The impact of fish oil derived n-3 fatty acids on antigen-presenting cells, particularly at the animal level, is unknown. We previously demonstrated B-cells isolated from mice fed with fish oil-suppressed naïve CD4(+) T-cell activation. Therefore, here we determined the mechanistic effects of fish oil on murine B-cell major histocompatibility complex (MHC) class II molecular distribution using a combination of total internal reflection fluorescence, Förster resonance energy transfer and confocal imaging. Fish oil had no impact on presynaptic B-cell MHC II clustering. Upon conjugation with transgenic T-cells, fish-oil suppressed MHC II accumulation at the immunological synapse. As a consequence, T-cell protein kinase C theta (PKCθ) recruitment to the synapse was also diminished. The effects were independent of changes in B-T cell adhesion, as measured with microscopy, flow cytometry and static cell adhesion assays with select immune ligands. Given that fish oil can reorganize the membrane by lowering membrane cholesterol levels, we then compared the results with fish oil to cholesterol depletion using methyl-B-cyclodextrin (MβCD). MβCD treatment of B-cells suppressed MHC II and T-cell PKCθ recruitment to the immunological synapse, similar to fish oil. Overall, the results reveal commonality in the mechanism by which fish oil manipulates protein lateral organization of B-cells compared to T-cells. Furthermore, the data establish MHC class II lateral organization on the B-cell side of the immunological synapse as a novel molecular target of fish oil. PMID:23791516

  13. Selective pressures on MHC class II genes in the guppy (Poecilia reticulata) as inferred by hierarchical analysis of population structure.

    PubMed

    Herdegen, M; Babik, W; Radwan, J

    2014-11-01

    Genes of the major histocompatibility complex, which are the most polymorphic of all vertebrate genes, are a pre-eminent system for the study of selective pressures that arise from host-pathogen interactions. Balancing selection capable of maintaining high polymorphism should lead to the homogenization of MHC allele frequencies among populations, but there is some evidence to suggest that diversifying selection also operates on the MHC. However, the pattern of population structure observed at MHC loci is likely to depend on the spatial and/or temporal scale examined. Here, we investigated selection acting on MHC genes at different geographic scales using Venezuelan guppy populations inhabiting four regions. We found a significant correlation between MHC and microsatellite allelic richness across populations, which suggests the role of genetic drift in shaping MHC diversity. However, compared to microsatellites, more MHC variation was explained by differences between populations within larger geographic regions and less by the differences between the regions. Furthermore, among proximate populations, variation in MHC allele frequencies was significantly higher compared to microsatellites, indicating that selection acting on MHC may increase population structure at small spatial scales. However, in populations that have significantly diverged at neutral markers, the population-genetic signature of diversifying selection may be eradicated in the long term by that of balancing selection, which acts to preserve rare alleles and thus maintain a common pool of MHC alleles. PMID:25244157

  14. PERB11 (MIC): a polymorphic MHC gene is expressed in skin and single nucleotide polymorphisms are associated with psoriasis

    PubMed Central

    Tay, G K; Hui, J; Gaudieri, S; Schmitt-Egenolf, M; Martinez, O P; Leelayuwat, C; Williamson, J F; Eiermann, T H; Dawkins, R L

    2000-01-01

    The susceptibility genes for psoriasis remain to be identified. At least one of these must be in the major histocompatibility complex (MHC) to explain associations with alleles at human leucocyte antigen (HLA)-A, -B, -C, -DR, -DQ and C4. In fact, most of these alleles are components of just two ancestral haplotypes (AHs) designated 13.1 and 57.1. Although relevant MHC gene(s) could be within a region of at least 4 Mb, most studies have favoured the area near HLA-B and -C. This region contains a large number of non-HLA genes, many of which are duplicated and polymorphic. Members of one such gene family, PERB11.1 and PERB11.2, are expressed in the skin and are encoded in the region between tumour necrosis factor and HLA-B. To investigate the relationship of PERB11.1 alleles to psoriasis, sequence based typing was performed on 97 patients classified according to age of onset and family history. The frequency of the PERB11.1*06 allele is 44% in type I psoriasis but only 7% in controls (Pc = 0.003 by Fisher's exact test, two-tailed). The major determinant of this association is a single nucleotide polymorphism (SNP) within intron 4. In normal and affected skin, expression of PERB11 is mainly in the basal layer of the epidermis including ducts and follicles. PERB11 is also present in the upper keratin layers but there is relative deficiency in the intermediate layers. These findings suggest a possible role for PERB11 and other MHC genes in the pathogenesis of psoriasis. PMID:10691930

  15. Regulation of major histocompatibility complex class II genes

    PubMed Central

    Choi, Nancy M.; Majumder, Parimal; Boss, Jeremy M.

    2010-01-01

    Summary The major histocompatibility complex class II (MHC-II) genes are regulated at the level of transcription. Recent studies have shown that chromatin modification is critical for efficient transcription of these genes, and a number of chromatin modifying complexes recruited to MHC-II genes have been described. The MHC-II genes are segregated from each other by a series of chromatin elements, termed MHC-II insulators. Interactions between MHC-insulators and the promoters of MHC-II genes are mediated by the insulator factor CCCTC-binding protein and are critical for efficient expression. This regulatory mechanism provides a novel view of how the entire MHC-II locus is assembled architecturally and can be coordinately controlled. PMID:20970972

  16. Transport of Streptococcus pneumoniae Capsular Polysaccharide in MHC Class II Tubules

    PubMed Central

    Stephen, Tom Li; Fabri, Mario; Groneck, Laura; Röhn, Till A; Hafke, Helena; Robinson, Nirmal; Rietdorf, Jens; Schrama, David; Becker, Jürgen C; Plum, Georg; Krönke, Martin; Kropshofer, Harald; Kalka-Moll, Wiltrud M

    2007-01-01

    Bacterial capsular polysaccharides are virulence factors and are considered T cell–independent antigens. However, the capsular polysaccharide Sp1 from Streptococcus pneumoniae serotype 1 has been shown to activate CD4+ T cells in a major histocompatibility complex (MHC) class II–dependent manner. The mechanism of carbohydrate presentation to CD4+ T cells is unknown. We show in live murine dendritic cells (DCs) that Sp1 translocates from lysosomal compartments to the plasma membrane in MHCII-positive tubules. Sp1 cell surface presentation results in reduction of self-peptide presentation without alteration of the MHCII self peptide repertoire. In DM-deficient mice, retrograde transport of Sp1/MHCII complexes resulting in T cell–dependent immune responses to the polysaccharide in vitro and in vivo is significantly reduced. The results demonstrate the capacity of a bacterial capsular polysaccharide antigen to use DC tubules as a vehicle for its transport as an MHCII/saccharide complex to the cell surface for the induction of T cell activation. Furthermore, retrograde transport requires the functional role of DM in self peptide–carbohydrate exchange. These observations open new opportunities for the design of vaccines against microbial encapsulated pathogens. PMID:17367207

  17. Domain structures and molecular evolution of class I and class II major histocompatibility gene complex (MHC) products deduced from amino acid and nucleotide sequence homologies

    NASA Astrophysics Data System (ADS)

    Ohnishi, Koji

    1984-12-01

    Domain structures of class I and class II MHC products were analyzed from a viewpoint of amino acid and nucleotide sequence homologies. Alignment statistics revealed that class I (transplantation) antigen H chains consist of four mutually homologous domains, and that class II (HLA-DR) antigen β and α chains are both composed of three mutually homologous ones. The N-terminal three and two domains of class I and class II (both β and α) gene products, respectively, all of which being ˜90 residues long, were concluded to be homologous to β2-microglobulin (β2M). The membraneembedded C-terminal shorter domains of these MHC products were also found to be homologous to one another and to the third domain of class I H chains. Class I H chains were found to be more closely related to class II α chains than to class II β chains. Based on these findings, an exon duplication history from a common ancestral gene encoding a β2M-like primodial protein of one-domain-length up to the contemporary MHC products was proposed.

