scholarly journals Utilization of a photoactivatable antigen system to examine B-cell probing termination and the B-cell receptor sorting mechanisms during B-cell activation

2016 ◽  
Vol 113 (5) ◽  
pp. E558-E567 ◽  
Author(s):  
Jing Wang ◽  
Shan Tang ◽  
Zhengpeng Wan ◽  
Yiren Gao ◽  
Yiyun Cao ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Single Molecule ◽  
Cell Activation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Antigen Binding ◽  
B Cell Activation ◽  
Class I Mhc ◽  
Antigen System

Antigen binding to the B-cell receptor (BCR) induces several responses, resulting in B-cell activation, proliferation, and differentiation. However, it has been difficult to study these responses due to their dynamic, fast, and transient nature. Here, we attempted to solve this problem by developing a controllable trigger point for BCR and antigen recognition through the construction of a photoactivatable antigen, caged 4-hydroxy-3-nitrophenyl acetyl (caged-NP). This photoactivatable antigen system in combination with live cell and single molecule imaging techniques enabled us to illuminate the previously unidentified B-cell probing termination behaviors and the precise BCR sorting mechanisms during B-cell activation. B cells in contact with caged-NP exhibited probing behaviors as defined by the unceasing extension of membrane pseudopods in random directions. Further analyses showed that such probing behaviors are cell intrinsic with strict dependence on F-actin remodeling but not on tonic BCR signaling. B-cell probing behaviors were terminated within 4 s after photoactivation, suggesting that this response was sensitive and specific to BCR engagement. The termination of B-cell probing was concomitant with the accumulation response of the BCRs into the BCR microclusters. We also determined the Brownian diffusion coefficient of BCRs from the same B cells before and after BCR engagement. The analysis of temporally segregated single molecule images of both BCR and major histocompatibility complex class I (MHC-I) demonstrated that antigen binding induced trapping of BCRs into the BCR microclusters is a fundamental mechanism for B cells to acquire antigens.

scholarly journals PTIP chromatin regulator controls development and activation of B cell subsets to license humoral immunity in mice

2017 ◽  
Vol 114 (44) ◽  
pp. E9328-E9337 ◽  
Author(s):  
Dan Su ◽  
Stijn Vanhee ◽  
Rebeca Soria ◽  
Elin Jaensson Gyllenbäck ◽  
Linda M. Starnes ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Humoral Immunity ◽  
Cell Activation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Transactivation Domain ◽  
B Cell Activation ◽  
Chromatin Regulator ◽  
B Cell Subsets

B cell receptor signaling and downstream NF-κB activity are crucial for the maturation and functionality of all major B cell subsets, yet the molecular players in these signaling events are not fully understood. Here we use several genetically modified mouse models to demonstrate that expression of the multifunctional BRCT (BRCA1 C-terminal) domain-containing PTIP (Pax transactivation domain-interacting protein) chromatin regulator is controlled by B cell activation and potentiates steady-state and postimmune antibody production in vivo. By examining the effects of PTIP deficiency in mice at various ages during ontogeny, we demonstrate that PTIP promotes bone marrow B cell development as well as the neonatal establishment and subsequent long-term maintenance of self-reactive B-1 B cells. Furthermore, we find that PTIP is required for B cell receptor- and T:B interaction-induced proliferation, differentiation of follicular B cells during germinal center formation, and normal signaling through the classical NF-κB pathway. Together with the previously identified role for PTIP in promoting sterile transcription at the Igh locus, the present results establish PTIP as a licensing factor for humoral immunity that acts at several junctures of B lineage maturation and effector cell differentiation by controlling B cell activation.

scholarly journals P063 B-cell receptor signalling in lymphoid tissues may be regulated by CEACAM1

2020 ◽  
Vol 14 (Supplement_1) ◽  
pp. S166-S166
Author(s):  
T Nagaishi ◽  
N Tsugawa ◽  
D Yamada ◽  
T Watabe ◽  
M Onizawa ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
Cell Surface ◽  
B Cell ◽  
Cell Activation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphoid Tissues ◽  
B Cell Activation ◽  
Activation Markers

