Integration of activatory and inhibitory signals in human B-cells (original) (raw)
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CD21 and FCRL5 form a receptor complex with robust B cell activating capacity
International immunology, 2018
The B cell response to antigen is critically regulated by co-receptors. CD21 (complement receptor 2) amplifies the response to antigen linked to its ligands, specific C3 fragments. In contrast, human Fc receptor-like 5 (FCRL5), a novel IgG receptor, was reported to inhibit B cell receptor (BCR) signaling. Here we show that CD21 and FCRL5 physically associate, suggesting that immune complexes containing both C3 fragment and IgG could simultaneously engage the pre-assembled receptors. We found that activating signaling molecules such as CD19, active PLCγ2 and BTK were rapidly recruited to FCRL5 upon engagement, suggesting a novel activating function for FCRL5. We confirmed that FCRL5 through its ITIM motifs inhibited BCR signaling in the absence of CD21 stimulation. In contrast, triple-engagement of FCRL5, CD21 and the BCR led to superior calcium response compared to CD21 and BCR co-stimulation, in both cell lines and tonsil B cells. Furthermore, the novel activating function was inde...
Immunology, 2014
B-cell antigen receptor (BCR) signalling and its regulation through negative and positive regulators are critical for balancing B-cell response and function. Human Fc receptor like-2 (FCRL2), a member of the newly identified FCRL family, could influence B-cell signalling due to possession of both immunoreceptor tyrosine-based activation and inhibitory motifs (ITAM and ITIM). Since the natural ligand of FCRL2 has not been identified, we generated FCRL2-specific monoclonal antibodies (mAbs) and employed them to investigate the influence of FCRL2 stimulation on BCR signalling in an FCRL2-expressing B-cell line. Two anti-FCRL2 mAb-producing hybridoma clones (5A7-E7 and 3D8-G8) were selected. None of the mAbs displayed any cross-reactivity with the other members of the FCRL family including recombinant FCRL1,-3,-4 and-5, as tested by FACS and ELISA techniques. Engagement of the FCRL2 by these mAbs resulted in significant inhibition of BCR signalling mediators such as calcium mobilization and phosphorylation of the mitogen-activated protein kinases Erk, p38 and Jnk. These findings indicate that the FCRL2 ITIM motifs are functional and the anti-FCRL2 mAbs may mimic the natural ligand of FCRL2 by induction of inhibitory signals in B cells.
Uncoupling CD21 and CD19 of the B-cell coreceptor
Proceedings of the National Academy of Sciences, 2009
Complement receptors (CRs) CD21 and CD35 form a coreceptor with CD19 and CD81 on murine B cells that when coligated with the B-cell receptor lowers the threshold of activation by several orders of magnitude. This intrinsic signaling role is thought to explain the impaired humoral immunity of mice bearing deficiency in CRs. However, CRs have additional roles on B cells independent of CD19, such as transport of C3-coated immune complexes and regulation of C4 and C3 convertase. To test whether association of CR with CD19 is necessary for their intrinsic activation-enhancing role, knockin mice expressing mutant receptors, Cr2 Δ/Δgfp , that bind C3 ligands but do not signal through CD19 were constructed. We found that uncoupling of CR and CD19 significantly diminishes survival of germinal center B cells and secondary antibody titers. However, B memory is less impaired relative to mice bearing a complete deficiency in CRs on B cells. These findings confirm the importance of interaction of...
The Journal of Immunology, 2005
C3dg is a cleavage product of the C3 component of complement that can facilitate the coligation of the complement receptor 2 (CR2/CD21) with the BCR via C3dg/Ag complexes. This interaction can greatly amplify BCR-mediated signaling events and acts to lower the threshold for B cell activation. Although previous studies have used anti-CR2 Abs or used chimeric Ags in the context of BCR transgenic mice as surrogate C3d-containing ligands, we have used a physiological form of C3d to study signaling in B cells from wild-type C57BL/6 mice. We find that while CR2-enhanced BCR signaling causes intracellular Ca2+ mobilization and total pTyr phosphorylation of an intensity comparable to optimal BCR ligation using anti-IgM Abs, it does so with limited activation of inhibitory effectors (such as CD22, Src homology region 2 domain containing phosphatase 1, and SHIP-1) and without substantial receptor cross-linking. In summary, we demonstrate that CR2-enhanced BCR signaling may proceed not only th...
International Immunology, 2002
The human and murine CD21 gene products have been functionally linked to B cell activation by the co-ligation of the BCR and the CD21/CD19/CD81 complexes. Binding of low levels of antigen complexed to the complement ligand(s) for CD21 enhances B cell activation compared to the stimulation caused by antigen alone. Mice lacking functional CD21 predispose to autoimmune responses suggesting that this receptor may also play a negative role: thus in the presence of excess complement-bearing immune complexes, B cell antigen-specific activation may be inhibited. This possibility was investigated using intracellular calcium elicitation analyses to follow BCR-mediated activation. Ligation of the BCR and limiting quantities of the CD21 receptor demonstrated the expected enhanced cellular response compared to BCR ligation alone: CD21 ligation alone demonstrated no alteration in calcium flux. However, co-ligation of the BCR with excess CD21 binding resulted in the elimination of the calcium response, suggesting that CD21 ligation was down-modulating the BCR response. Immunoprecipitation of kinases associated with the BCR and CD21/CD19/CD81 complexes demonstrated that Lyn is preferentially depleted from the BCR complex following excess binding of CD21. Localization of other kinases integral for B cell activation is not altered. These data suggest that excess CD21 ligand binding can negatively impact B cell activation by sequestering Lyn kinase away from the BCR complex.
mAbs, 2014
ACPA -Anti-citrullinated protein/peptide antibody, AEC -3-amino-9-ethylcarbazole, BCR -B cell receptor, FcγRIIb -Fcγ receptor IIb, ITAM -Immunoreceptor tyrosine-based activation motif, ITIM -Immunoreceptor tyrosine-based inhibitory motif, MAPK -Mitogen-activated protein kinase, PH -Pleckstrin homology, PI3-K -Phosphatidylinositol 3-kinase, PIP3 -Phosphatidylinositol (3,4,5)-triphosphate, RA -Rheumatoid arthritis, SHIP -SH2 domaincontaining inositol 5-phosphatase, SHP2 -SH2 domain containing protein tyrosine phosphatase, SLE -Systemic lupus erythematosus, TAK1 -TGF-activated kinase, TNF -Tumor necrosis factor.