Overexpression of the Autoimmunity-Associated Tyrosine Phosphatase PTPN22 in CLL Cells Enhances Antiapoptotic B-Cell Receptor Signals Through Selective Activation of the AKT Pathway

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 619-619
Author(s):  
Roberto Negro ◽  
Stefania Gobessi ◽  
Pablo G Longo ◽  
Luca Laurenti ◽  
Dimitar G Efremov
Keyword(s):  
B Cells ◽  
Autoimmune Diseases ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Negative Regulator ◽  
Gain Of Function ◽  
Akt Kinase ◽  
Bcr Signaling ◽  
Increased Activity

Abstract Abstract 619 The phosphatase PTPN22 is an important negative regulator of T cell receptor (TCR) signaling. A gain of function polymorphism in this phosphatase is strongly linked with increased risk for the development of several common autoimmune diseases, including rheumatoid arthritis, insulin-dependent diabetes mellitus and systemic lupus erythematosus. The exact mechanism how the more active PTPN22 variant contributes to the development of these autoimmune diseases is still unclear, but it is has been postulated that increased activity of this enzyme may prevent negative selection of autoreactive T cells by blocking proapoptotic TCR signals induced by autoantigen. We now report that PTPN22 is significantly overexpressed in CLL B cells. Of the 91 investigated CLL samples, in 66 (73%) the levels of PTPN22 were at least 2 fold higher and often exceeded by 10 fold the levels in normal tonsilar B cells (CLL, mean PTPN22 levels 5.7, SD +/−4.3; normal tonsilar B cells, mean PTPN22 levels 0.9, SD +/−0.1). PTPN22 levels were somewhat higher in CD38−positive than CD38−negative cases (P=0.031), whereas no association was observed with IGHV mutation status, ZAP-70 expression and time to treatment. To determine the functional consequences of PTPN22 overexpression, we performed RNA interference experiments in primary CLL B cells with high PTPN22. We evaluated the effects of PTPN22 silencing in unstimulated CLL cells as well as CLL cells stimulated through the B cell receptor (BCR), given the important role of the BCR signaling pathway in the pathophysiology of CLL. Both soluble and immobilized anti-IgM antibodies were used for BCR stimulation, as these two BCR crosslinking agents have opposing effects on leukemic cell survival. Silencing of PTPN22 had no effect on the viability of unstimulated CLL cells (n=11, viable CLL cells: siControl 49%, siPTPN22 48%, P=n.s.), but significantly inhibited the antiapoptotic effect of immobilized anti-IgM (viable CLL cells: siControl 55%, siPTPN22 48%, P=0.001) and enhanced the proapoptotic effect of soluble anti-IgM (viable CLL cells: siControl 44%, siPTPN22 31%, P<0.001). Consistent with these data, overexpression of PTPN22 in the lymphoma B cell line B104 significantly blocked apoptosis induced by soluble anti-IgM (control 22%, PTPN22 35%) without affecting the survival of unstimulated cells (control 52%, PTPN22 51%). The previous experiments suggested that PTPN22 functions as a molecular switch that enhances antiapoptotic and inhibits proapoptotic BCR signals. To establish the molecular mechanisms underlying these different activities, we studied BCR signal transduction in primary CLL cells transfected with control or PTPN22-specific siRNA. Downregulation of PTPN22 increased anti-IgM-induced activation of several BCR signaling molecules, including LYN, SYK, ERK, JNK and the proapoptotic p38MAPK, confirming that PTPN22 is primarily a negative regulator of BCR signaling. However, activation of the antiapoptotic AKT kinase was substantially reduced by silencing of PTPN22, as evidenced by decreased phosphorylation of AKT and its direct targets GSK3b and FOXO3a. Lack of PTPN22 expression was further associated with increased activity of the phosphatase SHIP, a key negative regulator of the AKT kinase. Opposite effects on the activity of these signaling molecules were observed when PTPN22 was overexpressed in B104 cells, thus validating the data obtained by PTPN22 knockdown in primary CLL cells. In summary, this study shows that the phosphatase PTPN22, which has been implicated in the pathogenesis of several common autoimmune diseases, is significantly overexpressed in CLL B-cells. Overexpression of PTPN22 inhibits activation of molecules that propagate the proapoptotic BCR signal, such as p38MAPK, but at the same time enhances the antiapoptotic signal delivered through the AKT kinase. The mechanism how PTPN22 enhances activation of AKT in response to BCR engagement is by preventing Lyn-mediated activation of the negative regulator SHIP. Collectively, these data show that PTPN22 is an important regulatory molecule in CLL and a potential therapeutic target. Inhibitors of this phosphatase, which are currently being developed for the treatment of autoimmune diseases associated with the gain of function PTPN22 variant, would be expected to convert the BCR survival signal into a death signal and may thus provide means for selective targeting of the malignant clone. Disclosures: No relevant conflicts of interest to declare.