Abstract Background It has been recently shown that the carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1) expressed in T cells may regulate immune responses in the gut. Moreover, it has also been reported that the treatments with either an agonistic monoclonal antibody (mAb) or natural ligands for this molecule can suppress colitis severity in murine models of inflammatory bowel diseases (IBD). On the other hand, in addition to T cells, B cells are also an important population in the gut-associated lymphoid tissues (GALT) that orchestrate mucosal homeostasis. However, the role of CEACAM1 in B cells has not been elucidated. Methods We analysed primary B-cell subsets in the lymphoid tissues of wild-type C57BL6 mice as well as a murine B-cell line, A20, to determine the expressions and functions of CEACAM1. Results FACS analysis of the lymphocyte subsets isolated from secondary lymphoid tissues such as spleen, mesenteric lymph nodes and Peyer’s patches of C57BL6 revealed higher expression level of CEACAM1 on B-cell surface than that of T cells. Bone marrow analysis showed that such CEACAM1 expression was increased during maturation and differentiation process of B cells. When isolated splenic B cells were stimulated with LPS, anti-CD40 or anti-μ chain Abs in the presence of agonistic anti-CEACAM1 mAb, the usual increased cytokine productions (such as IL-4 and IL-5 by activation via B cell receptor (BCR) signalling) were specifically suppressed by CEACAM1 signalling rather than B-cell activations via either TLR4 or CD40 signalling. Immunofluorescent studies using confocal microscopy revealed co-localisation of CEACAM1 and BCR when B cells were activated with anti-μ chain Ab. Given these results, A20 cells were transfected with CEACAM1 cDNA. Biochemical analysis showed that an inducible overexpression of CEACAM1 suppressed the BCR signalling in these cells when compared with that of vector alone-transfected control. Moreover, the overexpression of CEACAM1 in these cells resulted in reduced expressions of activation markers such as CD69, CD80, CD86, MHC-I and -II on the cell surface. These observations were associated with decreased Ca2+ influx and suppressed cytokine production by the overexpression of CEACAM1 after BCR signal activation. Conclusion These results suggest that CEACAM1 can regulate B-cell activation and differentiation specifically via BCR signalling in the lymphoid tissues. Therefore, this molecule can be a therapeutic target in IBD by regulating of both T-cell and B-cell activation in GALT.

scholarly journals The B Cell Receptor Itself Can Activate Complement to Provide the Complement Receptor 1/2 Ligand Required to Enhance B Cell Immune Responses In Vivo

2003 ◽  
Vol 198 (4) ◽  
pp. 591-602 ◽  
Author(s):  
Joerg Rossbacher ◽  
Mark J. Shlomchik
Keyword(s):  
Immune Response ◽  
B Cells ◽  
Immune Responses ◽  
B Cell ◽  
Cell Activation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Mutant Mice ◽  
B Cell Activation

B cells express complement receptors (CRs) that bind activated fragments of C3 and C4. Immunized CR knockout (KO) mice have lower antibody titers and smaller germinal centers (GCs), demonstrating the importance of CR signals for the humoral immune response. CR ligands were thought to be generated via complement fixation mediated by preexisting “natural” IgM or early Ab from inefficiently activated B cells. This concept was recently challenged by a transgenic (Tg) mouse model that lacks circulating antibody but still retains membrane IgM (mIgM) and mounts normal immune responses. To test whether CR ligands could be generated by the B cell receptor (BCR) itself, we generated similar mice carrying a mutated mIgM that was defective in C1q binding. We found that B cells from such mutant mice do not deposit C3 on B cells upon BCR ligation, in contrast to B cells from mIgM mice. This has implications for the immune response: the mutant mice have smaller GCs than mIgM mice, and they are particularly deficient in the maintenance of the GC response. These results demonstrate a new BCR-dependent pathway that is sufficient and perhaps necessary to provide a CR1/2 ligand that promotes efficient B cell activation.

scholarly journals APEX2 proximity biotinylation reveals protein dynamics triggered by B cell receptor activation