  18. Immunization with a Peptide Containing MHC Class I and II Epitopes Derived from the Tumor Antigen SIM2 Induces an Effective CD4 and CD8 T-Cell Response

    PubMed Central

    Kissick, Haydn T.; Sanda, Martin G.; Dunn, Laura K.; Arredouani, Mohamed S.

    2014-01-01

    Here, we sought to determine whether peptide vaccines designed harbor both class I as well as class II restricted antigenic motifs could concurrently induce CD4 and CD8 T cell activation against autologous tumor antigens. Based on our prior genome-wide interrogation of human prostate cancer tissues to identify genes over-expressed in cancer and absent in the periphery, we targeted SIM2 as a prototype autologous tumor antigen for these studies. Using humanized transgenic mice we found that the 9aa HLA-A*0201 epitope, SIM2237–245, was effective at inducing an antigen specific response against SIM2-expressing prostate cancer cell line, PC3. Immunization with a multi-epitope peptide harboring both MHC-I and MHC-II restricted epitopes induced an IFN-γ response in CD8 T cells to the HLA-A*0201-restricted SIM2237–245 epitope, and an IL-2 response by CD4 T cells to the SIM2240–254 epitope. This peptide was also effective at inducing CD8+ T-cells that responded specifically to SIM2-expressing tumor cells. Collectively, the data presented in this study suggest that a single peptide containing multiple SIM2 epitopes can be used to induce both a CD4 and CD8 T cell response, providing a peptide-based vaccine formulation for potential use in immunotherapy of various cancers. PMID:24690990

  19. Immunization with a peptide containing MHC class I and II epitopes derived from the tumor antigen SIM2 induces an effective CD4 and CD8 T-cell response.

    PubMed

    Kissick, Haydn T; Sanda, Martin G; Dunn, Laura K; Arredouani, Mohamed S

    2014-01-01

    Here, we sought to determine whether peptide vaccines designed harbor both class I as well as class II restricted antigenic motifs could concurrently induce CD4 and CD8 T cell activation against autologous tumor antigens. Based on our prior genome-wide interrogation of human prostate cancer tissues to identify genes over-expressed in cancer and absent in the periphery, we targeted SIM2 as a prototype autologous tumor antigen for these studies. Using humanized transgenic mice we found that the 9aa HLA-A*0201 epitope, SIM2(237-245), was effective at inducing an antigen specific response against SIM2-expressing prostate cancer cell line, PC3. Immunization with a multi-epitope peptide harboring both MHC-I and MHC-II restricted epitopes induced an IFN-γ response in CD8 T cells to the HLA-A*0201-restricted SIM2(237-245) epitope, and an IL-2 response by CD4 T cells to the SIM2(240-254) epitope. This peptide was also effective at inducing CD8+ T-cells that responded specifically to SIM2-expressing tumor cells. Collectively, the data presented in this study suggest that a single peptide containing multiple SIM2 epitopes can be used to induce both a CD4 and CD8 T cell response, providing a peptide-based vaccine formulation for potential use in immunotherapy of various cancers. PMID:24690990

  20. Immunologic Hierarchy, Class II MHC Promiscuity, and Epitope Spreading of a Melanoma Helper Peptide Vaccine

    PubMed Central

    Hu, Yinin; Petroni, Gina R.; Olson, Walter C.; Czarkowski, Andrea; Smolkin, Mark E.; Grosh, William W.; Chianese-Bullock, Kimberly A.; Slingluff, Craig L.

    2014-01-01

    Immunization with a combination melanoma helper peptide (6MHP) vaccine has been shown to induce CD4+ T-cell responses, which are associated with patient survival. In the present study, we define the relative immunogenicity and HLA allele promiscuity of individual helper peptides, and identify helper peptide-mediated augmentation of specific CD8+ T-cell responses. Thirty-seven participants with stage IIIB-IV melanoma were vaccinated with 6MHP in incomplete Freund’s adjuvant. The 6MHP vaccine is comprised of 6 peptides representing melanocytic differentiation proteins gp100, tyrosinase, Melan-A/MART-1 and cancer-testis antigens from the MAGE family. CD4+ and CD8+ T cell responses were assessed in peripheral blood and in sentinel immunized nodes (SIN) by thymidine uptake after exposure to helper peptides and by direct interferon-γ ELI spot assay against 14 MHC class I-restricted peptides. Vaccine-induced CD4+ T cell responses to individual epitopes were detected in the SIN of 63% (22/35) and in the peripheral blood of 38% (14/37) of participants for an overall response rate of 65% (24/37). The most frequently immunogenic peptides were MAGE-A3281-295 (49%) and tyrosinase 386-406 (32%). Responses were not limited to HLA restrictions originally described. Vaccine-associated CD8+ T-cell responses against class I-restricted peptides were observed in 45% (5/11) of evaluable participants. The 6MHP vaccine induces both CD4+ and CD8 + T cell responses against melanoma antigens. CD4+ T-cell responses were detected beyond reported HLA-DR restrictions. Induction of CD8+ T-cell responses suggests epitope spreading and systemic activity mediated at the tumor site. PMID:24756419

  1. Activated rat T cells synthesize and express functional major histocompatibility class II antigens.

    PubMed Central

    Broeren, C P; Wauben, M H; Lucassen, M A; Van Meurs, M; Van Kooten, P J; Boog, C J; Claassen, E; Van Eden, W

    1995-01-01

    In the present report, we studied the presence and functional significance of major histocompatibility complex (MHC) class II antigen on rat T cells. Most rat T-cell lines cultured in vitro were found to be MHC class II+. Also, these T-cell lines were shown to synthesize MHC class II molecules. Immunohistochemical and flow cytometric double stainings for T-cell receptor (TCR) and MHC class II showed that in vivo as well a large proportion of T cells was MHC class II+. The immunohistochemical staining of spleen sections enabled us to characterize the MHC class II+ and MHC class II- T cells. It was shown that resting T cells in vivo were MHC class II-. In contrast, activated T cells, as determined by their localization in the marginal zone of the spleen, proved to be MHC class II+. Finally, T-cell clones were found to be able to present peptidic antigens, but could only poorly present more complex exogenous antigens, probably due to inefficient uptake of such antigens. These features would endow activated rat T cells with the capacity to present cell-specific self-proteins, such as TCR, to regulatory CD4+ MHC class II-restricted T cells, as was described by our group elsewhere. Images Figure 2 Figure 5 PMID:7750994

  2. Interaction Analysis between HLA-DRB1 Shared Epitope Alleles and MHC Class II Transactivator CIITA Gene with Regard to Risk of Rheumatoid Arthritis

    PubMed Central

    Ronninger, Marcus; Seddighzadeh, Maria; Eike, Morten Christoph; Plant, Darren; Daha, Nina A.; Skinningsrud, Beate; Worthington, Jane; Kvien, Tore K.; Toes, Rene E. M.; Lie, Benedicte A.; Alfredsson, Lars; Padyukov, Leonid

    2012-01-01

    HLA-DRB1 shared epitope (SE) alleles are the strongest genetic determinants for autoantibody positive rheumatoid arthritis (RA). One of the key regulators in expression of HLA class II receptors is MHC class II transactivator (CIITA). A variant of the CIITA gene has been found to associate with inflammatory diseases. We wanted to explore whether the risk variant rs3087456 in the CIITA gene interacts with the HLA-DRB1 SE alleles regarding the risk of developing RA. We tested this hypothesis in a case-control study with 11767 individuals from four European Caucasian populations (6649 RA cases and 5118 controls). We found no significant additive interaction for risk alleles among Swedish Caucasians with RA (n = 3869, attributable proportion due to interaction (AP) = 0.2, 95%CI: −0.2–0.5) or when stratifying for anti-citrullinated protein antibodies (ACPA) presence (ACPA positive disease: n = 2945, AP = 0.3, 95%CI: −0.05–0.6, ACPA negative: n = 2268, AP = −0.2, 95%CI: −1.0–0.6). We further found no significant interaction between the main subgroups of SE alleles (DRB1*01, DRB1*04 or DRB1*10) and CIITA. Similar analysis of three independent RA cohorts from British, Dutch and Norwegian populations also indicated an absence of significant interaction between genetic variants in CIITA and SE alleles with regard to RA risk. Our data suggest that risk from the CIITA locus is independent of the major risk for RA from HLA-DRB1 SE alleles, given that no significant interaction between rs3087456 and SE alleles was observed. Since a biological link between products of these genes is evident, the genetic contribution from CIITA and class II antigens in the autoimmune process may involve additional unidentified factors. PMID:22461888

  3. Genetic variation of the MHC class II DRB genes in the Japanese weasel, Mustela itatsi, endemic to Japan, compared with the Siberian weasel, Mustela sibirica.

