scholarly journals CIN85 drives B cell responses by linking BCR signals to the canonical NF-κB pathway

2011 ◽  
Vol 208 (7) ◽  
pp. 1447-1457 ◽  
Author(s):  
Kohei Kometani ◽  
Takayuki Yamada ◽  
Yoshiteru Sasaki ◽  
Tadashi Yokosuka ◽  
Takashi Saito ◽  
...  
Keyword(s):  
T Cell ◽  
B Cell ◽  
Adaptor Protein ◽  
Cellular Responses ◽  
Cell Receptor ◽  
Antibody Responses ◽  
Cell Responses ◽  
Subsequent Degradation

CIN85, an adaptor protein which binds the C-terminal domain of tyrosine phosphorylated Cbl and Cbl-b, has been thought to be involved in the internalization and subsequent degradation of receptors. However, its physiological function remains unclear. To determine its role in B cells, we used Mb1-cre to generate mice with a B cell–specific deletion of CIN85. These mice had impaired T cell–independent type II antibody responses in vivo and diminished IKK-β activation and cellular responses to B cell receptor (BCR) cross-linking in vitro. Introduction of a constitutively active IKK-β construct corrected the defective antibody responses as well as cellular responses in the mutant mice. Together, our results suggest that CIN85 links the BCR to IKK-β activation, thereby contributing to T cell–independent immune responses.

scholarly journals Cd5 Maintains Tolerance in Anergic B Cells

2000 ◽  
Vol 191 (5) ◽  
pp. 883-890 ◽  
Author(s):  
Keli L. Hippen ◽  
Lina E. Tze ◽  
Timothy W. Behrens
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Immunoglobulin M ◽  
B Cell Tolerance ◽  
Cell Responses ◽  
B Cell Anergy ◽  
Clonal Anergy

Clonal anergy of autoreactive B cells is a key mechanism regulating tolerance. Here, we show that anergic B cells express significant surface levels of CD5, a molecule normally found on T cells and a subset of B-1 cells. Breeding of the hen egg lysozyme (HEL) transgenic model for B cell anergy onto the CD5 null background resulted in a spontaneous loss of B cell tolerance in vivo. Evidence for this included elevated levels of anti-HEL immunoglobulin M (IgM) antibodies in the serum of CD5−/− mice transgenic for both an HEL-specific B cell receptor (BCR) and soluble lysozyme. “Anergic” B cells lacking CD5 also showed enhanced proliferative responses in vitro and elevated intracellular Ca2+ levels at rest and after IgM cross-linking. These data support the hypothesis that CD5 negatively regulates Ig receptor signaling in anergic B cells and functions to inhibit autoimmune B cell responses.

scholarly journals Development of a System To Study CD4+-T-Cell Responses to Transgenic Ovalbumin-Expressing Toxoplasma gondii during Toxoplasmosis

2004 ◽  
Vol 72 (12) ◽  
pp. 7240-7246 ◽  
Author(s):  
Marion Pepper ◽  
Florence Dzierszinski ◽  
Amy Crawford ◽  
Christopher A. Hunter ◽  
David Roos
Keyword(s):  
T Cells ◽  
T Cell ◽  
Toxoplasma Gondii ◽  
T Cell Responses ◽  
Cell Receptor ◽  
In Vivo Studies ◽  
Cell Responses ◽  
Ifn Γ

ABSTRACT The study of the immune response to Toxoplasma gondii has provided numerous insights into the role of T cells in resistance to intracellular infections. However, the complexity of this eukaryote pathogen has made it difficult to characterize immunodominant epitopes that would allow the identification of T cells with a known specificity for parasite antigens. As a consequence, analysis of T-cell responses to T. gondii has been based on characterization of the percentage of T cells that express an activated phenotype during infection and on the ability of these cells to produce cytokines in response to complex mixtures of parasite antigens. In order to study specific CD4+ T cells responses to T. gondii, recombinant parasites that express a truncated ovalbumin (OVA) protein, in either a cytosolic or a secreted form, were engineered. In vitro and in vivo studies reveal that transgenic parasites expressing secreted OVA are able to stimulate T-cell receptor-transgenic OVA-specific CD4+ T cells to proliferate, express an activated phenotype, and produce gamma interferon (IFN-γ). Furthermore, the adoptive transfer of OVA-specific T cells into IFN-γ−/− mice provided enhanced protection against infection with the OVA-transgenic (but not parental) parasites. Together, these studies establish the utility of this transgenic system to study CD4+-T-cell responses during toxoplasmosis.