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.

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.

Overexpression of the Protein Tyrosine Phosphatase Lyp Reduces the Responsiveness of Chronic Lymphocytic Leukemia B-Cells to B-Cell Receptor Ligation.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 800-800
Author(s):  
Roberto Negro ◽  
Pablo G Longo ◽  
Michela Tarnani ◽  
Stefania Gobessi ◽  
Luca Laurenti ◽  
...  
Keyword(s):  
Signal Transduction ◽  
B Cells ◽  
Autoimmune Diseases ◽  
B Cell ◽  
Conflicts Of Interest ◽  
Signaling Molecules ◽  
Negative Regulator ◽  
Nucleotide Sequence Analysis ◽  
Aberrant Expression ◽  
Bcr Signaling

Abstract Abstract 800 CLL B cells display many features that suggest a role for antigen stimulation in the development and progression of the disease. These include the expression of stereotyped B-cell receptors (BCRs), the association between IgVH gene mutation status and prognosis, and the gene-expression profile of antigen-stimulated B cells. In addition, CLL B cells have other BCR-related features that distinguish them from normal B lymphocytes, such as lower levels of surface Ig, less efficient BCR signal transduction and increased basal activity of the proximal BCR signaling molecules Lyn and Syk. We have now investigated whether any of these features are related to aberrant expression or function of the phosphatases SHP-1, SHP-2 and Lyp (PTPN22), which regulate the amplitude and duration of the BCR signal by dephosphorylating various components of the BCR signal transduction unit. These phosphatases are also interesting because mutated or polymorphic variants have been linked to various malignant or autoimmune diseases. We started our study by performing nucleotide sequence analysis of the complete coding region of SHP1, SHP2 and Lyp in 8, 21 and 29 CLL B cell samples, respectively. Overall, only two mutations were identified (an R527C substitution in SHP2 and a Q456E substitution in Lyp, each in a single patient), suggesting that these phosphatases are infrequently mutated in CLL. The previously reported Lyp polymorphisms R620W and R263Q were observed in 2 additional cases. We next investigated expression of these phosphatases in purified CLL and normal B cells by immunoblotting. Expression of SHP1 and SHP2 was relatively uniform in the different CLL B-cells samples (n=42) and was not different from normal B cells (n=4). In contrast, expression of Lyp was markedly higher in most CLL samples, with 35 of the 49 investigated cases exhibiting 2 to more than 10 fold higher levels than normal B cells (n=5) (CLL, mean Lyp levels 4.7, SD +/−3.7; normal B cells, mean Lyp levels 0.9, SD +/−0.1, P=0.022). The mean Lyp levels were somewhat higher in U-CLL than M-CLL (6.0 vs. 3.9) and ZAP-70-positive than ZAP-70-negative cases (5.6 vs. 4.7), but these differences were not statistically significant. Analysis of Lyp expression in various lymphoma B-cell lines (n=9) also did not reveal significant differences with respect to normal B-cells, suggesting that Lyp overexpression is a specific feature of CLL. To determine what are the consequences of Lyp overexpression on BCR signaling, we downregulated Lyp in primary CLL B-cells by RNA interference and investigated activation of BCR signaling molecules following sIgM crosslinking. Downregulation of Lyp resulted in a substantial increase in BCR-induced phosphorylation of Lyn (Y397), Syk (Y352), BLNK (Y84) and ERK (T202/Y204), suggesting that overexpression of this phosphatase may be at least partially responsible for the lower BCR signaling capacity of CLL B-cells. Since Lyp expression can be induced in resting T cells by activation with anti-CD3, we investigated whether BCR stimulation will have a similar effect on CLL B-cells. A two-fold increase in Lyp levels was observed after 24 hours of sustained BCR stimulation with immobilized anti-IgM, whereas transient stimulation with soluble anti-IgM resulted in a 20% decrease in Lyp levels. These effects were specific for Lyp, since no such changes were observed in the expression of SHP1 and SHP2. In summary, this study shows that CLL B-cells specifically overexpress the phosphatase Lyp, and important negative regulator of BCR signaling that has been implicated in the pathogenesis of several common autoimmune diseases. Given the observation that Lyp can be induced by sustained BCR engagement and in view of recent findings that Lyp is also overexpressed in anergic B cells, these data further support the notion that CLL cells are continuously exposed to (auto)antigen in vivo. Disclosures: No relevant conflicts of interest to declare.