    PubMed

    Nishita, Y; Abramov, A V; Kosintsev, P A; Lin, L-K; Watanabe, S; Yamazaki, K; Kaneko, Y; Masuda, R

    2015-12-01

    Major histocompatibility complex (MHC) genes encode proteins that play a critical role in vertebrate immune system and are highly polymorphic. To further understand the molecular evolution of the MHC genes, we compared MHC class II DRB genes between the Japanese weasel (Mustela itatsi), a species endemic to Japan, and the Siberian weasel (Mustela sibirica), a closely related species on the continent. We sequenced a 242-bp region of DRB exon 2, which encodes antigen-binding sites (ABS), and found 24 alleles from 31 M. itatsi individuals and 17 alleles from 21 M. sibirica individuals, including broadly distributed, species-specific and/or geographically restricted alleles. Our results suggest that pathogen-driven balancing selection have acted to maintain the diversity in the DRB genes. For predicted ABS, nonsynonymous substitutions exceeded synonymous substitutions, also indicating positive selection, which was not seen at non-ABS. In a Bayesian phylogenetic tree, two M. sibirica DRB alleles were basal to the rest of the sequences from mustelid species and may represent ancestral alleles. Trans-species polymorphism was evident between many mustelid DRB alleles, especially between M. itatsi and M. sibirica. These two Mustela species divided about 1.7 million years ago, but still share many MHC alleles, indicative of their close phylogenetic relationship. PMID:26593752

  4. Genetic Variation at Exon 2 of the MHC Class II DQB Locus in Blue Whale (Balaenoptera musculus) from the Gulf of California.

    PubMed

    Moreno-Santillán, Diana D; Lacey, Eileen A; Gendron, Diane; Ortega, Jorge

    2016-01-01

    The genes of the Major Histocompatibility Complex (MHC) play an important role in the vertebrate immune response and are among the most polymorphic genes known in vertebrates. In some marine mammals, MHC genes have been shown to be characterized by low levels of polymorphism compared to terrestrial taxa; this reduction in variation is often explained as a result of lower pathogen pressures in marine habitats. To determine if this same reduction in variation applies to the migratory population of blue whales (Balaenoptera musculus) that occurs in the Gulf of California, we genotyped a 172 bp fragment of exon 2 of the MHC Class II DQB locus for 80 members of this population. Twenty-two putatively functional DQB allotypes were identified, all of which were homologous with DQB sequences from other cetacean species. Up to 5 putative alleles per individual were identified, suggesting that gene duplication has occurred at this locus. Rates of non-synonymous to synonymous substitutions (ω) and maximum likelihood analyses of models of nucleotide variation provided potential evidence of ongoing positive selection at this exon. Phylogenetic analyses of DQB alleles from B. musculus and 16 other species of cetaceans revealed trans-specific conservation of MHC variants, suggesting that selection has acted on this locus over prolonged periods of time. Collectively our findings reveal that immunogenic variation in blue whales is comparable to that in terrestrial mammals, thereby providing no evidence that marine taxa are subject to reduced pathogen-induced selective pressures. PMID:26761201

  5. Genetic Variation at Exon 2 of the MHC Class II DQB Locus in Blue Whale (Balaenoptera musculus) from the Gulf of California

    PubMed Central

    Moreno-Santillán, Diana D.; Lacey, Eileen A.; Gendron, Diane; Ortega, Jorge

    2016-01-01

    The genes of the Major Histocompatibility Complex (MHC) play an important role in the vertebrate immune response and are among the most polymorphic genes known in vertebrates. In some marine mammals, MHC genes have been shown to be characterized by low levels of polymorphism compared to terrestrial taxa; this reduction in variation is often explained as a result of lower pathogen pressures in marine habitats. To determine if this same reduction in variation applies to the migratory population of blue whales (Balaenoptera musculus) that occurs in the Gulf of California, we genotyped a 172 bp fragment of exon 2 of the MHC Class II DQB locus for 80 members of this population. Twenty-two putatively functional DQB allotypes were identified, all of which were homologous with DQB sequences from other cetacean species. Up to 5 putative alleles per individual were identified, suggesting that gene duplication has occurred at this locus. Rates of non-synonymous to synonymous substitutions (ω) and maximum likelihood analyses of models of nucleotide variation provided potential evidence of ongoing positive selection at this exon. Phylogenetic analyses of DQB alleles from B. musculus and 16 other species of cetaceans revealed trans-specific conservation of MHC variants, suggesting that selection has acted on this locus over prolonged periods of time. Collectively our findings reveal that immunogenic variation in blue whales is comparable to that in terrestrial mammals, thereby providing no evidence that marine taxa are subject to reduced pathogen-induced selective pressures. PMID:26761201

  6. Expression, refolding and crystallization of murine MHC class I H-2D{sup b} in complex with human β{sub 2}-microglobulin

    SciTech Connect

    Sandalova, Tatyana; Michaëlsson, Jakob; Harris, Robert A.; Ljunggren, Hans-Gustaf; Kärre, Klas; Schneider, Gunter; Achour, Adnane

    2005-12-01

    Mouse MHC class I H-2Db in complex with human β2m and the LCMV-derived peptide gp33 has been produced and crystallized. Resolution of the structure of this complex combined with the structural comparison with the previously solved crystal structure of H-2Db/mβ2m/gp33 should lead to a better understanding of how the β2m subunit affects the overall conformation of MHC complexes as well as the stability of the presented peptides. β{sub 2}-Microglobulin (β{sub 2}m) is non-covalently linked to the major histocompatibility (MHC) class I heavy chain and interacts with CD8 and Ly49 receptors. Murine MHC class I can bind human β{sub 2}m (hβ{sub 2}m) and such hybrid molecules are often used in structural and functional studies. The replacement of mouse β{sub 2}m (mβ{sub 2}m) by hβ{sub 2}m has important functional consequences for MHC class I complex stability and specificity, but the structural basis for this is unknown. To investigate the impact of species-specific β{sub 2}m subunits on MHC class I conformation, murine MHC class I H-2D{sup b} in complex with hβ{sub 2}m and the peptide gp33 derived from lymphocytic choriomeningitis virus (LCMV) has been expressed, refolded in vitro and crystallized. Crystals containing two complexes per asymmetric unit and belonging to the space group P2{sub 1}, with unit-cell parameters a = 68.1, b = 65.2, c = 101.9 Å, β = 102.4°, were obtained.

  7. Genome-wide association study identifies SNPs in the MHC class II loci that are associated with self-reported history of whooping cough

    PubMed Central

    McMahon, George; Ring, Susan M.; Davey-Smith, George; Timpson, Nicholas J.

