scholarly journals A B Cell Receptor with Two Igα Cytoplasmic Domains Supports Development of Mature But Anergic B Cells

2004 ◽  
Vol 199 (6) ◽  
pp. 855-865 ◽  
Author(s):  
Amy Reichlin ◽  
Anna Gazumyan ◽  
Hitoshi Nagaoka ◽  
Kathrin H. Kirsch ◽  
Manfred Kraus ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Cytoplasmic Domain ◽  
B Cell Receptor ◽  
B1a Cells ◽  
Bcr Signaling ◽  
Membrane Bound

B cell receptor (BCR) signaling is mediated through immunoglobulin (Ig)α and Igβ a membrane-bound heterodimer. Igα and Igβ are redundant in their ability to support early B cell development, but their roles in mature B cells have not been defined. To examine the function of Igα–Igβ in mature B cells in vivo we exchanged the cytoplasmic domain of Igα for the cytoplasmic domain of Igβ by gene targeting (Igβc→αc mice). Igβc→αc B cells had lower levels of surface IgM and higher levels of BCR internalization than wild-type B cells. The mutant B cells were able to complete all stages of development and were long lived, but failed to differentiate into B1a cells. In addition, Igβc→αc B cells showed decreased proliferative and Ca2+ responses to BCR stimulation in vitro, and were anergic to T-independent and -dependent antigens in vivo.

In Vivo Inhibition of BCR Activation in High-Risk CLL Patients On Therapy with Bruton's Tyrosine Kinase Inhibitor Ibrutinib: Correlative Studies from an Ongoing Phase 2 Clinical Trial

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 186-186
Author(s):  
Julia Hoellenriegel ◽  
Susan O'Brien ◽  
Michael J. Keating ◽  
William G. Wierda ◽  
Joseph J. Buggy ◽  
...  
Keyword(s):  
High Risk ◽  
Tyrosine Kinase ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Bcr Signaling ◽  
Correlative Studies ◽  
Pre Treatment

Abstract Abstract 186 B cell receptor (BCR) signaling is essential for normal B cell development and plays an important role in several B cell malignancies, including chronic lymphocytic leukemia (CLL). BrutonÕs tyrosine kinase (Btk) transmits B cell receptor (BCR) signaling and can be inhibited by ibrutinib, a selective, covalent Btk inhibitor. Because of highly encouraging results with ibrutinib in high-risk CLL patients in the Phase 1/2 trial, we explored the combination of ibrutinib and rituximab in forty high-risk CLL patients, characterized by the presence of 17p deletion or TP53 mutation (treated or untreated), previously treated patients with 11q deletion, or patients with a short remission duration (< 3 years) after first-line chemo-immunotherapy. Here, we present early correlative studies from this trial, focusing on BCR-related CLL responses (chemokine secretion, viability) and CLL cell migration, based on our recent experience with ibrutinib in preclinical CLL models (Ponader S et al., Blood 119:1182–9, 2012). The aim of this study was to determine if preclinical results of Btk inhibition with ibrutinib can be recapitulated in vivo with specimen from the high-risk population enrolled on this trial. Plasma levels of CLL3 and CCL4 (MIP-1α/β), two chemokines secreted by CLL cells in response to BCR activation, were assessed before, at 14, and at 28 days on treatment with ibrutinib. In 28 analyzed patients, we demonstrate robust, significant reductions in CCL3 and CCL4 plasma concentrations after 14 and 28 days of treatment. As shown in Figure 1, plasma CCL3 levels were reduced from 139.6 (± 40.4) pg/mL before treatment to 6.9 (± 0.9) pg/mL or 6.5 (± 0.7) pg/mL after 14 and 28 days of treatment, respectively. We also evaluated in vitro secretion of CCL3 and CCL4 into supernatants of CLL cells isolated from 12 patients before and during ibrutinib therapy in response to stimulation with anti-IgM. Compared to CLL cells from pre-treatment specimen, CLL cells from patients on Ibrutinib therapy showed reduced levels of CLL3 and CCL4 secretion, and additional treatment with Ibrutinib (0.5 μM – 1μM) led to reduced chemokine levels only in pre-treatment samples, indicating complete Btk target inhibition. We next evaluated the effect of ibrutinib on CLL cell viability after anti-IgM stimulation. In pre-treatment samples, ibrutinib abrogated BCR-triggered CLL cell survival. Surprisingly, CLL cells from ibrutinib-treated patients remained anti-IgM responsive in these viability assays. These Btk-independent pro-survival effects could not be inhibited by in vitro treatment with ibrutinib, indicating that some of the anti-IgM-triggered pro-survival signaling can bypass Btk. Next, we analyzed migration of CLL cells towards the chemokines CXCL12 and CXCL13 in transwell chemotaxis assays. Pre-treatment samples displayed significant higher chemotaxis towards CXCL12 and CXCL13 when compared to CLL samples from patients on therapy with ibrutinib. The mean relative migration of such samples toward CXCL12 or CXCL13 was reduced to 28% (± 5%) or 35% (±15%) of respective CLL cells isolated before ibrutinib therapy (100%), n=6. Collectively, our results demonstrate that Ibrutinib blocks BCR-dependent survival and migration responses in high-risk CLL patients in vivo. They also corroborate the validity and robustness of CCL3 and CCL4 as biomarkers for BCR inhibition in CLL patients. Figure 1 Figure 1. Disclosures: O'Brien: Pharmacyclics: Research support Other. Buggy:Pharmacyclics: Employment, Equity Ownership. Burger:Pharmacyclics: Consultancy, Research Funding.