scholarly journals Regulation and Function of the CD3γ DxxxLL Motif: A Binding Site for Adaptor Protein-1 and Adaptor Protein-2 in Vitro

1997 ◽  
Vol 138 (2) ◽  
pp. 271-281 ◽  
Author(s):  
Jes Dietrich ◽  
Jesper Kastrup ◽  
Bodil L. Nielsen ◽  
Niels Ødum ◽  
Carsten Geisler
Keyword(s):  
T Cell ◽  
T Cell Activation ◽  
Cell Activation ◽  
Tolerance Induction ◽  
Adaptor Protein ◽  
Adaptor Proteins ◽  
Cell Receptor ◽  
Coated Vesicle

Several receptors are downregulated by internalization after ligand binding. Regulation of T cell receptor (TCR) expression is an important step in T cell activation, desensitization, and tolerance induction. One way T cells regulate TCR expression is by phosphorylation/dephosphorylation of the TCR subunit clusters of differentiation (CD)3γ. Thus, phosphorylation of CD3γ serine 126 (S126) causes a downregulation of the TCR. In this study, we have analyzed the CD3γ internalization motif in three different systems in parallel: in the context of the complete multimeric TCR; in monomeric CD4/CD3γ chimeras; and in vitro by binding CD3γ peptides to clathrin-coated vesicle adaptor proteins (APs). We find that the CD3γ D127xxxLL131/132 sequence represents one united motif for binding of both AP-1 and AP-2, and that this motif functions as an active sorting motif in monomeric CD4/ CD3γ molecules independently of S126. An acidic amino acid is required at position 127 and a leucine (L) is required at position 131, whereas the requirements for position 132 are more relaxed. The spacing between aspartic acid 127 (D127) and L131 is crucial for the function of the motif in vivo and for AP binding in vitro. Furthermore, we provide evidence indicating that phosphorylation of CD3γ S126 in the context of the complete TCR induces a conformational change that exposes the DxxxLL sequence for AP binding. Exposure of the DxxxLL motif causes an increase in the TCR internalization rate and we demonstrate that this leads to an impairment of TCR signaling. On the basis of the present results, we propose the existence of at least three different types of L-based receptor sorting motifs.

scholarly journals Regulated Antigen Expression in Live RecombinantSalmonella enterica Serovar Typhimurium Strongly Affects Colonization Capabilities and Specific CD4+-T-Cell Responses

2001 ◽  
Vol 69 (12) ◽  
pp. 7493-7500 ◽  
Author(s):  
Dirk Bumann
Keyword(s):  
T Cell ◽  
T Cell Responses ◽  
Antigen Expression ◽  
Cell Receptor ◽  
Cell Responses ◽  
Wide Range ◽  
Serovar Typhimurium ◽  
Cellular Immune

ABSTRACT Regulated antigen expression can influence the immunogenicity of live recombinant Salmonella vaccines, but a rational optimization has remained difficult since important aspects of this effect are incompletely understood. Here, attenuated Salmonella enterica serovar Typhimurium SL3261 strains expressing the model antigen GFP_OVA were used to quantify in vivo antigen levels by flow cytometry and to simultaneously follow the crucial early steps of antigen-specific T-cell responses in mice that are transgenic for a T-cell receptor recognizing ovalbumin. Among seven tested promoters,P pagC has the highest activity in murine tissues combined with low in vitro expression, whereasP tac has a comparable in vivo and a very high in vitro activity. Both SL3261 (pPpagCGFP_OVA) and SL3261 (pPtacGFP_OVA) cells can induce potent ovalbumin-specific cellular immune responses following oral administration, but doses almost 1,000-fold lower are sufficient for the in vivo-inducible construct SL3261 (pPpagCGFP_OVA) compared to SL3261 (pPtacGFP_OVA). This efficacy difference is largely explained by impaired early colonization capabilities of SL3261 (pPtacGFP_OVA) cells. Based on the findings of this study, appropriate in vivo expression levels for any given antigen can be rationally selected from the increasing set of promoters with defined properties. This will allow the improvement of recombinantSalmonella vaccines against a wide range of pathogens.