    2015-01-01

    Whooping cough is currently seeing resurgence in countries despite high vaccine coverage. There is considerable variation in subject-specific response to infection and vaccine efficacy, but little is known about the role of human genetics. We carried out a case–control genome-wide association study of adult or parent-reported history of whooping cough in two cohorts from the UK: the ALSPAC cohort and the 1958 British Birth Cohort (815/758 cases and 6341/4308 controls, respectively). We also imputed HLA alleles using dense SNP data in the MHC region and carried out gene-based and gene-set tests of association and estimated the amount of additive genetic variation explained by common SNPs. We observed a novel association at SNPs in the MHC class II region in both cohorts [lead SNP rs9271768 after meta-analysis, odds ratio [95% confidence intervals (CIs)] 1.47 (1.35, 1.6), P-value 1.21E − 18]. Multiple strong associations were also observed at alleles at the HLA class II loci. The majority of these associations were explained by the lead SNP rs9271768. Gene-based and gene-set tests and estimates of explainable common genetic variation could not establish the presence of additional associations in our sample. Genetic variation at the MHC class II region plays a role in susceptibility to whooping cough. These findings provide additional perspective on mechanisms of whooping cough infection and vaccine efficacy. PMID:26231221

  8. Genome-wide association study identifies SNPs in the MHC class II loci that are associated with self-reported history of whooping cough.

    PubMed

    McMahon, George; Ring, Susan M; Davey-Smith, George; Timpson, Nicholas J

    2015-10-15

    Whooping cough is currently seeing resurgence in countries despite high vaccine coverage. There is considerable variation in subject-specific response to infection and vaccine efficacy, but little is known about the role of human genetics. We carried out a case-control genome-wide association study of adult or parent-reported history of whooping cough in two cohorts from the UK: the ALSPAC cohort and the 1958 British Birth Cohort (815/758 cases and 6341/4308 controls, respectively). We also imputed HLA alleles using dense SNP data in the MHC region and carried out gene-based and gene-set tests of association and estimated the amount of additive genetic variation explained by common SNPs. We observed a novel association at SNPs in the MHC class II region in both cohorts [lead SNP rs9271768 after meta-analysis, odds ratio [95% confidence intervals (CIs)] 1.47 (1.35, 1.6), P-value 1.21E - 18]. Multiple strong associations were also observed at alleles at the HLA class II loci. The majority of these associations were explained by the lead SNP rs9271768. Gene-based and gene-set tests and estimates of explainable common genetic variation could not establish the presence of additional associations in our sample. Genetic variation at the MHC class II region plays a role in susceptibility to whooping cough. These findings provide additional perspective on mechanisms of whooping cough infection and vaccine efficacy. PMID:26231221

  9. Trypanosoma cruzi Infection Down-Modulates the Immunoproteasome Biosynthesis and the MHC Class I Cell Surface Expression in HeLa Cells

    PubMed Central

    Camargo, Ricardo; Faria, Liliam O.; Kloss, Alexander; Favali, Cecília B. F.; Kuckelkorn, Ulrike; Kloetzel, Peter-Michael; de Sá, Cezar Martins; Lima, Beatriz D.

    2014-01-01

    Generally, Trypanosoma cruzi infection in human is persistent and tends to chronicity, suggesting that the parasite evade the immune surveillance by down regulating the intracellular antigen processing routes. Within the MHC class I pathway, the majority of antigenic peptides are generated by the proteasome. However, upon IFN-γ stimulation, the catalytic constitutive subunits of the proteasome are replaced by the subunits β1i/LMP2, β2i/MECL-1 and β5i/LMP7 to form the immunoproteasome. In this scenario, we analyzed whether the expression and activity of the constitutive and the immunoproteasome as well as the expression of other components of the MHC class I pathway are altered during the infection of HeLa cells with T. cruzi. By RT-PCR and two-dimensional gel electrophoresis analysis, we showed that the expression and composition of the constitutive proteasome is not affected by the parasite. In contrast, the biosynthesis of the β1i, β2i, β5i immunosubunits, PA28β, TAP1 and the MHC class I molecule as well as the proteasomal proteolytic activities were down-regulated in infected-IFN-γ-treated cell cultures. Taken together, our results provide evidence that the protozoan T. cruzi specifically modulates its infection through an unknown posttranscriptional mechanism that inhibits the expression of the MHC class I pathway components. PMID:24752321

  10. Acidosis increases MHC class II-restricted presentation of a protein endowed with a pH-dependent heparan sulfate-binding ability.

    PubMed

    Knittel, Delphine; Savatier, Alexandra; Upert, Grégory; Lortat-Jacob, Hugues; Léonetti, Michel

    2015-04-15

    Heparan sulfate proteoglycans (HSPGs) are ubiquitously expressed molecules that participate in numerous biological processes. We previously showed that HSPGs expressed on the surface of APCs can serve as receptors for a hybrid protein containing an HS ligand and an Ag, which leads to more efficient stimulation of Th cells. To investigate whether such behavior is shared by proteins with inherent HS-binding ability, we looked for proteins endowed with this characteristic. We found that diphtheria toxin and its nontoxic mutant, called CRM197, can interact with HS. However, we observed that their binding ability is higher at pH 6 than at pH 7.4. Therefore, as extracellular acidosis occurs during infection by various micro-organisms, we assessed whether HS-binding capacity affects MHC class II-restricted presentation at different pHs. We first observed that pH decrease allows CRM197 binding to HSPG-expressing cells, including APCs. Then, we showed that this interaction enhances Ag uptake and presentation to Th cells. Lastly, we observed that pH decrease does not affect processing and presentation abilities of the APCs. Our findings show that acidic pH causes an HSPG-mediated uptake and an enhancement of T cell stimulation of Ags with the inherent ability to bind HSPGs pH-dependently. Furthermore, they suggest that proteins from micro-organisms with this binding characteristic might be supported more efficiently by the adaptive immune system when acidosis is triggered during infection. PMID:25754736

  11. Expression and purification of human MHC class I-related chain molecule B-α1 domain.

    PubMed

    Wang, Shufen; Xiang, Zemin; Wang, Ya; Xu, Huanhuan; Zhang, Dengyang; Wang, Xuanjun; Sheng, Jun

    2016-07-01

    Major histocompatibility complex (MHC) class I-related chain A/B (MICA/B) is a type of stress-induced molecule that plays an important role in tumor surveillance. MICA/B shares a similar structure with MHC class I molecules, but MICA/B contains a closed cleft, not an open one, in its N-terminal alpha1 domain. The alpha 1 domain was believed to have no roles in antigen presentation, because the closed cleft provides limited space for binding with known molecules, and the cleft of MICA/B have been reported no known functions. To study the possible function of the cleft located in human MICA/B's alpha 1 domain, we attempted to express the human MICB-α1 (hMICB-α1) domain allele protein, which is approximately 20.5 kDa, by utilizing an Escherichia coli (E. coli) secretory pathway. Protein expression was accomplished through the phosphate-limited inducible promoter. After purification using ammonium sulfate precipitation, phenyl hydrophobic Sepharose, SP Sepharose and HisTrap affinity Sepharose, recombinant human MICB-α1 (rhMICB-α1) was obtained with 94.3% purity. The binding capacity of rhMICB-α1 with natural killer group 2, member D (NKG2D) was evaluated in vitro. The results demonstrated that rhMICB-α1 can be prepared through the E. coli secretory pathway. Purified rhMICB-α1 protein was able to functionally bind with NKG2D. This method can be further used to obtain functionally active rhMICB-α1 protein, which can served as the basis for further studies of the possible function of the MICB cleft. PMID:27036081

  12. ZXDC, a novel zinc finger protein that binds CIITA and activates MHC gene transcription

    PubMed Central

    Al-Kandari, Wafa; Jambunathan, Srikarthika; Navalgund, Vandana; Koneni, Rupa; Freer, Margot; Parimi, Neeta; Mudhasani, Rajini; Fontes, Joseph D.