scholarly journals Igβ tyrosine residues contribute to the control of B cell receptor signaling by regulating receptor internalization

2006 ◽  
Vol 203 (7) ◽  
pp. 1785-1794 ◽  
Author(s):  
Anna Gazumyan ◽  
Amy Reichlin ◽  
Michel C. Nussenzweig
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Sh2 Domain ◽  
B Cell Receptor ◽  
Receptor Internalization ◽  
Bcr Signaling ◽  
B Cell Receptor Signaling ◽  
B1 Cells

Immunoglobulin (Ig)α and Igβ initiate B cell receptor (BCR) signaling through immune receptor tyrosine activation motifs (ITAMs) that are targets of SH2 domain–containing kinases. To examine the function of Igβ ITAM tyrosine resides in mature B cells in vivo, we exchanged these residues for alanine by gene targeting (IgβAA). Mutant mice showed normal development of all B cell subtypes with the exception of B1 cells that were reduced by fivefold. However, primary B cells purified from IgβAA mice showed significantly decreased steady-state and ligand-mediated BCR internalization and higher levels of cell surface IgM and IgD. BCR cross-linking resulted in decreased Src and Syk activation but paradoxically enhanced and prolonged BCR signaling, as measured by cellular tyrosine phosphorylation, Ca++ flux, AKT, and ERK activation. In addition, B cells with the ITAM mutant receptor showed an enhanced response to a T-independent antigen. Thus, Igβ ITAM tyrosines help set BCR signaling threshold by regulating receptor internalization.

B-cell receptor signaling in chronic lymphocytic leukemia

Blood ◽  
2011 ◽  
Vol 118 (16) ◽  
pp. 4313-4320 ◽  
Author(s):  
Freda K. Stevenson ◽  
Sergey Krysov ◽  
Andrew J. Davies ◽  
Andrew J. Steele ◽  
Graham Packham
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Clinical Activity ◽  
Variable Regions ◽  
Bcr Signaling

Abstract The B-cell receptor (BCR) is a key survival molecule for normal B cells and for most B-cell malignancies. Recombinatorial and mutational patterns in the clonal immunoglobulin (Ig) of chronic lymphocytic leukemia (CLL) have revealed 2 major IgMD-expressing subsets and an isotype-switched variant, each developing from distinct B-cell populations. Tracking of conserved stereotypic features of Ig variable regions characteristic of U-CLL indicate circulating naive B cells as the likely cells of origin. In CLL, engagement of the BCR by antigen occurs in vivo, leading to down-regulated expression and to an unanticipated modulation of glycosylation of surface IgM, visible in blood cells, especially in U-CLL. Modulated glycoforms of sIgM are signal competent and could bind to environmental lectins. U-CLL cases express more sIgM and have increased signal competence, linking differential signaling responses to clinical behavior. Mapping of BCR signaling pathways identifies targets for blockade, aimed to deprive CLL cells of survival and proliferative signals. New inhibitors of BCR signaling appear to have clinical activity. In this Perspective, we discuss the functional significance of the BCR in CLL, and we describe strategies to target BCR signaling as an emerging therapeutic approach.