scholarly journals The Ligand-binding Domain of CD22 Is Needed for Inhibition of the B Cell Receptor Signal, as Demonstrated by a Novel Human CD22-specific Inhibitor Compound

2002 ◽  
Vol 195 (9) ◽  
pp. 1207-1213 ◽  
Author(s):  
Soerge Kelm ◽  
Judith Gerlach ◽  
Reinhard Brossmer ◽  
Claus-Peter Danzer ◽  
Lars Nitschke
Keyword(s):  
B Cells ◽  
Cell Surface ◽  
Ligand Binding ◽  
B Cell ◽  
Intracellular Signaling ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Negative Regulator ◽  
Target Cells ◽  
Lectin Domain

CD22 is a B cell–specific transmembrane protein of the Siglec family. It binds specifically to α2,6-linked sialic acid (Sia) residues, which are also present on glycoproteins on the B cell surface. CD22 acts as a negative regulator in B cell receptor–mediated signaling by recruitment of Src homology 2 domain–containing tyrosine phosphatase (SHP)-1 to its intracellular tail. To analyze how ligand-binding of CD22 influences its intracellular signaling domain, we designed synthetic sialosides as inhibitors for the lectin domain of CD22. One of these compounds inhibited binding of human CD22-Fc to target cells over 200-fold better than Sia and was highly selective for human CD22. When Daudi cells or primary B cells were stimulated with anti-immunoglobulin (Ig)M in presence of this sialoside inhibitor, a higher Ca2+ response was observed, similar to CD22-deficient B cells. Accordingly, a lower tyrosine-phosphorylation of CD22 and SHP-1 recruitment was demonstrated in presence of the sialoside. Thus, by interfering with ligand binding of CD22 on the B cell surface, we have shown for the first time that the lectin domain of CD22 has a direct, positive influence on its intracellular inhibitory domain. Also, we have developed a novel low molecular weight compound which can enhance the response of human B cells.

Rapid, Sustained B Cell Receptor Signaling in Lymphoma B Cells Differs from Normal Signaling in Tumor Infiltrating Nonmalignant B Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 283-283
Author(s):  
Jonathan M. Irish ◽  
Debra K. Czerwinski ◽  
Garry P. Nolan ◽  
Ronald Levy
Keyword(s):  
B Cells ◽  
Cell Signaling ◽  
B Cell ◽  
Peripheral Blood ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Life And Death ◽  
Heavy Chains ◽  
Bcr Signaling ◽  
Tumor Biopsies

