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Takeshi Tsubata - One of the best experts on this subject based on the ideXlab platform.
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Inhibitory B cell co-receptors and autoimmune diseases.
Immunological medicine, 2019Co-Authors: Takeshi TsubataAbstract:B cells express various inhibitory co-receptors including CD22 (also known as Siglec-2), Siglec-10 (Siglec-G in mice), CD72, LILRB (PIR-B in mice) and FcγRIIB that contain immunoreceptor tyrosine-based inhibition motifs (ITIMs) in the cytoplasmic region and negatively regulate BCR signaling by recruiting phosphatases to the ITIMs. Some of the inhibitory B cell co-receptors suppress development of SLE. Among these, CD72 most strongly regulates SLE. CD72 recognizes Sm/RNP, a lupus self-antigen and an endogenous TLR7 ligand, as a specific ligand, and suppresses B cell response to this TLR7 ligand. This suppression may inhibit development of SLE because TLR7 is indispensable in multiple mouse SLE models. In contrast, inhibitory B cell co-receptors such as CD22 and CD72 inhibit expansion of regulatory B cells that are known to regulate development of autoimmune diseases including type 1 diabetes (T1D) and multiple sclerosis. CD72 strongly exacerbate development of T1D in NOD mice probably by limiting expansion of regulatory B cells. Thus, inhibitory B cell co-receptors especially CD72 regulates distinct autoimmune diseases either positively or negatively. As B cell depletion therapy clearly reveals crucial roles of B cells in the regulation of various autoimmune diseases, CD72 may be a novel therapeutic target for treatment of autoimmune diseases.
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Ligand Recognition Determines the Role of Inhibitory B Cell Co-receptors in the Regulation of B Cell Homeostasis and Autoimmunity.
Frontiers in immunology, 2018Co-Authors: Takeshi TsubataAbstract:B cells express various inhibitory co-receptors including CD22, CD72, and Siglec-G. These receptors contain immunoreceptor tyrosine-based inhibition motifs (ITIMs) in the cytoplasmic region. Although many of the inhibitory co-receptors negatively regulate BCR signaling by activating SH2-containing protein tyrosine phosphatase 1 (SHP-1), different inhibitory co-receptors have distinct functional properties. CD22, Siglec-G, and CD72 preferentially regulate tonic signaling in conventional B cells, B-1 cell homeostasis, and development of lupus-like disease, respectively. CD72 recognizes RNA-related lupus self-antigen Sm/RNP as a ligand. This ligand recognition recruits CD72 to BCR in Sm/RNP-reactive B cells thereby suppressing production of anti-Sm/RNP autoantibody involved in the pathogenesis of lupus. In contrast, Siglec-G recognizes α2,3 as well as α2,6 sialic acids whereas CD22 recognizes α2,6 sialic acid alone. Because glycoproteins including BCR are dominantly glycosylated with α2,3 sialic acids in B-1 cells, Siglec-G but not CD22 recruits BCR as a ligand specifically in B-1 cells, and regulates B-1 cell homeostasis by suppressing BCR signaling in B-1 cells. Thus, recognition of distinct ligands determines functional properties of different inhibitory B cell co-receptors.
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Human CD72 splicing isoform responsible for resistance to systemic lupus erythematosus regulates serum immunoglobulin level and is localized in endoplasmic reticulum
BMC immunology, 2012Co-Authors: Yuki Hitomi, Takahiro Adachi, Naoyuki Tsuchiya, Zen-ichiro Honda, Katsushi Tokunaga, Takeshi TsubataAbstract:Background CD72 is an inhibitory co-receptor expressed on B cells. We previously demonstrated significant association of the polymorphism of the CD72 gene with susceptibility to human systemic lupus erythematosus (SLE) in individuals carrying a SLE-susceptible FCGR2B genotype (FCGR2B-232Thr/Thr). The human CD72 locus generates a splicing isoform that lacks exon 8 (CD72Δex8) as well as full-length CD72 (CD72fl), and the CD72 polymorphism regulates exon 8 skipping.
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Centromeric interval of chromosome 4 derived from C57BL/6 mice accelerates type 1 diabetes in NOD.CD72b congenic mice.
Biochemical and biophysical research communications, 2009Co-Authors: Rong Hou, Takahiro Adachi, Mareki Ohtsuji, Naomi Ohtsuji, Li Zhang, Sachiko Hirose, Takeshi TsubataAbstract:Abstract The nonobese diabetic (NOD) mouse is a useful model of autoimmune type 1 diabetes exhibiting many similarities to human type 1 diabetes patients including the presence of auto-reactive T cells and pancreas-specific autoantiboies. Multiple Idd loci control the development of diabetes in NOD mice. CD72, a B cell membrane-bound glycoprotein carrying a C-type lectin-like domain, is an inhibitory co-receptor of the B cell antigen receptor (BCR) that negatively regulates BCR signaling. Among four known haplotypes of mouse CD72, NOD mice carry the CD72c haplotype, whereas most of the other inbred strains of mice carry either CD72a or CD72b. In this study, we generated congenic NOD.CD72b mice that carry C57BL/6 (B6) mouse-derived centromeric chromosome 4 interval (24–45 cM) surrounding the CD72b locus. Unexpectedly, NOD.CD72b mice were not protected from diabetes, but rather exhibited accelerated development of both insulitis and diabetes. Our result defines novel locus or loci in the vicinity of CD72 gene that negatively control diabetes, indicating that NOD disease is under complex genetic controls of not only Idd genes but also disease-resistant genes.