    2006-01-01

    The class II trans-activator (CIITA) is recognized as the master regulator of major histocompatibility complex (MHC) class II gene transcription and contributes to the transcription of MHC class I genes. To better understand the function of CIITA, we performed yeast two-hybrid with the C-terminal 807 amino acids of CIITA, and cloned a novel human cDNA named zinc finger, X-linked, duplicated family member C (ZXDC). The 858 amino acid ZXDC protein contains 10 zinc fingers and a transcriptional activation domain, and was found to interact with the region of CIITA containing leucine-rich repeats. Over-expression of ZXDC in human cell lines resulted in super-activation of MHC class I and class II promoters by CIITA. Conversely, silencing of ZXDC expression reduced the ability of CIITA to activate transcription of MHC class II genes. Given the specific interaction between the ZXDC and CIITA proteins, as well as the effect of ZXDC on MHC gene transcription, it appears that ZXDC is an important regulator of both MHC class I and class II transcription. PMID:16600381

  13. MHC class II diversity of koala (Phascolarctos cinereus) populations across their range.

    PubMed

    Lau, Q; Jaratlerdsiri, W; Griffith, J E; Gongora, J; Higgins, D P

    2014-10-01

    Major histocompatibility complex class II (MHCII) genes code for proteins that bind and present antigenic peptides and trigger the adaptive immune response. We present a broad geographical study of MHCII DA β1 (DAB) and DB β1 (DBB) variants of the koala (Phascolarctos cinereus; n=191) from 12 populations across eastern Australia, with a total of 13 DAB and 7 DBB variants found. We identified greater MHCII variation and, possibly, additional gene copies in koala populations in the north (Queensland and New South Wales) relative to the south (Victoria), confirmed by STRUCTURE analyses and genetic differentiation using analysis of molecular variance. The higher MHCII diversity in the north relative to south could potentially be attributed to (i) significant founder effect in Victorian populations linked to historical translocation of bottlenecked koala populations and (ii) increased pathogen-driven balancing selection and/or local genetic drift in the north. Low MHCII genetic diversity in koalas from the south could reduce their potential response to disease, although the three DAB variants found in the south had substantial sequence divergence between variants. This study assessing MHCII diversity in the koala with historical translocations in some populations contributes to understanding the effects of population translocations on functional genetic diversity. PMID:24690756

  14. Measurement of Peptide Binding to MHC Class II Molecules by Fluorescence Polarization.

    PubMed

    Yin, Liusong; Stern, Lawrence J

    2014-01-01

    Peptide binding to major histocompatibility complex class II (MHCII) molecules is a key process in antigen presentation and CD4+ T cell epitope selection. This unit describes a fairly simple but powerful fluorescence polarization-based binding competition assay to measure peptide binding to soluble recombinant MHCII molecules. The binding of a peptide of interest to MHCII molecules is assessed based on its ability to inhibit the binding of a fluorescence-labeled probe peptide, with the strength of binding characterized as IC50 (concentration required for 50% inhibition of probe peptide binding). Data analysis related to this method is discussed. In addition, this unit includes a support protocol for fluorescence labeling peptide using an amine-reactive probe. The advantage of this protocol is that it allows simple, fast, and high-throughput measurements of binding for a large set of peptides to MHCII molecules. PMID:25081912

  15. Vemurafenib enhances MHC induction in BRAFV600E homozygous melanoma cells

    PubMed Central

    Sapkota, Bishu; Hill, Charles E.; Pollack, Brian P.

    2013-01-01

    To optimally integrate targeted kinase inhibitors and immunotherapies in the treatment of melanoma, it will be critical to understand how BRAFV600E mutational status and BRAFV600E inhibition influence the expression of genes that govern antitumor immune responses. Because major histocompatibility complex (MHC) molecules are critical for interactions between tumor cells and lymphocytes, we investigated the impact of BRAFV600E-selective inhibitors on the expression of MHC molecules. We found that the treatment of A375 melanoma cells with vemurafenib enhances the induction of MHC Class I and Class II molecules by interferon γ and IFNα2b. Consistent with these findings, we observed that the forced overexpression of BRAFV600E has the opposite effect and can repress the baseline expression of MHC Class I molecules in A375 cells. Further studies utilizing eight other melanoma cell lines revealed that the vemurafenib-mediated enhancement of MHC induction by IFNγ only occurs in the context of homozygous, but not heterozygous, BRAFV600E mutation. These findings suggest that BRAFV600Eactivity directly influences the expression of MHC molecules and the response to Type I and Type II IFNs. Furthermore, our data suggest that the effect of vemurafenib on the expression of immune system-relevant genes may depend on the zygosity of the BRAFV600E mutation, which is not routinely assessed in melanoma patients. PMID:23483066

  16. MHC class II compatibility in aborted fetuses and term infants of couples with recurrent spontaneous abortion.

    PubMed

    Ober, C; Steck, T; van der Ven, K; Billstrand, C; Messer, L; Kwak, J; Beaman, K; Beer, A

    1993-12-01

    Maternal-fetal histocompatibility for alleles at HLA class II loci, HLA-DQA1 and HLA-DQB1, was examined in 40 abortuses and 31 liveborn children of 68 couples with a history of idiopathic recurrent spontaneous abortion (RSAB) who underwent leukocyte immunization prior to the index pregnancy. Significantly more couples with RSAB shared two HLA-DQA1 alleles as compared with fertile control couples (0.18 vs. 0.03, respectively; P = 0.031). There were no differences in HLA sharing between couples with RSAB who experienced a repeat abortion in the index pregnancy as compared with couples with RSAB who were delivered of a liveborn child. Non-significant deficits of abortuses who were compatible for alleles at the HLA-DQA1 (6 observed vs. 8.5 expected; P = 0.225) and the HLA-DQB1 (7 observed vs. 9.2 expected; P = 0.254) loci were observed. A significant deficit of HLA-DQA1 compatible liveborn children was observed (1 observed vs. 5.5 expected; P = 0.0069). The overall deficit of HLA-DQA1 compatible fetuses (7 observed vs. 14.0 expected; P = 0.0018) after approximately 8 weeks gestation suggests that HLA-DQA1 compatible fetuses may be aborted early in pregnancy, prior to the time when fetal tissue can be recovered for genetic studies. PMID:8207709

  17. ITAM signaling in dendritic cells controls T helper cell priming by regulating MHC class II recycling

    PubMed Central

    Graham, Daniel B.; Akilesh, Holly M.; Gmyrek, Grzegorz B.; Piccio, Laura; Gilfillan, Susan; Sim, Julia; Belizaire, Roger; Carrero, Javier A.; Wang, Yinan; Blaufuss, Gregory S.; Sandoval, Gabriel; Fujikawa, Keiko; Cross, Anne H.; Russell, John H.; Cella, Marina

    2010-01-01

    Immature dendritic cells (DCs) specialize in antigen capture and maintain a highly dynamic pool of intracellular major histocompatibility complex class II (MHCII) that continuously recycles from peptide loading compartments to the plasma membrane and back again. This process facilitates sampling of environmental antigens for presentation to T helper cells. Here, we show that a signaling pathway mediated by the DC immunoreceptor tyrosine-based activation motif (ITAM)–containing adaptors (DAP12 and FcRγ) and Vav family guanine nucleotide exchange factors controls the half-life of surface peptide-MHCII (pMHCII) complexes and is critical for CD4 T-cell triggering in vitro. Strikingly, mice with disrupted DC ITAMs show defective T helper cell priming in vivo and are protected from experimental autoimmune encephalitis. Mechanistically, we show that deficiency in ITAM signaling results in increased pMHCII internalization, impaired recycling, and an accumulation of ubiquitinated MHCII species that are prematurely degraded in lysosomes. We propose a novel mechanism for control of T helper cell priming. PMID:20634378

  18. Processing and MHC class II presentation of exogenous soluble antigen involving a proteasome-dependent cytosolic pathway in CD40-activated B cells.