Abstract The B cell receptor (BCR) drives life and death signaling throughout B cell development, and dysregulation of BCR signaling might be expected to play a role in aberrant proliferation of lymphoma B cells. We have previously used flow cytometry based cell signaling profiles to identify patterns of altered signaling in acute myeloid leukemia that were informative of clinical outcome (Irish et al., Cell, 2004). Here we used a similar signaling profiles approach to compare BCR signaling in normal and lymphoma B cells. However, in addition to comparing follicular lymphoma (FL) B cells with peripheral blood B cells from normal donors, we also interrogated signaling within individual non-tumor B cells infiltrating FL tumor biopsies. By staining for CD20 and BCR light chain isotype (κ vs. λ), we could distinguish tumor and normal B cells within each patient biopsy. Following crosslinking of BCR heavy chains (shared by tumor and non-tumor B cells), we measured phosphorylation of Syk and Btk proteins, as markers of early BCR signaling activity, and Erk1/2 and p38, as markers of downstream BCR signaling effector activity. The BCR signaling network in FL tumor B cells was activated more rapidly than infiltrating non-tumor B cells, achieved greater levels of per-cell signaling, and sustained high levels of signaling over a period of hours. In lymphoma B cells, BCR-mediated Btk and Erk1/2 phosphorylation could reach the normal maximum in as little as 4 minutes, which was much more rapid than the 30–60 minutes required for peak signaling in non-tumor B cells. Strikingly, the timing and magnitude of BCR pathway protein phosphorylation we measured in non-tumor B cells within tumor biopsies was the same as that of normal, mature B cells from peripheral blood. These results suggest that the altered BCR signaling we identified in lymphoma is cell-intrinsic and associated with lymphomagenesis, as opposed to being a general change in tumor microenviornment affecting all B cells within a biopsy. FL tumor B cells from different patients were distinguished by the degree and number of changes to BCR signaling, such that variable profiles of lymphoma signaling kinetics distinguished each patient from the consistent signaling of normal B cells. These results identify cell-intrinsic changes to BCR signaling that may contribute to immortalization of lymphoma B cells and suggest that single cell profiles could identify lymphoma specific BCR-mediated signaling responsible for clinical outcomes.

Targeting Early Events of B-Cell Receptor Signaling in Chronic Lymphocytic Leukemia: Suppressed Syk and PLCγ2 Activities Predict Apoptotic Response of Leukemic Cells to Dasatinib

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 5023-5023
Author(s):  
Y. Lynn Wang ◽  
Zibo Song ◽  
Pin Lu ◽  
John P. Leonard ◽  
Morton Coleman ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Kinase Inhibitor ◽  
Ex Vivo ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Leukemic Cells ◽  
Bcr Signaling

Abstract B cell receptor (BCR) signaling plays an essential role in the pathogenesis of chronic lymphocytic leukemia. In a subset of patients with a poor clinical outcome, BCR ligation leads to increased cell metabolism and cell survival (Cancer Research66, 7158–66, 2006). Based on these findings, we tested whether targeting BCR signaling with dasatinib, an inhibitor of Src kinase, would interfere with the signaling cascade and cause death of CLL B cells. CLL leukemic cells were isolated from 34 patients and were incubated with or without dasatinib at a low dose of 128 nM. Among 34 cases, viability of leukemic cells was reduced by 2% to 90%, with an average of ~50% reduction on day 4 of ex vivo culture. Further study showed that CLL B cells undergo death by apoptosis via the intrinsic pathway which involves the generation of reactive oxygen species. Analysis of the Src family kinases showed that phosphorylation of Src, Lyn and Hck was inhibited by dasatinib not only in those cases that responded to dasatinib with apoptosis, but also in those that did not respond well (&lt;20% apoptosis). Further analysis revealed that suppressed activity of two downstream molecules, Syk and PLC Statistical analysis showed a significant correlation between CLL dasatinib response and their IgVH mutation and ZAP70 status. Cases with worse prognoses by these criteria have a better response to the kinase inhibitor. Lastly, we have also found that ZAP70 positive cases showed a greater degree of PLC

The Human Pre-B Cell Receptor Signaling Cascade Is Regulated Via PI-3Kinase and MAPK Pathway

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1314-1314
Author(s):  
Kolandaswamy Anbazhagan ◽  
Vincent Fuentes ◽  
Eliane Bissac ◽  
Remy Nyga ◽  
Naomi Taylor ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cells ◽  
B Cell ◽  
Nuclear Translocation ◽  
Accessory Pathway ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Signaling Cascade ◽  
Down Regulation ◽  
Bcr Signaling