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Molecular interactions regulate BCR signal inhibition by CD22 and CD72
Trends in immunology, 2004Co-Authors: Lars Nitschke, Takeshi TsubataAbstract:The inhibitory coreceptors CD22 and CD72 downmodulate B-cell receptor (BCR) signaling and function as a molecular switch, determining whether antigen-stimulated B cells undergo apoptosis or proliferation. These coreceptors carry an intrinsic property for associating with the BCR, and this association is crucial for the initiation of signal inhibition through phosphorylation of these coreceptors by BCR-associated kinases. Recent findings have demonstrated that signal inhibition by these coreceptors is regulated by ligands for the coreceptors and by molecules binding to the coreceptors or the BCR. Moreover, signal inhibition by CD22 depends on the BCR isotype. These findings suggest a dynamic regulation of these coreceptors through molecular interactions on the B-cell surface.
Jane R. Parnes - One of the best experts on this subject based on the ideXlab platform.
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Modulation of Peripheral B Cell Tolerance by CD72 in a Murine Model
Arthritis and rheumatism, 2008Co-Authors: Monte M. Winslow, Thai M. Cao, Albert Chen, Corrine R. Davis, Elizabeth D. Mellins, Paul J. Utz, Gerald R. Crabtree, Jane R. ParnesAbstract:B cell self tolerance is maintained by several mechanisms, including deletion, receptor editing, and anergy (1–4). Mechanisms of peripheral tolerance are thought to be important, because a considerable proportion of self-reactive B cells escape central tolerance mechanisms and emerge into the periphery (5). Breakdown of peripheral B cell tolerance results in autoantibody production, which contributes to autoimmune diseases such as systemic lupus erythematosus (SLE), multiple sclerosis, Sjogren's syndrome, and rheumatoid arthritis. SLE is an autoimmune disorder characterized by multiorgan inflammation and deregulated production of autoantibodies, including antinuclear antibodies (ANAs) and anti–double-stranded DNA (anti-dsDNA) antibodies (6). B cell–targeted therapies have shown great promise in the treatment of SLE (7). B cell coreceptors, such as CD19, CD22, CD72, Fcγ receptor type IIb (FcγRIIb), CD45, and CD5, tightly regulate B cell receptor (BCR) signaling to fine-tune B cell responses in encounters with antigens in the periphery (8,9). In fact, several of these coreceptors, including CD22, FcγRIIb, and CD45, have been directly linked to SLE (10–13). CD72, a 45-kd type II transmembrane glycoprotein, is constitutively expressed on developing and mature B cells, but not on terminally differentiated plasma cells. The cytoplasmic domain of CD72 contains an immunoreceptor tyrosine–based inhibition motif (ITIM) that binds SH2-containing protein tyrosine phosphatase 1 (SHP-1), and an ITIM-like motif that binds Grb2 (14). Our recent data indicate that CD72 plays a negative role in regulating BCR-induced signaling in primary mature B cells (9), but CD72 may also transmit a positive signal independent of the BCR (15). CD100, a ligand for CD72, can turn off the negative effect of CD72 by inhibiting phosphorylation of CD72 and, in turn, disrupting the SHP-1/CD72 complex (16,17). Although a link between CD72 and SLE has been suggested (18–20), the role of CD72 in autoimmunity is not well defined and the molecular mechanisms by which CD72 might influence B cell tolerance are unknown. Cbl-b, an E3 ubiquitin ligase, has been shown to regulate B cell tolerance. Cbl-b–deficient B cells are hyperreactive to BCR ligation, and mice lacking Cbl-b produce anti-dsDNA autoantibodies (21). Clinically, Cbl-b–deficient mice are susceptible to spontaneous and induced autoimmune diseases (22–24), and mice with B cell–specific ablation of both Cbl and Cbl-b manifest SLE-like autoimmune disease (25), suggesting that Cbl-b modulates peripheral T cell and B cell tolerance. However, the proteins that anchor Cbl-b to the membrane in close proximity to the BCR signalosome and that regulate Cbl-b function in B cells have not been identified. In the present study we examined the direct role of CD72 in peripheral B cell tolerance and in the development of autoimmunity. Our findings indicate that CD72 is an essential negative regulator of BCR signaling in self-reactive B cells, and that CD72 interacts with and regulates Cbl-b. As indicated by multiple criteria, CD72 functions as a physiologic regulator of B cell anergy, and inappropriate regulation of the CD72-signaling network may contribute to the development of autoimmune diseases such as SLE.