    PubMed

    Becker, Hans Jiro; Kondo, Eisei; Shimabukuro-Vornhagen, Alexander; Theurich, Sebastian; von Bergwelt-Baildon, Michael S

    2016-08-01

    Activated B cells have the capacity to present antigen and induce immune responses as potent antigen-presenting cells (APCs). As in other APCs, antigen presentation by B cells involves antigen internalization, antigen processing, and peptide loading onto MHC molecules. However, while the mechanism of antigen processing has been studied extensively in other APCs, this pathway remains elusive in B cells. The aim of this study was to investigate the MHC class II processing pathway in CD40-activated B cells (CD40Bs), as a model for activated, antigen-presenting B cells. Using CMV pp65 as a model antigen, we evaluated processing and presentation of the CD4 + T-cell epitope 509-523 (K509) by human CD40Bs in ELISPOT assays. As expected, stimulation of specific CD4 + T-cell clones was attenuated after pretreatment of CD40Bs with inhibitors of classic class II pathway components. However, proteasome inhibitors such as epoxomicin limited antigen presentation as well. This suggests that the antigen is processed in a non-classical, cytosolic MHC class II pathway. Further experiments with truncated protein variants revealed involvement of the proteasome in processing of the N and C extensions of the epitope. Access to the cytosol was shown to be size dependent. Epoxomicin sensitivity exclusively in CD40B cells, but not in dendritic cells, suggests a novel processing mechanism unique to this APC. Our data suggest that B cells process antigen using a distinct, non-classical class II pathway. PMID:26561366

  19. Structure and evolution of a new avian MHC class II B gene in a sub-Antarctic seabird, the thin-billed prion (Procellariiformes: Pachyptila belcheri).

    PubMed

    Silva, Mónica C; Edwards, Scott V

    2009-03-01

    The major histocompatibility complex encodes molecules that present foreign peptides to T cells of the immune system. The peptide binding region (PBR) of these molecules is among the most polymorphic regions found in vertebrate taxa. Genomic cloning approaches are improving our understanding of the evolution of this multigene family in nonmodel avian groups. By building a cosmid library, a new MHC class II B gene, Pabe-DAB1, was isolated and characterized at the genomic level in a sub-Antarctic seabird, the thin-billed prion (Pachyptila belcheri). Pabe-DAB1 exhibits the hallmark structural features of functional MHC class II loci. Direct sequencing of the PBR encoding exon in a panel of prions revealed significantly higher levels of genetic diversity compared to two noncoding neutral loci, with most alleles differing by at least one replacement substitution in the peptide binding codons. We estimated evolutionary dynamics for Pabe-DAB1 using a variety of Bayesian and other approaches. Evidence for balancing selection comes from a spatially variable ratio of nonsynonymous-to-synonymous substitutions (mean d (N)/d (S) = 2.87) in the PBR, with sites predicted to be functionally relevant exhibiting the highest omega values. We estimate the population recombination rate to be approximately 0.3 per site per generation, indicating an important role for recombination in generating polymorphism at this locus. Pabe-DAB1 is among the few avian class II loci characterized at the genomic level and with a known intron-exon structure, a feature that greatly facilitated the amplification and sequencing of a single MHC locus in this species. PMID:19209378

  20. Changes in variation at the MHC class II DQA locus during the final demise of the woolly mammoth

    PubMed Central

    Pečnerová, Patrícia; Díez-del-Molino, David; Vartanyan, Sergey; Dalén, Love

    2016-01-01

    According to the nearly-neutral theory of evolution, the relative strengths of selection and drift shift in favour of drift at small population sizes. Numerous studies have analysed the effect of bottlenecks and small population sizes on genetic diversity in the MHC, which plays a central role in pathogen recognition and immune defense and is thus considered a model example for the study of adaptive evolution. However, to understand changes in genetic diversity at loci under selection, it is necessary to compare the genetic diversity of a population before and after the bottleneck. In this study, we analyse three fragments of the MHC DQA gene in woolly mammoth samples radiocarbon dated to before and after a well-documented bottleneck that took place about ten thousand years ago. Our results indicate a decrease in observed heterozygosity and number of alleles, suggesting that genetic drift had an impact on the variation on MHC. Based on coalescent simulations, we found no evidence of balancing selection maintaining MHC diversity during the Holocene. However, strong trans-species polymorphism among mammoths and elephants points to historical effects of balancing selection on the woolly mammoth lineage. PMID:27143688

  1. Changes in variation at the MHC class II DQA locus during the final demise of the woolly mammoth.

    PubMed

    Pečnerová, Patrícia; Díez-Del-Molino, David; Vartanyan, Sergey; Dalén, Love

    2016-01-01

    According to the nearly-neutral theory of evolution, the relative strengths of selection and drift shift in favour of drift at small population sizes. Numerous studies have analysed the effect of bottlenecks and small population sizes on genetic diversity in the MHC, which plays a central role in pathogen recognition and immune defense and is thus considered a model example for the study of adaptive evolution. However, to understand changes in genetic diversity at loci under selection, it is necessary to compare the genetic diversity of a population before and after the bottleneck. In this study, we analyse three fragments of the MHC DQA gene in woolly mammoth samples radiocarbon dated to before and after a well-documented bottleneck that took place about ten thousand years ago. Our results indicate a decrease in observed heterozygosity and number of alleles, suggesting that genetic drift had an impact on the variation on MHC. Based on coalescent simulations, we found no evidence of balancing selection maintaining MHC diversity during the Holocene. However, strong trans-species polymorphism among mammoths and elephants points to historical effects of balancing selection on the woolly mammoth lineage. PMID:27143688

  2. Changes in variation at the MHC class II DQA locus during the final demise of the woolly mammoth

    NASA Astrophysics Data System (ADS)

    Pečnerová, Patrícia; Díez-Del-Molino, David; Vartanyan, Sergey; Dalén, Love

    2016-05-01

    According to the nearly-neutral theory of evolution, the relative strengths of selection and drift shift in favour of drift at small population sizes. Numerous studies have analysed the effect of bottlenecks and small population sizes on genetic diversity in the MHC, which plays a central role in pathogen recognition and immune defense and is thus considered a model example for the study of adaptive evolution. However, to understand changes in genetic diversity at loci under selection, it is necessary to compare the genetic diversity of a population before and after the bottleneck. In this study, we analyse three fragments of the MHC DQA gene in woolly mammoth samples radiocarbon dated to before and after a well-documented bottleneck that took place about ten thousand years ago. Our results indicate a decrease in observed heterozygosity and number of alleles, suggesting that genetic drift had an impact on the variation on MHC. Based on coalescent simulations, we found no evidence of balancing selection maintaining MHC diversity during the Holocene. However, strong trans-species polymorphism among mammoths and elephants points to historical effects of balancing selection on the woolly mammoth lineage.

  3. Clonal expansion of double-positive intraepithelial lymphocytes by MHC class I-related chain A expressed in mouse small intestinal epithelium.

    PubMed

    Park, Eun Jeong; Takahashi, Ichiro; Ikeda, Junko; Kawahara, Kazuko; Okamoto, Tetsuji; Kweon, Mi-Na; Fukuyama, Satoshi; Groh, Veronika; Spies, Thomas; Obata, Yuichi; Miyazaki, Jun-Ichi; Kiyono, Hiroshi

    2003-10-15

    Expression of a distant homologue MHC class I molecule, MHC class I-related chain A (MICA), has been found to be stress inducible and limited to the intestinal epithelium. This nonclassical MHC molecule is associated with various carcinomas in humans. To understand the biological consequences of MICA expression in the gut, we generated transgenic (Tg) mice (T3(b)-MICA Tg) under the control of the T3(b) promoter. The T3(b)-MICA Tg mice expressed MICA selectively in the intestine and had an increased number of TCRalphabeta CD4CD8alphaalpha, double-positive (DP) intraepithelial lymphocytes (IELs) in the small bowel. These MICA-expanded DP IELs exhibited a bias to Vbeta8.2 and overlapped motifs of the complementarity-determining region 3 region among various Tg mice. Hence, the overexpression of MICA resulted in a clonal expansion of DP IELs. Studies in model of inflammatory bowel disease showed that transgenic MICA was able to attenuate the acute colitis induced by dextran sodium sulfate administration. Therefore, this unique in vivo model will enable investigation of possible influences of stress-inducible MICA on the gut immune surveillance. PMID:14530335

  4. Expression of the mouse MHC class Ib H2-T11 gene product, a paralog of H2-T23 (Qa-1) with shared peptide-binding specificity.