scholarly journals Subversion of Pkca and Pkcbii Upregulation Play an Essential Role in Chronic Lymphocytic Leukaemia Development, Inducing Constitutive B Cell Receptor-Mediated Signals

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 488-488
Author(s):  
Anuradha Tarafdar ◽  
Ashfia Fatima Khan ◽  
Emilio Cosimo ◽  
Hothri A. Moka ◽  
Karen Dunn ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukaemia ◽  
B Cell ◽  
Progenitor Cells ◽  
Essential Role ◽  
Lymphocytic Leukaemia ◽  
Cell Receptor ◽  
B Cell Receptor

Abstract B cell receptor (BCR)-mediated signals orchestrate key events during the life cycle of B lymphocytes enabling normal B cell development and maturation. Chronic lymphocytic leukaemia (CLL), an incurable malignancy of mature B cells, displays deregulated BCR-mediated signalling, the intensity of which correlates disease pathogenesis. As such, signals generated upon BCR engagement represent promising targets for novel therapies. Of note, the expression profile of selected protein kinase C (PKC) isoforms, which link proximal BCR mediated signals with downstream pathways, exhibit altered expression patterns in CLL cells: upregulation of PKCβII, PKCε, PKCζ and downregulation of PKCα and PKCβI compared with normal B cells. We previously demonstrated that introduction of a kinase inactive PKCα (PKCα-KR) construct in mouse lymphoid progenitor cells resulted in development of a CLL-like disease both in vitro and in vivo. This model resembles the more aggressive subset of CLL, exhibiting an upregulation of ZAP-70 and elevated ERK-MAPK-mTOR signalling resulting in enhanced proliferation and increased tumor load in the lymphoid organs. Interestingly, reduced PKCα function resulted in PKCβII upregulation, a key pathogenic feature of CLL. Inhibition of PKCβII in these cells with enzastaurin resulted in cell cycle arrest in vitro, reduced tumour load and elevated apoptosis in vivo indicating that PKCβII plays a vital role in maintaining cell survival in our model. To further elucidate the role of PKCβ in leukaemogenesis, we have performed sequential prkcb knockdown (KD) and PKCα-KR expression in the lymphoid progenitor cells. prkcb KD resulted in reduced proliferation and survival concurrent with reduced expression of leukaemic markers (CD23 and CD5) compared to control cells indicating that prkcb plays an essential role in initiation of leukaemogenesis in our model. To identify targets responsible for the regulation for prkcb transcription we performed global gene analysis (Affymetrix GeneChip mouse gene 1.0 ST) comparing MIEV (empty vector control) and PKCα-KR transduced cells at successive time-points mapping critical stages of B-cell transformation, pre- and post PKCβII upregulation. MetaCoreTM analysis revealed that upon upregulation of PKCβII, the BCR-mediated signalling pathway was significantly upregulated in PKCα-KR expressing cells. At the protein level, key hubs proximal to the BCR - Lyn, Btk and Akt - were upregulated, indicating constitutive activation of BCR signalling in the CLL-like PKCα-KR expressing cells. Additionally, proliferative signals downstream of the BCR (mTOR, NF-kB and c-myc) were also upregulated. Treatment of PKCα-KR expressing cells with the Btk inhibitor Ibrutinib (PCI-32765), which has recently been approved for the treatment of CLL, reduced cellular proliferation and inhibited phosphorylation of BtkY223, AktS473 and S6S235/236. Overall, we demonstrate that PKCβII expression is essential for leukaemogenesis and identify key signalling pathways that drive the initiation/development of CLL in the PKCα-KR model. Disclosures No relevant conflicts of interest to declare.

scholarly journals Genome-wide screens identify calcium signaling as a key regulator of IgE+ plasma cell differentiation and survival

2021 ◽  
Author(s):  
Rebecca Newman ◽  
Pavel Tolar
Keyword(s):  
B Cells ◽  
Calcium Signaling ◽  
B Cell ◽  
Plasma Cells ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Genome Wide ◽  
Antibody Isotypes ◽  
Bcr Signaling

SummaryIgE antibodies protect against toxins and parasites, however, they also mediate allergic reactions. In contrast to other antibody isotypes, B cells switched to IgE respond transiently and do not give rise to long-lived plasma cells (PCs) or memory B cells. Although the intrinsic differences of IgE+ B cells have been linked to signaling by the IgE-B cell receptor (BCR), the molecular pathways controlling their behavior remain poorly understood. Here we employ whole-genome CRISPR screening to identify genes regulating IgE+ B cell proliferation, survival and differentiation into PCs. We show that IgE+ B cells are selectively suppressed by the IgE-BCR signaling to intracellular calcium, which inhibits PC differentiation and limits their lifespan after differentiation. Consequently, manipulation of calcium signaling in vivo enhances IgE+ PC responses. Insights from this pathway shed new light on the self-limiting character of IgE responses and open new avenues to eliminate IgE+ PCs in allergy.