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a unique CD72 epitope suggests a potential interaction with fcγrii cd32 on b lineage lymphocytes
Hybridoma, 2006Co-Authors: Yoshio Yamashita, Jane R. Parnes, Hyewon Phee, Kim-sue R.s. Tudor, Maria Isabel D. Rossi, Mark K Coggeshall, Paul W. KincadeAbstract:It has long been known that ligation of the transmembrane CD72 glycoprotein delivers signals to B lymphocytes, with the outcome depending on context. Of particular interest is its ability to function as a counter-receptor/ ligand for the CD100 semaphorin protein. We have now obtained evidence that CD72 physically interacts on the lymphocyte membrane with Fcγ receptor II (CD32). The association was first revealed with a new monoclonal antibody that recognizes polymorphic determinants on murine CD72. Although the specificity for CD72 was clear from immunoblotting, transfection and other experiments, staining with this reagent was inhibited when cells were pretreated with an Fc receptor-blocking antibody (CD16/CD32 specific). Furthermore, confocal microscopy revealed that the two molecules co-distributed on viable B cells. We also used the antibody to determine when CD72 becomes available to maturing lymphocytes. The marker is first acquired as large pre-B cells and enter the IL-7 independent phase of matura...
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CD72 Down-Modulates BCR-Induced Signal Transduction and Diminishes Survival in Primary Mature B Lymphocytes
Journal of immunology (Baltimore Md. : 1950), 2006Co-Authors: James W. Tung, Ingo H. Tarner, Andrew L. Snow, Tsuyoshi Yukinari, Rachel Ngernmaneepothong, Olivia M. Martinez, Jane R. ParnesAbstract:CD72, a 45-kDa type II transmembrane glycoprotein carrying an ITIM motif, is believed to be an inhibitory coreceptor of the BCR. Mature B cells lacking CD72 show enhanced Ca2+ mobilization and are hyperproliferative in response to BCR ligation. However, the signal transduction pathways downstream of BCR signaling that transmit the inhibitory effect of CD72 in mature B cells remain unknown. To address this question, we used hen egg lysozyme-specific BCR transgenic mice to elucidate the differential cell signaling between wild-type and CD72-deficient B cells in response to hen egg lysozyme Ag stimulation. Our results demonstrate that CD72 predominantly down-regulates the major signal transduction pathways downstream of the BCR, including NF-AT, NF-κB, ERK, JNK, p38-MAPK, and PI3K/Akt in mature B cells. CD72 ligation with anti-CD72 Ab (K10.6), which mimics the binding of CD100 (a natural ligand for CD72) to release the inhibitory function of CD72, augments cell proliferation, Ca2+ flux, IκBα activation, and ERK MAPK activity upon Ag stimulation in wild-type B cells. In addition, we show direct evidence that CD72 promotes cell cycle arrest and apoptosis after Ag stimulation in mature B cells. Taken together, our findings conclude that CD72 plays a dominant role as a negative regulator of BCR signaling in primary mature B lymphocytes.
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A Unique CD72 Epitope Suggests a Potential Interaction with FcγRII/CD32 on B Lineage Lymphocytes
Hybridoma (2005), 2006Co-Authors: Yoshio Yamashita, Jane R. Parnes, Hyewon Phee, Kim-sue R.s. Tudor, Maria Isabel D. Rossi, K. Mark Coggeshall, Paul W. KincadeAbstract:It has long been known that ligation of the transmembrane CD72 glycoprotein delivers signals to B lymphocytes, with the outcome depending on context. Of particular interest is its ability to function as a counter-receptor/ ligand for the CD100 semaphorin protein. We have now obtained evidence that CD72 physically interacts on the lymphocyte membrane with Fcγ receptor II (CD32). The association was first revealed with a new monoclonal antibody that recognizes polymorphic determinants on murine CD72. Although the specificity for CD72 was clear from immunoblotting, transfection and other experiments, staining with this reagent was inhibited when cells were pretreated with an Fc receptor-blocking antibody (CD16/CD32 specific). Furthermore, confocal microscopy revealed that the two molecules co-distributed on viable B cells. We also used the antibody to determine when CD72 becomes available to maturing lymphocytes. The marker is first acquired as large pre-B cells and enter the IL-7 independent phase of matura...
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CD72-deficient mice reveal nonredundant roles of CD72 in B cell development and activation.
Immunity, 1999Co-Authors: Chin Pan, Nicole Baumgarth, Jane R. ParnesAbstract:Abstract CD72, a B cell surface protein of the C-type lectin superfamily, recruits the tyrosine phosphatase SHP-1 through its ITIM motif(s). Using CD72-deficient ( CD72 −/− ) mice, we demonstrate that CD72 is a nonredundant regulator of B cell development. In the bone marrow of CD72 −/− mice, there was a reduction in the number of mature recirculating B cells and an accumulation of pre-B cells. In the periphery of CD72 −/− mice, there were fewer mature B-2 cells and more B-1 cells. In addition, CD72 is a negative regulator of B cell activation, as CD72 −/− B cells were hyperproliferative in response to various stimuli and showed enhanced kinetics in their intracellular Ca 2+ response following IgM cross-linking.