Prediction of MHC Class I-Restricted Immunogenic Epitopes of B-Cell Receptors Indicates T Cell-Mediated Immunosurveillance of Malignant B Cell Clones.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 1949-1949
Author(s):  
Anna-Maria Strothmeyer ◽  
Marcus Duehren-von Minden ◽  
Marcelo A Navarrete ◽  
Kristina Heining-Mikesch ◽  
Hendrik Veelken
Keyword(s):  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Mhc Class I ◽  
Cell Activity ◽  
Class I ◽  
Cell Responses ◽  
Hla Haplotypes

Abstract Abstract 1949 Poster Board I-972 Tumor-specific immune responses can be induced in patients with indolent B cell lymphomas (iNHL) by active immunization against the individual B cell receptor (BCR) expressed by the malignant B cell clone, the so-called “idiotype” (Id). In subsequent trials of intradermal vaccination with recombinant lymphoma-derived Fab fragment in iNHL, we have studied the specificity of MHC class I-restricted anti-Id T cell responses by epitope mapping experiments with synthetic Id-derived peptides predicted to be presented by the respective patient's HLA complex. While such peptides exist in hypervariable and conserved Id regions, these assays have shown consistently that in vivo-induced T cell responses occur preferentially against individual Id epitopes located in complementarity-determining regions (CDR), whereas framework (FR) and constant region-derived epitopes are ignored (Bertinetti et al., Cancer Res. 2006; Navarrete et al., ASH 2008). These results contrast with in vitro studies showing that FR-derived peptides are excellent targets for cytotoxic T cells in iNHL patients (Trojan et al., Nat Med 2000). To gain further insight into the relative predominance and immunological role of MHC class I-restricted Id epitopes, we conducted a comprehensive reverse immunology study in follicular lymphoma (FL). Clonal and functional IgH chain transcript sequences were identified from tumor biopsies of 39 FL patients by A-PCR (Bertinetti et al., EJH 2006). The HLA-A and B haplotype of the patients was determined by conventional serological testing and high-resolution PCR genotyping. Potentially MHC-presentable peptides from all Id sequences and their corresponding germ-line (GL) VH genes were identified for the HLA haplotypes of all 39 patients by reverse immunology (bimas.cit.nih.gov). Identified peptides were ranked for each haplotype according to their predicted score, and the sum of the scores for the 20 highest ranking peptides was calculated. The sum score for any given Id was compared to the mean of the sum scores of the other 38 Ids on the respective patient's HLA haplotypes. Separate analyses were performed for CDR peptides (containing at least 2 AA in any CDR) versus non-CDR-peptides (allocated through imgt.cines.fr), Id versus GL sequences, and Id versus contaminating sporadic Ig sequences that represent bona fide normal B cells in the biopsies. 72% of all peptides with BIMAS scores of ≥50 and ≥10, respectively, were located in FR, expecially in FR3. The ranked sum Id scores were lower for the patients' own tumor Id than for the mean of the allogeneic Ids (Table; Wilcoxon's matched pair test). This difference was mostly attributable to CDR-derived epitopes throughout all CDRs despite overall lower immunogenicity compared to FR. There was no evidence for differential immunogenicity between a hypermutated FL Id and the corresponding GL (p=0.58). Finally, a preliminary survey of IgH sequences from non-clonal B cells indicated similar immunogenicity compared to FL Id (p=0.31). These bioinformatic findings indicate T cell-mediated immunosurveillance against the BCR of malignant and perhaps nonmalignant B cells. T cell activity appears to be directed predominantly against individual CDR peptides despite their lesser predicted HLA binding capacity compared to FR peptides. Existing CDR epitopes are not generated during the hypermutation process of BCRs, raising the possibility that randomly generated, more immunogenic hypervariable peptides are not permitted by the immune system. In conjunction with the T cell activity observed in in vivo and in vitro studies cited above, these findings are consistent with strong peripheral tolerance to shared Id structures. On the other hand, T cell control of individual Id peptides may play a role in immunosurveillance of malignant B cells and may be exploited for active immunotherapy of lymphoma. In contrast, generic or pan-B-cell epitopes are predicted to be less effective in inducing anti-lymphoma T cell responses.Median (range) BIMASPatient IdMean of allogeneic IdscomparisonAll peptides213 (40-5920)369 (56-5520)p=0.0012FR peptides157 (20-5415)239 (18-3891)p=0.045CDR peptides74 (7-648)175 (21-1760)p<0.0001- CDR1 peptides21 (0.7-144)52 (1.9-630)p=0.0007- CDR2 peptides7.6 (0.2-345)30 (2.2-212)p=0.0089- CDR3 peptides16 (1.3-506)37 (6-980)p=0.0008 Disclosures: No relevant conflicts of interest to declare.