    PubMed

    Chen, Lili; Reyes-Vargas, Eduardo; Dai, Hu; Escobar, Hernando; Rudd, Brant; Fairbanks, Jared; Ho, Alexander; Cusick, Mathew F; Kumánovics, Attila; Delgado, Julio; He, Xiao; Jensen, Peter E

    2014-08-01

    The mouse MHC class Ib gene H2-T11 is 95% identical at the DNA level to H2-T23, which encodes Qa-1, one of the most studied MHC class Ib molecules. H2-T11 mRNA was observed to be expressed widely in tissues of C57BL/6 mice, with the highest levels in thymus. To circumvent the availability of a specific mAb, cells were transduced with cDNA encoding T11 with a substituted α3 domain. Hybrid T11D3 protein was expressed at high levels similar to control T23D3 molecules on the surface of both TAP(+) and TAP(-) cells. Soluble T11D3 was generated by folding in vitro with Qa-1 determinant modifier, the dominant peptide presented by Qa-1. The circular dichroism spectrum of this protein was similar to that of other MHC class I molecules, and it was observed to bind labeled Qa-1 determinant modifier peptide with rapid kinetics. By contrast to the Qa-1 control, T11 tetramers did not react with cells expressing CD94/NKG2A, supporting the conclusion that T11 cannot replace Qa-1 as a ligand for NK cell inhibitory receptors. T11 also failed to substitute for Qa-1 in the presentation of insulin to a Qa-1-restricted T cell hybridoma. Despite divergent function, T11 was observed to share peptide-loading specificity with Qa-1. Direct analysis by tandem mass spectrometry of peptides eluted from T11D3 and T23D3 isolated from Hela cells demonstrated a diversity of peptides with a clear motif that was shared between the two molecules. Thus, T11 is a paralog of T23 encoding an MHC class Ib molecule that shares peptide-binding specificity with Qa-1 but differs in function. PMID:24958902

  5. DC-expressed MHC class I single-chain trimer-based vaccines prime cytotoxic T lymphocytes against exogenous but not endogenous antigens.

    PubMed

    Ordaz, Maria L; Larmonier, Nicolas; Lybarger, Lonnie

    2010-01-01

    The poor immunogenicity of many tumors can be partly explained by the inefficiency of the MHC class I peptide presentation pathway. MHC-I-based single-chain trimers (SCT) represent a new class of molecules with the potential to overcome this limitation. We here evaluated the ability of SCT presenting a melanoma antigen peptide (TRP-2) to prime cytotoxic T lymphocyte (CTL) responses in mice when given as DNA vaccines via Gene Gun or when expressed by dendritic cells. The SCT was unable to induce detectable priming or significant anti-tumor activity of CTL using either vaccination strategy, whereas control SCT (with an exogenous peptide) primed strong responses. This study thus provides the first data related to the use of SCT in combination with DC and their application toward self antigens and suggest this potent technology, alone, is insufficient to overcome self tolerance. PMID:20199770

  6. Genetic Variation of the Major Histocompatibility Complex (MHC Class II B Gene) in the Threatened Hume’s Pheasant, Syrmaticus humiae

    PubMed Central

    Chen, Weicai; Bei, Yongjian; Li, Hanhua

    2015-01-01

    Major histocompatibility complex (MHC) genes are the most polymorphic genes in vertebrates and encode molecules that play a crucial role in pathogen resistance. As a result of their diversity, they have received much attention in the fields of evolutionary and conservation biology. Here, we described the genetic variation of MHC class II B (MHCIIB) exon 2 in a wild population of Hume’s pheasant (Syrmaticus humiae), which has suffered a dramatic decline in population over the last three decades across its ranges in the face of heavy exploitation and habitat loss. Twenty-four distinct alleles were found in 73 S. humiae specimens. We found seven shared alleles among four geographical groups as well as six rare MHCIIB alleles. Most individuals displayed between one to five alleles, suggesting that there are at least three MHCIIB loci of the Hume’s pheasant. The dN ⁄ dS ratio at putative antigen-binding sites (ABS) was significantly greater than one, indicating balancing selection is acting on MHCIIB exon 2. Additionally, recombination and gene conversion contributed to generating MHCIIB diversity in the Hume’s pheasant. One to three recombination events and seventy-five significant gene conversion events were observed within the Hume’s pheasant MHCIIB loci. The phylogenetic tree and network analysis revealed that the Hume’s pheasant alleles do not cluster together, but are scattered through the tree or network indicating a trans-species evolutionary mode. These findings revealed the evolution of the Hume’s pheasant MHC after suffering extreme habitat fragmentation. PMID:25629763

  7. Epigenetic Mechanisms Regulate MHC and Antigen Processing Molecules in Human Embryonic and Induced Pluripotent Stem Cells

    PubMed Central

    Suárez-Álvarez, Beatriz; Rodriguez, Ramón M.; Calvanese, Vincenzo; Blanco-Gelaz, Miguel A.; Suhr, Steve T.; Ortega, Francisco; Otero, Jesus; Cibelli, Jose B.; Moore, Harry; Fraga, Mario F.; López-Larrea, Carlos

    2010-01-01

    Background Human embryonic stem cells (hESCs) are an attractive resource for new therapeutic approaches that involve tissue regeneration. hESCs have exhibited low immunogenicity due to low levels of Mayor Histocompatibility Complex (MHC) class-I and absence of MHC class-II expression. Nevertheless, the mechanisms regulating MHC expression in hESCs had not been explored. Methodology/Principal Findings We analyzed the expression levels of classical and non-classical MHC class-I, MHC class-II molecules, antigen-processing machinery (APM) components and NKG2D ligands (NKG2D-L) in hESCs, induced pluripotent stem cells (iPSCs) and NTera2 (NT2) teratocarcinoma cell line. Epigenetic mechanisms involved in the regulation of these genes were investigated by bisulfite sequencing and chromatin immunoprecipitation (ChIP) assays. We showed that low levels of MHC class-I molecules were associated with absent or reduced expression of the transporter associated with antigen processing 1 (TAP-1) and tapasin (TPN) components in hESCs and iPSCs, which are involved in the transport and load of peptides. Furthermore, lack of β2-microglobulin (β2m) light chain in these cells limited the expression of MHC class I trimeric molecule on the cell surface. NKG2D ligands (MICA, MICB) were observed in all pluripotent stem cells lines. Epigenetic analysis showed that H3K9me3 repressed the TPN gene in undifferentiated cells whilst HLA-B and β2m acquired the H3K4me3 modification during the differentiation to embryoid bodies (EBs). Absence of HLA-DR and HLA-G expression was regulated by DNA methylation. Conclusions/Significance Our data provide fundamental evidence for the epigenetic control of MHC in hESCs and iPSCs. Reduced MHC class I and class II expression in hESCs and iPSCs can limit their recognition by the immune response against these cells. The knowledge of these mechanisms will further allow the development of strategies to induce tolerance and improve stem cell allograft acceptance

  8. The Dendritic Cell Major Histocompatibility Complex II (MHC II) Peptidome Derives from a Variety of Processing Pathways and Includes Peptides with a Broad Spectrum of HLA-DM Sensitivity.