Takahiro Adachi - One of the best experts on this subject based on the ideXlab platform.
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CD72 negatively regulates B lymphocyte responses to the lupus-related endogenous toll-like receptor 7 ligand Sm/RNP.
The Journal of experimental medicine, 2016Co-Authors: Chizuru Akatsu, Takahiro Adachi, Kenro Shinagawa, Nobutaka Numoto, Zhihong Liu, Ayse Konuskan Ucar, Mohammad Aslam, Shirly Phoon, Koji Furukawa, Nobutoshi ItoAbstract:Toll-like receptor 7 (TLR7) plays an essential role in development of systemic lupus erythematosus by co-stimulating B cells reactive to the endogenous TLR7 ligand Sm/ribonucleoprotein (RNP), a crucial lupus self-antigen. However, how the TLR7-mediated autoimmune response is regulated is not yet known. In this study, we demonstrate that CD72, an inhibitory B cell co-receptor known to prevent development of lupus, recognizes Sm/RNP at the extracellular C-type lectin-like domain (CTLD) and specifically inhibits B cell response to Sm/RNP. Moreover, the CTLD of CD72c , a lupus-susceptible allele, binds to Sm/RNP less strongly than that of lupus-resistant CD72a . Reduced binding of CD72c is supported by x-ray crystallographic analysis that reveals a considerable alteration in charge at the putative ligand-binding site. Thus, CD72 appears to specifically inhibit B cell response to the endogenous TLR7 ligand Sm/RNP through CTLD-mediated recognition of Sm/RNP, thereby preventing production of anti-Sm/RNP antibody crucial for development of lupus.
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Human CD72 splicing isoform responsible for resistance to systemic lupus erythematosus regulates serum immunoglobulin level and is localized in endoplasmic reticulum
BMC immunology, 2012Co-Authors: Yuki Hitomi, Takahiro Adachi, Naoyuki Tsuchiya, Zen-ichiro Honda, Katsushi Tokunaga, Takeshi TsubataAbstract:Background CD72 is an inhibitory co-receptor expressed on B cells. We previously demonstrated significant association of the polymorphism of the CD72 gene with susceptibility to human systemic lupus erythematosus (SLE) in individuals carrying a SLE-susceptible FCGR2B genotype (FCGR2B-232Thr/Thr). The human CD72 locus generates a splicing isoform that lacks exon 8 (CD72Δex8) as well as full-length CD72 (CD72fl), and the CD72 polymorphism regulates exon 8 skipping.
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Centromeric interval of chromosome 4 derived from C57BL/6 mice accelerates type 1 diabetes in NOD.CD72b congenic mice.
Biochemical and biophysical research communications, 2009Co-Authors: Rong Hou, Takahiro Adachi, Mareki Ohtsuji, Naomi Ohtsuji, Li Zhang, Sachiko Hirose, Takeshi TsubataAbstract:Abstract The nonobese diabetic (NOD) mouse is a useful model of autoimmune type 1 diabetes exhibiting many similarities to human type 1 diabetes patients including the presence of auto-reactive T cells and pancreas-specific autoantiboies. Multiple Idd loci control the development of diabetes in NOD mice. CD72, a B cell membrane-bound glycoprotein carrying a C-type lectin-like domain, is an inhibitory co-receptor of the B cell antigen receptor (BCR) that negatively regulates BCR signaling. Among four known haplotypes of mouse CD72, NOD mice carry the CD72c haplotype, whereas most of the other inbred strains of mice carry either CD72a or CD72b. In this study, we generated congenic NOD.CD72b mice that carry C57BL/6 (B6) mouse-derived centromeric chromosome 4 interval (24–45 cM) surrounding the CD72b locus. Unexpectedly, NOD.CD72b mice were not protected from diabetes, but rather exhibited accelerated development of both insulitis and diabetes. Our result defines novel locus or loci in the vicinity of CD72 gene that negatively control diabetes, indicating that NOD disease is under complex genetic controls of not only Idd genes but also disease-resistant genes.