scholarly journals Co-potentiation of antigen recognition: A mechanism to boost weak T cell responses and provide immunotherapy in vivo

2015 ◽  
Vol 1 (9) ◽  
pp. e1500415 ◽  
Author(s):  
Michele M. Hoffmann ◽  
Carlos Molina-Mendiola ◽  
Alfreda D. Nelson ◽  
Christopher A. Parks ◽  
Edwin E. Reyes ◽  
...  
Keyword(s):  
T Cell ◽  
Lung Metastases ◽  
T Cell Responses ◽  
Cell Receptor ◽  
Antigen Recognition ◽  
Immune Reactivity ◽  
Antigen Receptors ◽  
Cell Responses

Adaptive immunity is mediated by antigen receptors that can induce weak or strong immune responses depending on the nature of the antigen that is bound. In T lymphocytes, antigen recognition triggers signal transduction by clustering T cell receptor (TCR)/CD3 multiprotein complexes. In addition, it hypothesized that biophysical changes induced in TCR/CD3 that accompany receptor engagement may contribute to signal intensity. Nonclustering monovalent TCR/CD3 engagement is functionally inert despite the fact that it may induce changes in conformational arrangement or in the flexibility of receptor subunits. We report that the intrinsically inert monovalent engagement of TCR/CD3 can specifically enhance physiologic T cell responses to weak antigens in vitro and in vivo without stimulating antigen-unengaged T cells and without interrupting T cell responses to strong antigens, an effect that we term as “co-potentiation.” We identified Mono-7D6-Fab, which biophysically altered TCR/CD3 when bound and functionally enhanced immune reactivity to several weak antigens in vitro, including a gp100-derived peptide associated with melanoma. In vivo, Mono-7D6-Fab induced T cell antigen–dependent therapeutic responses against melanoma lung metastases, an effect that synergized with other anti-melanoma immunotherapies to significantly improve outcome and survival. We conclude that Mono-7D6-Fab directly co-potentiated TCR/CD3 engagement by weak antigens and that such concept can be translated into an immunotherapeutic design. The co-potentiation principle may be applicable to other receptors that could be regulated by otherwise inert compounds whose latent potency is only invoked in concert with specific physiologic ligands.

scholarly journals Regulation of In Vitro and In Vivo Immune Functions by the Cytosolic Adaptor Protein SKAP-HOM

2005 ◽  
Vol 25 (18) ◽  
pp. 8052-8063 ◽  
Author(s):  
M. Togni ◽  
K. D. Swanson ◽  
S. Reimann ◽  
S. Kliche ◽  
A. C. Pearce ◽  
...  
Keyword(s):  
Immune System ◽  
T Cell ◽  
Physiological Role ◽  
Adaptor Protein ◽  
T Cell Responses ◽  
Specific Substrate ◽  
Immune Functions ◽  
Cell Responses