Chronic lymphocytic leukemia: revelations from the B-cell receptor

Blood ◽  
2004 ◽  
Vol 103 (12) ◽  
pp. 4389-4395 ◽  
Author(s):  
Freda K. Stevenson ◽  
Federico Caligaris-Cappio
Keyword(s):  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Cell Of Origin ◽  
Regulatory B Cell ◽  
V Genes

Abstract The finding that chronic lymphocytic leukemia (CLL) consists of 2 clinical subsets, distinguished by the incidence of somatic mutations in the immunoglobulin (Ig) variable region (V) genes, has clearly linked prognosis to biology. Antigen encounter by the cell of origin is indicated in both subsets by selective but distinct expression of V genes, with evidence for continuing stimulation after transformation. The key to distinctive tumor behavior likely relates to the differential ability of the B-cell receptor (BCR) to respond. Both subsets may be undergoing low-level signaling in vivo, although analysis of blood cells limits knowledge of critical events in the tissue microenvironment. Analysis of signal competence in vitro reveals that unmutated CLL generally continues to respond, whereas mutated CLL is anergized. Differential responsiveness may reflect the increased ability of post-germinal center B cells to be triggered by antigen, leading to long-term anergy. This could minimize cell division in mutated CLL and account for prognostic differences. Unifying features of CLL include low responsiveness, expression of CD25, and production of immunosuppressive cytokines. These properties are reminiscent of regulatory T cells and suggest that the cell of origin of CLL might be a regulatory B cell. Continuing regulatory activity, mediated via autoantigen, could suppress Ig production and lead to disease-associated hypogammaglobulinemia. (Blood. 2004;103:4389-4395)

scholarly journals Rewiring of B cell receptor signaling by Epstein–Barr virus LMP2A

2020 ◽  
Vol 117 (42) ◽  
pp. 26318-26327
Author(s):  
Kamonwan Fish ◽  
Federico Comoglio ◽  
Arthur L. Shaffer ◽  
Yanlong Ji ◽  
Kuan-Ting Pan ◽  
...  
Keyword(s):  
Cell Cycle ◽  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Barr Virus ◽  
Human B Cells ◽  
Bcr Signaling ◽  
Epstein Barr

Epstein–Barr virus (EBV) infects human B cells and reprograms them to allow virus replication and persistence. One key viral factor in this process is latent membrane protein 2A (LMP2A), which has been described as a B cell receptor (BCR) mimic promoting malignant transformation. However, how LMP2A signaling contributes to tumorigenesis remains elusive. By comparing LMP2A and BCR signaling in primary human B cells using phosphoproteomics and transcriptome profiling, we identified molecular mechanisms through which LMP2A affects B cell biology. Consistent with the literature, we found that LMP2A mimics a subset of BCR signaling events, including tyrosine phosphorylation of the kinase SYK, the calcium initiation complex consisting of BLNK, BTK, and PLCγ2, and its downstream transcription factor NFAT. However, the majority of LMP2A-induced signaling events markedly differed from those induced by BCR stimulation. These included differential phosphorylation of kinases, phosphatases, adaptor proteins, transcription factors such as nuclear factor κB (NF-κB) and TCF3, as well as widespread changes in the transcriptional output of LMP2A-expressing B cells. LMP2A affected apoptosis and cell-cycle checkpoints by dysregulating the expression of apoptosis regulators such as BCl-xL and the tumor suppressor retinoblastoma-associated protein 1 (RB1). LMP2A cooperated with MYC and mutant cyclin D3, two oncogenic drivers of Burkitt lymphoma, to promote proliferation and survival of primary human B cells by counteracting MYC-induced apoptosis and by inhibiting RB1 function, thereby promoting cell-cycle progression. Our results indicate that LMP2A is not a pure BCR mimic but rather rewires intracellular signaling in EBV-infected B cells that optimizes cell survival and proliferation, setting the stage for oncogenic transformation.