    PubMed

    Clement, Cristina C; Becerra, Aniuska; Yin, Liusong; Zolla, Valerio; Huang, Liling; Merlin, Simone; Follenzi, Antonia; Shaffer, Scott A; Stern, Lawrence J; Santambrogio, Laura

    2016-03-11

    The repertoire of peptides displayed in vivo by MHC II molecules derives from a wide spectrum of proteins produced by different cell types. Although intracellular endosomal processing in dendritic cells and B cells has been characterized for a few antigens, the overall range of processing pathways responsible for generating the MHC II peptidome are currently unclear. To determine the contribution of non-endosomal processing pathways, we eluted and sequenced over 3000 HLA-DR1-bound peptides presented in vivo by dendritic cells. The processing enzymes were identified by reference to a database of experimentally determined cleavage sites and experimentally validated for four epitopes derived from complement 3, collagen II, thymosin β4, and gelsolin. We determined that self-antigens processed by tissue-specific proteases, including complement, matrix metalloproteases, caspases, and granzymes, and carried by lymph, contribute significantly to the MHC II self-peptidome presented by conventional dendritic cells in vivo. Additionally, the presented peptides exhibited a wide spectrum of binding affinity and HLA-DM susceptibility. The results indicate that the HLA-DR1-restricted self-peptidome presented under physiological conditions derives from a variety of processing pathways. Non-endosomal processing enzymes add to the number of epitopes cleaved by cathepsins, altogether generating a wider peptide repertoire. Taken together with HLA-DM-dependent and-independent loading pathways, this ensures that a broad self-peptidome is presented by dendritic cells. This work brings attention to the role of "self-recognition" as a dynamic interaction between dendritic cells and the metabolic/catabolic activities ongoing in every parenchymal organ as part of tissue growth, remodeling, and physiological apoptosis. PMID:26740625

  9. HLA-DRB1*11 and variants of the MHC class II locus are strong risk factors for systemic juvenile idiopathic arthritis.

    PubMed

    Ombrello, Michael J; Remmers, Elaine F; Tachmazidou, Ioanna; Grom, Alexei; Foell, Dirk; Haas, Johannes-Peter; Martini, Alberto; Gattorno, Marco; Özen, Seza; Prahalad, Sampath; Zeft, Andrew S; Bohnsack, John F; Mellins, Elizabeth D; Ilowite, Norman T; Russo, Ricardo; Len, Claudio; Hilario, Maria Odete E; Oliveira, Sheila; Yeung, Rae S M; Rosenberg, Alan; Wedderburn, Lucy R; Anton, Jordi; Schwarz, Tobias; Hinks, Anne; Bilginer, Yelda; Park, Jane; Cobb, Joanna; Satorius, Colleen L; Han, Buhm; Baskin, Elizabeth; Signa, Sara; Duerr, Richard H; Achkar, J P; Kamboh, M Ilyas; Kaufman, Kenneth M; Kottyan, Leah C; Pinto, Dalila; Scherer, Stephen W; Alarcón-Riquelme, Marta E; Docampo, Elisa; Estivill, Xavier; Gül, Ahmet; de Bakker, Paul I W; Raychaudhuri, Soumya; Langefeld, Carl D; Thompson, Susan; Zeggini, Eleftheria; Thomson, Wendy; Kastner, Daniel L; Woo, Patricia

    2015-12-29

    Systemic juvenile idiopathic arthritis (sJIA) is an often severe, potentially life-threatening childhood inflammatory disease, the pathophysiology of which is poorly understood. To determine whether genetic variation within the MHC locus on chromosome 6 influences sJIA susceptibility, we performed an association study of 982 children with sJIA and 8,010 healthy control subjects from nine countries. Using meta-analysis of directly observed and imputed SNP genotypes and imputed classic HLA types, we identified the MHC locus as a bona fide susceptibility locus with effects on sJIA risk that transcended geographically defined strata. The strongest sJIA-associated SNP, rs151043342 [P = 2.8 × 10(-17), odds ratio (OR) 2.6 (2.1, 3.3)], was part of a cluster of 482 sJIA-associated SNPs that spanned a 400-kb region and included the class II HLA region. Conditional analysis controlling for the effect of rs151043342 found that rs12722051 independently influenced sJIA risk [P = 1.0 × 10(-5), OR 0.7 (0.6, 0.8)]. Meta-analysis of imputed classic HLA-type associations in six study populations of Western European ancestry revealed that HLA-DRB1*11 and its defining amino acid residue, glutamate 58, were strongly associated with sJIA [P = 2.7 × 10(-16), OR 2.3 (1.9, 2.8)], as was the HLA-DRB1*11-HLA-DQA1*05-HLA-DQB1*03 haplotype [6.4 × 10(-17), OR 2.3 (1.9, 2.9)]. By examining the MHC locus in the largest collection of sJIA patients assembled to date, this study solidifies the relationship between the class II HLA region and sJIA, implicating adaptive immune molecules in the pathogenesis of sJIA. PMID:26598658

  10. HLA-DRB1*11 and variants of the MHC class II locus are strong risk factors for systemic juvenile idiopathic arthritis

    PubMed Central

    Ombrello, Michael J.; Remmers, Elaine F.; Tachmazidou, Ioanna; Grom, Alexei; Foell, Dirk; Haas, Johannes-Peter; Martini, Alberto; Gattorno, Marco; Özen, Seza; Prahalad, Sampath; Zeft, Andrew S.; Bohnsack, John F.; Mellins, Elizabeth D.; Ilowite, Norman T.; Russo, Ricardo; Len, Claudio; Hilario, Maria Odete E.; Oliveira, Sheila; Yeung, Rae S. M.; Rosenberg, Alan; Wedderburn, Lucy R.; Anton, Jordi; Schwarz, Tobias; Hinks, Anne; Bilginer, Yelda; Park, Jane; Cobb, Joanna; Satorius, Colleen L.; Han, Buhm; Baskin, Elizabeth; Signa, Sara; Duerr, Richard H.; Achkar, J. P.; Kamboh, M. Ilyas; Kaufman, Kenneth M.; Kottyan, Leah C.; Pinto, Dalila; Scherer, Stephen W.; Alarcón-Riquelme, Marta E.; Docampo, Elisa; Estivill, Xavier; Gül, Ahmet; de Bakker, Paul I. W.; Raychaudhuri, Soumya; Langefeld, Carl D.; Thompson, Susan; Zeggini, Eleftheria; Thomson, Wendy; Kastner, Daniel L.; Woo, Patricia

    2015-01-01

    Systemic juvenile idiopathic arthritis (sJIA) is an often severe, potentially life-threatening childhood inflammatory disease, the pathophysiology of which is poorly understood. To determine whether genetic variation within the MHC locus on chromosome 6 influences sJIA susceptibility, we performed an association study of 982 children with sJIA and 8,010 healthy control subjects from nine countries. Using meta-analysis of directly observed and imputed SNP genotypes and imputed classic HLA types, we identified the MHC locus as a bona fide susceptibility locus with effects on sJIA risk that transcended geographically defined strata. The strongest sJIA-associated SNP, rs151043342 [P = 2.8 × 10−17, odds ratio (OR) 2.6 (2.1, 3.3)], was part of a cluster of 482 sJIA-associated SNPs that spanned a 400-kb region and included the class II HLA region. Conditional analysis controlling for the effect of rs151043342 found that rs12722051 independently influenced sJIA risk [P = 1.0 × 10−5, OR 0.7 (0.6, 0.8)]. Meta-analysis of imputed classic HLA-type associations in six study populations of Western European ancestry revealed that HLA-DRB1*11 and its defining amino acid residue, glutamate 58, were strongly associated with sJIA [P = 2.7 × 10−16, OR 2.3 (1.9, 2.8)], as was the HLA-DRB1*11—HLA-DQA1*05—HLA-DQB1*03 haplotype [6.4 × 10−17, OR 2.3 (1.9, 2.9)]. By examining the MHC locus in the largest collection of sJIA patients assembled to date, this study solidifies the relationship between the class II HLA region and sJIA, implicating adaptive immune molecules in the pathogenesis of sJIA. PMID:26598658