Abstract Abstract 1314 Background: Pre-B cell receptor (pre-BCR) constitutes a major check point in the early steps of mouse and human B cell development. Several functions have been attributed to this receptor which include a delivery of proliferation and survival signals, increased sensitivity to interleukin-7 (IL-7) and down modulation of recombinase activating genes (RAG) and surrogate light chain (SLC) encoding genes. Pre-BCR is also involved in shaping the VH repertoire and preventing autoimmunity. Finally, there is increasing evidence that pre-BCR might be implicated in leukemogenesis. Most of the functions of pre-BCR have been predicted based on studies in knockout mice and leukemic cell lines. In a previous study we have shown that pre-BCR aggregation resulted in the activation of src and Syk kinases which in turn activated the PI-3K/Akt, Btk, PLCγ-2 and Ras/MAPK. In this study, we examined the pre-BCR signalling cascade using human normal primary pre-B cells with a particular focus on transcription factors activation and Rag modulation and their regulatory aspects. Methods: Pre-B cells were sorted from adult human bone marrow samples, treated or not with inhibitors of Syk (BAY61–3606), Akt (LY294002) and MEKK1 (UO126) prior to crosslink the pre-BCR by means of F(ab')2 anti-μHC. The effect of Pre-BCR signaling was examined by quantifying the transcript levels of Rag1, Rag2, E2A, EBF1, Pax5, FoxO1 and FoxO3, IRF4/8. Activation of transcription factors such as NF-κB p50, c-Fos, IRF4 and FoxO3A, was assessed by analyzing their nuclear translocation by immunofluorescence microscopy. Results: We show that NF-κB p50 is translocated into nucleus within 3h after pre-BCR stimulation. Crosslinking of pre-BCR also resulted in an enhancement of nuclear c-Fos translocation. BAY61-3606 (Syk inhibitor) treatment resulted in complete apoptosis (100 % cell death within 48h). Although treatment of normal pre-B cells with LY294002 or U0126 did not alter cell survival, nuclear translocation of pre-BCR-induced p50 NF-κB was prevented by former and enhanced by later. Conversely, c-Fos nuclear expression was inhibited by U0126 and slightly but consistently enhanced by LY294002 in association with a decrease in its cytoplasmic location. Pre-BCR stimulation also induced IRF4 translocation to the nucleus. Pre-BCR stimulation also resulted in the down regulation of Rag1 (− 48 %, P<0.01), Pax5 (− 40%, P<0.01) and E2A (− 35 %, P< 0.01) transcripts, whereas EBF1 and FoxO1 and 3 expression remained unchanged. In LY294002-treated cells, Rag1/Rag2 expression was up regulated (+130%, P< 0.01 and +251%, P< 0.01, respectively) following pre-BCR crosslinking, whereas in the presence of U0126 the pre-BCR induced Rag1/Rag2 down modulation remained unchanged. Conclusion: Our results indicate that the pre-BCR has the potential to promote pre-B cell proliferation, survival and differentiation by activating NF-kB, c-Fos and IRF4. It also has the ability to protect pre-B cells from genome instability by down-regulating Rag1/2, probably through down modulation of Pax5 and E2A. We bring evidence that PI-3 K/Akt pathway plays a crucial role in the regulation of the pre-BCR signaling cascade and that Akt-mediated NF-kB and c-Fos activation is antagonized by MAPK. Up-regulation of Rag transcripts upon Akt inhibition suggests either a feed-back negative loop or a dual effect of pre-BCR on Rag expression with an Akt-dependent Rag down regulation and an accessory pathway that enhances Rag expression. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Silencing of B Cell Receptor Signals in Human Naive B Cells

2002 ◽  
Vol 196 (10) ◽  
pp. 1291-1305 ◽  
Author(s):  
Niklas Feldhahn ◽  
Ines Schwering ◽  
Sanggyu Lee ◽  
Maria Wartenberg ◽  
Florian Klein ◽  
...  
Keyword(s):  
Gene Expression ◽  
B Cells ◽  
B Cell ◽  
Cell Receptor ◽  
Memory B Cells ◽  
B Cell Receptor ◽  
Functional Assay ◽  
Hematopoietic Stem ◽  
Large Group ◽  
Bcr Signaling