2020 ◽  
Author(s):  
Luqman O Awoniyi ◽  
Vid Šuštar ◽  
Sara Hernández-Pérez ◽  
Marika Vainio ◽  
Alexey V Sarapulov ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Lipid Raft ◽  
Cell Activation ◽  
Kinetic Resolution ◽  
Cell Receptor ◽  
Receptor Activation ◽  
B Cell Receptor ◽  
B Cell Activation ◽  
Bcr Signaling

ABSTRACTB lymphocytes form a central part of the adaptive immune system, helping to clear infections by mounting antibody responses and immunological memory. B cell activation is critically controlled by a specific antigen receptor, the B cell receptor (BCR), which triggers a complex, multibranched signaling cascade initiating various cellular changes. While parts of these pathways are reasonably well characterized, we still lack a comprehensive protein-level view of the very dynamic and robust cellular response triggered by antigen engagement. Ability to track, with sufficient kinetic resolution, the protein machineries responding to BCR signaling is imperative to provide new understanding into this complex cell activation event. We address this challenge by using APEX2 proximity labeling technique, that allows capture a major fraction of proteins in a given location with 20nm range and 1min time window, and target the APEX2 enzyme to the plasma membrane lipid raft domain, where BCR efficiently translocates upon activation. Our data provides unprecedented insights into the protein composition of lipid raft environment in B cells, and the changes triggered there upon BCR cross-linking and translocation. In total, we identified 1677 proteins locating at the vicinity of lipid raft domains in cultured mouse B cells. The data includes a majority of proteins known to be involved in proximal BCR signaling. Interestingly, our differential enrichment analysis identified various proteins that underwent dynamic changes in their localization but that had no previously known linkage to early B cell activation. As expected, we also identified, for example, a wealth of proteins linked to clathrin-mediated endocytosis that were recruited to the lipid rafts upon cell activation. We believe that his data serves as a valuable record of proteins involved in BCR activation response and aid various future studies in the field.

scholarly journals Functional Outcome of B Cell Activation by Chromatin Immune Complex Engagement of the B Cell Receptor and TLR9

2007 ◽  
Vol 179 (11) ◽  
pp. 7397-7405 ◽  
Author(s):  
Liliana Busconi ◽  
Jason W. Bauer ◽  
Joseph R. Tumang ◽  
Amy Laws ◽  
Kristin Perkins-Mesires ◽  
...  
Keyword(s):  
B Cell ◽  
Functional Outcome ◽  
Immune Complex ◽  
Cell Activation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
B Cell Activation

scholarly journals Monovalent engagement of the BCR activates ovalbumin-specific transnuclear B cells

2014 ◽  
Vol 211 (2) ◽  
pp. 365-379 ◽  
Author(s):  
Ana M. Avalos ◽  
Angelina M. Bilate ◽  
Martin D. Witte ◽  
Albert K. Tai ◽  
Jiang He ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Synthetic Peptides ◽  
Cell Activation ◽  
Cell Receptor ◽  
Fine Tuning ◽  
B Cell Activation ◽  
Cd40 Ligation ◽  
Peptide Antigen ◽  
Peptide Epitope

Valency requirements for B cell activation upon antigen encounter are poorly understood. OB1 transnuclear B cells express an IgG1 B cell receptor (BCR) specific for ovalbumin (OVA), the epitope of which can be mimicked using short synthetic peptides to allow antigen-specific engagement of the BCR. By altering length and valency of epitope-bearing synthetic peptides, we examined the properties of ligands required for optimal OB1 B cell activation. Monovalent engagement of the BCR with an epitope-bearing 17-mer synthetic peptide readily activated OB1 B cells. Dimers of the minimal peptide epitope oriented in an N to N configuration were more stimulatory than their C to C counterparts. Although shorter length correlated with less activation, a monomeric 8-mer peptide epitope behaved as a weak agonist that blocked responses to cell-bound peptide antigen, a blockade which could not be reversed by CD40 ligation. The 8-mer not only delivered a suboptimal signal, which blocked subsequent responses to OVA, anti-IgG, and anti-kappa, but also competed for binding with OVA. Our results show that fine-tuning of BCR-ligand recognition can lead to B cell nonresponsiveness, activation, or inhibition.

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