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Inhibitory Coreceptors Activated by Antigens But Not by Anti-Ig Heavy Chain Antibodies Install Requirement of Costimulation Through CD40 for Survival and Proliferation of B Cells
Journal of immunology (Baltimore Md. : 1950), 2003Co-Authors: Yasuhisa Hokazono, Takahiro Adachi, Nobuhiko Tada, Matthias Wabl, Teruo Amagasa, Takeshi TsubataAbstract:Ag-induced B cell proliferation in vivo requires a costimulatory signal through CD40, whereas B cell Ag receptor (BCR) ligation by anti-Ig H chain Abs, such as anti-Ig μ H chain Ab and anti-Ig δ H chain Ab, alone induces proliferation of B cells in vitro, even in the absence of CD40 ligation. In this study, we demonstrate that CD40 signaling is required for survival and proliferation of B cells stimulated by protein Ags in vitro as well as in vivo. This indicates that the in vitro system represents B cell activation in vivo, and that protein Ags generate BCR signaling distinct from that by anti-Ig H chain Abs. Indeed, BCR ligation by Ags, but not by anti-Ig H chain Abs, efficiently phosphorylates the inhibitory coreceptors CD22 and CD72. When these coreceptors are activated, anti-Ig H chain Ab-stimulated B cells can survive and proliferate only in the presence of CD40 signaling. Conversely, treatment of Ag-stimulated B cells with anti-CD72 mAb blocks CD72 phosphorylation and induces proliferation, even in the absence of CD40 signaling. These results strongly suggest that activation of B cells by anti-Ig H chain Abs involves their ability to silence the inhibitory coreceptors, and that the inhibitory coreceptors install requirement of CD40 signaling for survival and proliferation of Ag-stimulated B cells.
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SHP-1 Requires Inhibitory Co-receptors to Down-modulate B Cell Antigen Receptor-mediated Phosphorylation of Cellular Substrates
The Journal of biological chemistry, 2001Co-Authors: Takahiro Adachi, Chisato Wakabayashi, Hidetaka Yakura, Jürgen Wienands, Michael Reth, Takeshi TsubataAbstract:Abstract Signaling through the B cell antigen receptor (BCR) is negatively regulated by the SH2 domain-containing protein-tyrosine phosphatase SHP-1, which requires association with tyrosine-phosphorylated proteins for activation. Upon BCR ligation, SHP-1 has been shown to associate with the BCR, the cytoplasmic protein-tyrosine kinases Lyn and Syk, and the inhibitory co-receptors CD22 and CD72. How SHP-1 is activated by BCR ligation and regulates BCR signaling is, however, not fully understood. Here we demonstrate that, in the BCR-expressing myeloma line J558Lμm3, CD72 expression reduces the BCR ligation-induced phosphorylation of the BCR component Igα/Igβ and its cytoplasmic effectors Syk and SLP-65. Substrate phosphorylation was restored by expression of dominant negative mutants of SHP-1, whereas the SHP-1 mutants failed to enhance phosphorylation of the cellular substrates in the absence of CD72. This indicates that SHP-1 is efficiently activated by CD72 but not by other pathways in J558Lμm3 cells and that inhibition of SHP-1 specifically activated by CD72 reverses CD72-induced dephosphorylation of cellular substrates in these cells. Taken together, BCR-induced SHP-1 activation is likely to require inhibitory co-receptors such as CD72, and SHP-1 appears to mediate the negative regulatory effect of CD72 on BCR signaling by dephosphorylating Igα/Igβ and its downstream signaling molecules Syk and SLP-65.
Subbarao Bondada - One of the best experts on this subject based on the ideXlab platform.
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Positive and negative roles of CD72 in B cell function
Immunologic Research, 2002Co-Authors: Subbarao BondadaAbstract:Regulation of B cell activation depends on integration of signals transmitted by the B cell receptor (BCR) and a variety of co-receptors. CD72 is a B cell co-receptor that is expressed in all stages of B cell development except plasma cells. Ligation of CD72 enhances B cell growth and differentiation. Recently, the class IV semaphoring, CD 100, has been identified as the natural ligand for CD72. Cytoplasmic domain of CD72 has been shown to be associated with SHP-1 leading to the proposal that the positive effects of CD72 on B cell response may result from sequestration of negative signals from BCR. However, association of CD72 with Grb2 and/or CD19 suggests that CD72 could transmit positive signals. Based on these data, we propose a dual signaling model of CD72.
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Positive signaling through CD72 induces mitogen-activated protein kinase activation and synergizes with B cell receptor signals to induce X-linked immunodeficiency B cell proliferation.
Journal of immunology (Baltimore Md. : 1950), 2001Co-Authors: Chandrasekar Venkataraman, Steven Estus, Chen Dong, Roger J. Davis, Richard A Flavell, Subbarao BondadaAbstract:CD72 is a 45-kDa B cell transmembrane glycoprotein that has been shown to be important for B cell activation. However, whether CD72 ligation induces B cell activation by delivering positive signals or sequestering negative signals away from B cell receptor (BCR) signals remains unclear. Here, by comparing the late signaling events associated with the mitogen-activated protein kinase pathway, we identified many similarities and some differenes between CD72 and BCR signaling. Thus, CD72 and BCR activated the extracellular signal-regulated kinase (ERK) and the c-Jun N-terminal kinase (JNK) but not p38 mitogen-activated protein kinase. Both CD72- and BCR-mediated ERK and JNK activation required protein kinase C activity, which was equally important for CD72- and BCR-induced B cell proliferation. However, CD72 induced stronger JNK activation compared with BCR. Surprisingly, the JNK activation induced by both BCR and CD72 is Btk independent. Although both CD72 and BCR induced Btk-dependent ERK activation, CD72-mediated proliferation is more resistent to blocking of ERK activity than that of BCR, as shown by the proliferation response of B cells treated with PD98059 and dibutyryl cAMP, agents that inhibit ERK activity. Most importantly, CD72 signaling compensated for defective BCR signaling in X-linked immunodeficiency B cells and partially restored the proliferation response of X-linked immunodeficiency B cells to anti-IgM ligation. These results suggest that CD72 signals B cells by inducing BCR-independent positive signaling pathways.