To identify changes in the regulation of B cell receptor (BCR) signals during the development of human B cells, we generated genome-wide gene expression profiles using the serial analysis of gene expression (SAGE) technique for CD34+ hematopoietic stem cells (HSCs), pre-B cells, naive, germinal center (GC), and memory B cells. Comparing these SAGE profiles, genes encoding positive regulators of BCR signaling were expressed at consistently lower levels in naive B cells than in all other B cell subsets. Conversely, a large group of inhibitory signaling molecules, mostly belonging to the immunoglobulin superfamily (IgSF), were specifically or predominantly expressed in naive B cells. The quantitative differences observed by SAGE were corroborated by semiquantitative reverse transcription–polymerase chain reaction (RT-PCR) and flow cytometry. In a functional assay, we show that down-regulation of inhibitory IgSF receptors and increased responsiveness to BCR stimulation in memory as compared with naive B cells at least partly results from interleukin (IL)-4 receptor signaling. Conversely, activation or impairment of the inhibitory IgSF receptor LIRB1 affected BCR-dependent Ca2+ mobilization only in naive but not memory B cells. Thus, LIRB1 and IL-4 may represent components of two nonoverlapping gene expression programs in naive and memory B cells, respectively: in naive B cells, a large group of inhibitory IgSF receptors can elevate the BCR signaling threshold to prevent these cells from premature activation and clonal expansion before GC-dependent affinity maturation. In memory B cells, facilitated responsiveness upon reencounter of the immunizing antigen may result from amplification of BCR signals at virtually all levels of signal transduction.

scholarly journals Mst1 positively regulates B-cell receptor signaling via CD19 transcriptional levels

2016 ◽  
Vol 1 (3) ◽  
pp. 219-230 ◽  
Author(s):  
Xiaoming Bai ◽  
Lu Huang ◽  
Linlin Niu ◽  
Yongjie Zhang ◽  
Jinzhi Wang ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Lipid Bilayers ◽  
Messenger Rna ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Developmental Defect ◽  
Hippo Signaling ◽  
Bcr Signaling ◽  
Genetically Manipulated

Abstract As a key regulator of hippo signaling pathway, Mst kinases are emerging as one of the key signaling molecules that influence cell proliferation, organ size, cell migration, and cell polarity. In B lymphocytes, Mst1 deficiency causes the developmental defect of marginal zone (MZ) B cells, but how Mst1 regulates B-cell receptor (BCR) activation and differentiation remains elusive. Using genetically manipulated mouse models and total internal reflection fluorescence microscopy, we have demonstrated that Mst1 positively regulates BCR signaling via modulating CD19 transcriptional levels. Consistent with this, Mst1-deficient mice exhibited reduced BCR signaling, which is concurrent with defective BCR clustering and B-cell spreading on stimulatory lipid bilayers. The disruption of CD19-mediated Btk signaling by Mst1 deficiency leads to the severe defect in the differentiation of MZ and germinal center B cells. Mechanistic analysis showed that Mst1 upregulates the messenger RNA level of CD19 via regulating the transcriptional factor TEAD2 that directly binds to the consensus motif in the 3′ untranslated region of cd19. Overall, our results reveal a new function of Mst1 in B cells and the mechanism by which Mst1 regulates the activation and differentiation of peripheral B cells.

scholarly journals Altered B-cell receptor signaling kinetics distinguish human follicular lymphoma B cells from tumor-infiltrating nonmalignant B cells

Blood ◽  
2006 ◽  
Vol 108 (9) ◽  
pp. 3135-3142 ◽  
Author(s):  
Jonathan M. Irish ◽  
Debra K. Czerwinski ◽  
Garry P. Nolan ◽  
Ronald Levy
Keyword(s):  
B Cells ◽  
Cell Signaling ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphoma Cells ◽  
B Lymphoma Cells ◽  
Bcr Signaling ◽  
B Cell Subsets

Abstract The B-cell receptor (BCR) transmits life and death signals throughout B-cell development, and altered BCR signaling may be required for survival of B-lymphoma cells. We used single-cell signaling profiles to compare follicular lymphoma (FL) B cells and nonmalignant host B cells within individual patient biopsies and identified BCR-mediated signaling events specific to lymphoma B cells. Expression of CD20, Bcl-2, and BCR light chain isotype (κ or λ) distinguished FL tumor B-cell and nontumor host B-cell subsets within FL patient biopsies. BCR-mediated signaling via phosphorylation of Btk, Syk, Erk1/2, and p38 occurred more rapidly in tumor B cells from FL samples than in infiltrating nontumor B cells, achieved greater levels of per-cell signaling, and sustained this level of signaling for hours longer than nontumor B cells. The timing and magnitude of BCR-mediated signaling in nontumor B cells within an FL sample instead resembled that observed in mature B cells from the peripheral blood of healthy subjects. BCR signaling pathways that are potentiated specifically in lymphoma cells should provide new targets for therapeutic attention.

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