ABSTRACT SKAP-HOM is a cytosolic adaptor protein representing a specific substrate for the Src family protein tyrosine kinase Fyn. Previously, several groups have provided experimental evidence that SKAP-HOM (most likely in cooperation with the cytosolic adaptor protein ADAP) is involved in regulating leukocyte adhesion. To further assess the physiological role of SKAP-HOM, we investigated the immune system of SKAP-HOM-deficient mice. Our data show that T-cell responses towards a variety of stimuli are unaffected in the absence of SKAP-HOM. Similarly, B-cell receptor (BCR)-mediated total tyrosine phosphorylation and phosphorylation of Erk, p38, and JNK, as well as immunoreceptor-mediated Ca2+ responses, are normal in SKAP-HOM−/− animals. However, despite apparently normal membrane-proximal signaling events, BCR-mediated proliferation is strongly attenuated in the absence of SKAP-HOM−/−. In addition, adhesion of activated B cells to fibronectin (a ligand for β1 integrins) as well as to ICAM-1 (a ligand for β2 integrins) is strongly reduced. In vivo, the loss of SKAP-HOM results in a less severe clinical course of experimental autoimmune encephalomyelitis following immunization of mice with the encephalitogenic peptide of MOG (myelin oligodendrocyte glycoprotein). This is accompanied by strongly reduced serum levels of MOG-specific antibodies and lower MOG-specific T-cell responses. In summary, our data suggest that SKAP-HOM is required for proper activation of the immune system, likely by regulating the cross-talk between immunoreceptors and integrins.

Immunomodulating activity of meningococcal antigens

1983 ◽  
Vol 29 (12) ◽  
pp. 1611-1618 ◽  
Author(s):  
Brian G. Sparkes
Keyword(s):  
T Cell ◽  
B Cell ◽  
Outer Membrane Proteins ◽  
Cell Responses ◽  
T Cell Suppression ◽  
Cell Suppression ◽  
Cell Mitogen

A preparation of meningococcal antigens (MA) extracted in CaCl2, and containing mostly outer membrane proteins, was strongly mitogenic for murine B lymphocytes. Given to mice in vitro, MA markedly impaired subsequent in vivo T-cell responses of splenocytes. Suppression of normal T splenocytes in vitro occurred with both adherent (Ad) and nonadherent (NA) splenocytes from MA-sensitized mice. B cells were much less affected by the suppression induced by MA, and only Ad cells could convey in vitro the low level impairment of B-cell proliferation. Strong T-cell suppression associated with a B-cell mitogen is also produced by bacillus Calmette-Guérin (BCG) and Corynebacterium parvum. The possible role of these phenomena in meningococcal disease is discussed.

Antigen-loaded exosomes alone induce Th1-type memory through a B cell–dependent mechanism

Blood ◽  
2009 ◽  
Vol 113 (12) ◽  
pp. 2673-2683 ◽  
Author(s):  
Khaleda Rahman Qazi ◽  
Ulf Gehrmann ◽  
Emilie Domange Jordö ◽  
Mikael C. I. Karlsson ◽  
Susanne Gabrielsson
Keyword(s):  
Cell Proliferation ◽  
T Cell ◽  
B Cell ◽  
T Cell Responses ◽  
T Cell Response ◽  
T Cell Proliferation ◽  
Cell Responses ◽  
Proliferation In Vitro

Abstract Exosomes are nanovesicles harboring proteins important for antigen presentation. We compared the potency of differently loaded exosomes, directly loaded with OVA323-339 peptide (Pep-Exo) or exosomes from OVA-pulsed DCs (OVA-Exo), for their ability to induce specific T-cell proliferation in vitro and in vivo. Both Pep-Exo and OVA-Exo elicited specific transgenic T-cell proliferation in vitro, with the Pep-Exo being more efficient. In contrast, only OVA-Exo induced specific T-cell responses in vivo highlighting the importance of indirect loading strategies in clinical applications. Coadministration of whole OVA overcame the unresponsiveness with Pep-Exo but still elicited a lower response compared with OVA-Exo. In parallel, we found that OVA-Exo not only augmented the specific T-cell response but also gave a Th1-type shift and an antibody response even in the absence of whole OVA. We detected IgG2a and interferon-γ production from splenocytes showing the capability of exosomes to provide antigen for B-cell activation. Furthermore, we found that B cells are needed for exosomal T-cell stimulation because Bruton tyrosine kinase–deficient mice showed abrogated B- and T-cell responses after OVA-Exo immunization. These findings reveal that exosomes are potent immune regulators and are relevant for the design of vaccine adjuvants and therapeutic intervention strategies to modulate immune responses.

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