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CD72 mediated b cell activation involves recruitment of cd19 and activation of phosphatidylinositol 3 kinase
European Journal of Immunology, 1998Co-Authors: Chandrasekar Venkataraman, Anne Mette Buhl, Ching Shih Chen, John C. Cambier, Subbarao BondadaAbstract:Occupancy of the B cell glycoprotein, CD72 results in syk-independent activation of phospholipase-C gamma and calcium mobilization. The cytoplasmic tail of CD72 does not contain an immunoreceptor tyrosine-based activation motif to directly transduce signals into the B lymphocyte. Hence, we investigated whether other coreceptors such as CD19 and its associated phosphatidylinositol 3-kinase (PI 3-K) were involved in CD72 signaling. Two specific inhibitors of PI 3-K inhibited CD72-stimulated B cell proliferation in a dose-dependent manner. Activation of B lymphocytes via CD72 resulted in recruitment and activation of PI 3-K, which was mediated by CD19. Accordingly, CD72 ligation induced CD19 tyrosine phosphorylation. Thus, lipid products generated as a result of PI 3-K activation may have an important function in CD72-mediated B lymphocyte activation. The kinetics of CD19 tyrosine phosphorylation induced by CD72 ligation were strikingly different from those seen following B cell antigen receptor (BCR) stimulation. A transient increase in the tyrosine phosphorylation of the complement receptors, CD21 and CD35 was observed in BCR- but not CD72-stimulated cells. Co-cross-linking of CD72 and CD19 failed to induce syk tyrosine phosphorylation suggesting that even under these conditions, CD72 signaling was independent of syk activation. A transient and stimulation-dependent physical association between CD19 and CD72 was observed in CD72-ligated cells. These observations suggest a mechanism by which CD72 can recruit CD19 and influence activation of CD19-associated PI 3-K, which appears to be critical for CD72-mediated B cell activation.
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Activation of Lyn, Blk, and Btk But Not Syk in CD72-Stimulated B Lymphocytes
Journal of immunology (Baltimore Md. : 1950), 1998Co-Authors: Chandrasekar Venkataraman, Natarajan Muthusamy, Subramanian Muthukkumar, Subbarao BondadaAbstract:CD72 is a B cell-specific glycoprotein that has been shown to be important for activation of mature B cells. Previously we showed that some of the early signaling events, such as calcium mobilization and phospholipase-γ activation, were similar in B cell Ag receptor (BCR)- and CD72-stimulated B cells and that BCR- but not CD72-mediated early signaling events were blocked by protein kinase A activation. The present report shows that CD72 ligation induces a variety of tyrosine-phosphorylated proteins, most of which were of the same molecular mass as those seen in anti-IgM-treated B cells, except for a 72-kDa protein. Further analysis showed that the tyrosine kinases lyn and blk were activated in CD72-ligated B cells. Interestingly, the non-src kinase syk was not activated in CD72-stimulated cells whereas the tec family kinase btk was activated in both CD72- and BCR-stimulated B cells. Furthermore, B cells from xid mice were unresponsive to CD72-induced proliferation, indicating an essential role for btk in CD72-induced signaling events. Surprisingly, tyrosine phosphorylation of phospholipase C-γ2 was normal in CD72-stimulated cells in spite of a lack of activation of syk. Furthermore, B cell proliferation through CD72 was blocked by the immunosuppressive agents cyclosporin A and FK506, indicating the important role for Ca 2+ -regulated activation events similar to BCR-stimulated cells. We propose that btk can substitute for syk in inducing phospholipase C-γ2 tyrosine phosphorylation and initiating calcium mobilization in CD72-stimulated B lymphocytes.