scholarly journals A Gammaherpesvirus Bcl-2 Ortholog Blocks B Cell Receptor-Mediated Apoptosis and Promotes the Survival of Developing B Cells In Vivo

2014 ◽  
Vol 10 (2) ◽  
pp. e1003916 ◽  
Author(s):  
Carrie B. Coleman ◽  
Jennifer E. McGraw ◽  
Emily R. Feldman ◽  
Alexa N. Roth ◽  
Lisa R. Keyes ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor

scholarly journals Epstein-Barr Virus LMP2A Alters In Vivo and In Vitro Models of B-Cell Anergy, but Not Deletion, in Response to Autoantigen

2005 ◽  
Vol 79 (12) ◽  
pp. 7355-7362 ◽  
Author(s):  
Michelle A. Swanson-Mungerson ◽  
Robert G. Caldwell ◽  
Rebecca Bultema ◽  
Richard Longnecker
Keyword(s):  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Nuclear Translocation ◽  
Cell Receptor ◽  
Barr Virus ◽  
Low Levels ◽  
Epstein Barr

ABSTRACT A significant percentage of the population latently harbors Epstein-Barr virus (EBV) in B cells. One EBV-encoded protein, latent membrane protein 2A (LMP2A), is expressed in tissue culture models of EBV latent infection, in human infections, and in many of the EBV-associated proliferative disorders. LMP2A constitutively activates proteins involved in the B-cell receptor (BCR) signal transduction cascade and inhibits the antigen-induced activation of these proteins. In the present study, we investigated whether LMP2A alters B-cell receptor signaling in primary B cells in vivo and in vitro. LMP2A does not inhibit antigen-induced tolerance in response to strong stimuli in an in vivo tolerance model in which B cells are reactive to self-antigen. In contrast, LMP2A bypasses anergy induction in response to low levels of soluble hen egg lysozyme (HEL) both in vivo and in vitro as determined by the ability of LMP2A-expressing HEL-specific B cells to proliferate and induce NF-κB nuclear translocation after exposure to low levels of antigen. Furthermore, LMP2A induces NF-κB nuclear translocation independent of BCR cross-linking. Since NF-κB is required to bypass tolerance induction, this LMP2A-dependent NF-κB activation may complete the tolerogenic signal induced by low levels of soluble HEL. Overall, the findings suggest that LMP2A may not inhibit BCR-induced signals under all conditions as previously suggested by studies with EBV immortalized B cells.

Rituximab inhibits B-cell receptor signaling

Blood ◽  
2010 ◽  
Vol 115 (5) ◽  
pp. 985-994 ◽  
Author(s):  
Samar Kheirallah ◽  
Pierre Caron ◽  
Emilie Gross ◽  
Anne Quillet-Mary ◽  
Justine Bertrand-Michel ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Mammalian Target Of Rapamycin ◽  
Cell Receptor ◽  
Inhibitory Effect ◽  
B Cell Receptor ◽  
B Cell Survival ◽  
Bcr Signaling ◽  
Dependent Inhibition ◽  
Lipid Raft Microdomains

Abstract Rituximab (RTX), a monoclonal antibody directed against the CD20 protein, is a drug commonly used in the treatment of B-cell–derived lymphoid neoplasias and of antibody-mediated autoimmune diseases. In addition to cell- and complement-mediated B-cell depletion, RTX is thought to inhibit B-cell survival and proliferation through negative regulation of canonical signaling pathways involving Akt, ERK, and mammalian target of rapamycin. However, surprisingly, although B-cell receptor (BCR) signaling has been considered critical for normal and more recently, for neoplastic B cells, the hypothesis that RTX could target BCR has never been investigated. Using follicular lymphoma cell lines as models, as well as normal B cells, we show here, for the first time, that pretreatment with RTX results in a time-dependent inhibition of the BCR-signaling cascade involving Lyn, Syk, PLCγ2, Akt, and ERK, and calcium mobilization. The inhibitory effect of RTX correlates with decrease of raft-associated cholesterol, complete inhibition of BCR relocalization into lipid raft microdomains, and down-regulation of BCR immunoglobulin expression. Thus, RTX-mediated alteration of BCR expression, dynamics, and signaling might contribute to the immunosuppressive activity of the drug.

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