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CD72‐mediated B cell activation involves recruitment of CD19 and activation of phosphatidylinositol 3‐kinase
European journal of immunology, 1998Co-Authors: Chandrasekar Venkataraman, Anne Mette Buhl, Ching Shih Chen, John C. Cambier, Subbarao BondadaAbstract:Occupancy of the B cell glycoprotein, CD72 results in syk-independent activation of phospholipase-C gamma and calcium mobilization. The cytoplasmic tail of CD72 does not contain an immunoreceptor tyrosine-based activation motif to directly transduce signals into the B lymphocyte. Hence, we investigated whether other coreceptors such as CD19 and its associated phosphatidylinositol 3-kinase (PI 3-K) were involved in CD72 signaling. Two specific inhibitors of PI 3-K inhibited CD72-stimulated B cell proliferation in a dose-dependent manner. Activation of B lymphocytes via CD72 resulted in recruitment and activation of PI 3-K, which was mediated by CD19. Accordingly, CD72 ligation induced CD19 tyrosine phosphorylation. Thus, lipid products generated as a result of PI 3-K activation may have an important function in CD72-mediated B lymphocyte activation. The kinetics of CD19 tyrosine phosphorylation induced by CD72 ligation were strikingly different from those seen following B cell antigen receptor (BCR) stimulation. A transient increase in the tyrosine phosphorylation of the complement receptors, CD21 and CD35 was observed in BCR- but not CD72-stimulated cells. Co-cross-linking of CD72 and CD19 failed to induce syk tyrosine phosphorylation suggesting that even under these conditions, CD72 signaling was independent of syk activation. A transient and stimulation-dependent physical association between CD19 and CD72 was observed in CD72-ligated cells. These observations suggest a mechanism by which CD72 can recruit CD19 and influence activation of CD19-associated PI 3-K, which appears to be critical for CD72-mediated B cell activation.
Thomas Dorner - One of the best experts on this subject based on the ideXlab platform.
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op0273 inhibition of b cell receptor signaling with epratuzumab and the effects of alpha 2 6 sialic acid removal
Annals of the Rheumatic Diseases, 2013Co-Authors: N Sieger, Karin Reiter, Sarah J Fleischer, Capucine Daridon, Anthony Shock, Gerd-rüdiger Burmester, Thomas DornerAbstract:Background CD22 is a trans-membrane protein exclusively expressed on B cells which regulates adhesion/migration and inhibits cellular responses to B-cell receptor (BCR) stimulation[1]. Spleen tyrosine kinase (Syk) and subsequent phospholipase C (PLC)-γ2-triggered Ca 2+ fluxes are rapidly activated after BCR stimulation. CD22 binds specifically to α-2,6-sialic acid residues present on a broad number of proteins, including CD22 itself. Sialo-interactions appear to be crucial for optimal function of CD22 and can occur in cis (to ligands on the same cell surface) or trans (to ligands on neighboring cells). Objectives The humanized anti-CD22 monoclonal antibody epratuzumab, currently being tested in clinical trials, modulates adhesion molecule expression and B cell migration in vitro [2] however, the potential of CD22-ligation with cis and trans sialic acid to regulate BCR signaling has not been delineated. Methods In vitro effects of epratuzumab on BCR-induced signaling were evaluated by analyzing the phosphorylation of BCR-signaling molecules (Syk/PLC-γ2) and Ca 2+ flux. The influence of epratuzumab on the phosphorylation of the BCR signaling molecules Syk and PLC-γ2 was studied in vitro by flow cytometry in PBMCs from healthy donors. To investigate the effect of sialo-interaction on the effects by epratuzumab, PBMCs were treated with neuraminidase (to remove sialic acic) and BCR induced phosphorylation (Syk and PLC-γ2) was detected with or without epratuzumab incubation. To monitor the concentration of intracellular Ca 2+ , B cells pre-loaded with Indo-1AM were incubated with or without F(ab’) 2 -epratuzumab for 60 minutes at 37°C and were subsequently stimulated with F(ab’) 2 anti-IgM/IgG. Intracellular Ca 2+ concentrations were monitored by flow cytometry over 10 minutes after activation. Results Epratuzumab inhibited Syk and PLC-γ2 phosphorylation after in vitro BCR stimulation by 23±9% and 33±5, respectively. Interestingly, preventing sialo-interactions of CD22 partially reduced the inhibitory effect of epratuzumab on Syk and PLC-γ2 phosphorylation to 12±6% and 22±4%, respectively. The inhibition of kinase phosphorylation was demonstrated in both CD27 - and CD27 + memory B cells and appeared to be independent of FcR signaling since a F(ab’) 2 fragment of epratuzumab induced comparable results to the full antibody. In addition, a F(ab’) 2 fragment of epratuzumab reduced the BCR-induced calcium flux. These data are consistent with the potential of targeting CD22 to raise the threshold of BCR activation which also provides additional control of B cell function. Conclusions Intracellular BCR signals can be partially inhibited by CD22 ligation with epratuzumab, providing further understanding of targeting CD22 biology in human autoimmune diseases. This study was supported by Sonderforschungsbereich 650 and DFG491/7-1 and in part by UCB Pharma Inc. References Nitschke L. The role of CD22 and other inhibitory co-receptors in B-cell activation. Current opinion in immunology 2005;17:290–7. Daridon C, Blassfeld D, Reiter K, et al. Epratuzumab targeting of CD22 affects adhesion molecule expression and migration of B-cells in systemic lupus erythematosus. Arthritis Research & Therapy 2010;12:R204. Disclosure of Interest N. Sieger: None Declared, S. Fleischer: None Declared, H. Mei: None Declared, K. Reiter: None Declared, A. Shock Employee of: UCB Pharma inc., G. Burmester: None Declared, C. Daridon: None Declared, T. Dorner Grant/Research support from: Immunomedics
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CD22 ligation inhibits downstream B cell receptor signaling and Ca2+ flux upon activation
Arthritis & Rheumatism, 2013Co-Authors: N Sieger, Karin Reiter, Sarah J Fleischer, Capucine Daridon, Anthony Shock, Gerd-rüdiger Burmester, Thomas DornerAbstract:Objective CD22 is a surface molecule exclusively expressed on B cells that regulates adhesion and B cell receptor (BCR) signaling as an inhibitory coreceptor of the BCR. Central downstream signaling molecules that are activated upon BCR engagement include spleen tyrosine kinase (Syk) and, subsequently, phospholipase Cγ2 (PLCγ2), which results in calcium (Ca2+) mobilization. The humanized anti-CD22 monoclonal antibody epratuzumab is currently being tested in clinical trials. This study was undertaken to determine the potential mechanism by which this drug regulates B cell activation. Methods Purified B cells were preincubated with epratuzumab, and the colocalization of CD22 and CD79α, without BCR engagement, was assessed by confocal microscopy. The phosphorylation of Syk (Y348, Y352) and PLCγ2 (Y759) as well as the Ca2+ flux in the cells were analyzed by flow cytometry upon stimulation of the BCR and/or Toll-like receptor 9 (TLR-9). The influence of CD22 ligation on BCR signaling was assessed by pretreating the cells with epratuzumab or F(ab′)2 fragment of epratuzumab, in comparison with control cells (medium alone or isotype-matched IgG1). Results Epratuzumab induced colocalization of CD22 and components of the BCR independent of BCR engagement, and also reduced intracellular Ca2+ mobilization and diminished the phosphorylation of Syk and PLCγ2 after BCR stimulation in vitro. Inhibition of kinase phosphorylation was demonstrated in both CD27− and CD27+ B cells, and this appeared to be independent of Fc receptor signaling. Preactivation of the cells via the stimulation of TLR-9 did not circumvent the inhibitory effect of epratuzumab on BCR signaling. Conclusion These findings are consistent with the concept of targeting CD22 to raise the threshold of BCR activation, which could offer therapeutic benefit in patients with autoimmune diseases.
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epratuzumab affects b cells trafficking in systemic lupus erythematosus
Annals of the Rheumatic Diseases, 2011Co-Authors: Capucine Daridon, Daniela Blaβfeld, Karin Reiter, Claudia Giesecke, Daniela Frolich, Arwed Hostmann, Arne Hansen, David M. Goldenberg, Thomas DornerAbstract:Background Epratuzumab, a humanised anti-CD22 monoclonal antibody, is under investigation as a therapeutic antibody in non-Hodgkin9s lymphoma and systemic lupus erythematosus (SLE), but its mechanism of action on B cells remains elusive. Treatment of SLE patients with epratuzumab leads to a reduction of circulating CD27 negative B cells, although epratuzumab is weakly cytotoxic to B cells in vitro. Therefore, potential effects of epratuzumab on adhesion molecule expression and the migration of B cells have been evaluated. Methods Epratuzumab binding specificity and the surface expression of adhesion molecules (CD62L, β7 integrin and β1 integrin) after culture with epratuzumab was studied on B cell subsets of SLE patients by flow cytometry. In addition, in vitro transwell migration assays were performed to analyse the effects of epratuzumab on migration towards different chemokines such as CXCL12, CXCL13 or to CXCR3 ligands, and to assess the functional consequences of altered adhesion molecule expression. Results Epratuzumab binding was considerably higher on B cells relative to other cell types assessed. No binding of epratuzumab was observed on T cells, while weak non-specific binding of epratuzumab on monocytes was noted. On B cells, binding of epratuzumab was particularly enhanced on CD27 negative B cells compared to CD27 positive B cells, primarily related to a higher expression of CD22 on CD27 negative B cells. Moreover, epratuzumab binding led to a decrease in the cell surface expression of CD62L and β7 integrin, while the expression of β1 integrin was enhanced. The effects on the pattern of adhesion molecule expression observed with epratuzumab were principally confined to a fraction of the CD27 negative B cell subpopulation and were associated with enhanced spontaneous migration of B cells. Furthermore, epratuzumab also enhanced the migration of CD27 negative B cells towards the chemokine CXCL12. Conclusions The current data suggest that epratuzumab has effects on the expression of the adhesion molecules CD62L, β7 integrin and β1 integrin as well as on migration towards CXCL12, primarily of CD27 negative B cells. Therefore, induced changes in migration appear to be part of the mechanism of action of epratuzumab and are consistent with the observation that CD27 negative B cells are preferentially removed from the peripheral blood under treatment.