Overexpression of TOSO in CLL Is Triggered by B-Cell Receptor Signaling and Associated with Progressive Disease.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2070-2070
Author(s):  
Christian P Pallasch ◽  
Alexandra Schulz ◽  
Nadine Kutsch ◽  
Janine Schwamb ◽  
Susanne Hagist ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
B Cells ◽  
B Cell ◽  
Healthy Donor ◽  
Progressive Disease ◽  
Marginal Zone ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Quantitative Real Time Pcr ◽  
Follicular Lymphomas

Abstract Resistance towards apoptotic stimuli mediated by overexpression of antiapoptotic factors or extracellular survival signals like B-cell receptor stimulation (BCR) are considered to be responsible for accumulation of malignant B cells in CLL . TOSO, also known as Fas-inhibitory molecule 3 (FAIM3), was identified as overexpressed candidate gene in CLL based on re-evaluation of publicly available microarray data sets. Based on primary CLL samples from 106 patients, TOSO expression was compared to healthy donor B cells using quantitative real-time PCR, western-blot, flow cytometry and immunohistochemistry. To reveal underlying mechanisms of TOSO overexpression, B-cell receptor (BCR) and CD40Ligand stimulation as well as bone marrow stroma cell co-incubation was performed. Apoptotic resistance was assessed by annexin V/7-AAD flow cytometry in context of CH11-Fas-agonistic antibody. TOSO was identified to exhibit elevated relative expression of 6.8 compared to healthy donor B cells using quantitative real-time PCR (p=0.004). High levels of TOSO expression in CLL correlated with high leukocyte count, advanced Binet stage, previous need for chemotherapy and unmutated IgVH status. CD38+ CLL subsets harboring proliferative activity showed significantly enhanced TOSO expression. Immunohistochemistry revealed upregulation of TOSO in lymph nodes of CLL patients. In lymph nodes derived from healthy donors TOSO was detected in single plasmocytoid cells within the germinal center and in the marginal zone. No specific staining was seen in follicular lymphomas. CLL-specific upregulation of TOSO was confirmed by RT-PCR, samples of follicular lymphomas, DLBCL, marginal zone lymphoma and Hodgkin cell lines did not reveal TOSO up-regulation. We evaluated functional mechanisms of aberrant TOSO expression in CLL cells and identified TOSO expression significantly being induced by BCR-stimulation compared to control cells (relative expression (RE) 8.25 vs. 4.86, p=0.013). In contrast, CD40L signaling significantly reduced TOSO expression (RE 2.60; p=0.007). Spontaneous apoptosis of CLL cells was significantly reduced by BCR-stimulus, in CD40Ligandstimulated CLL samples a slight sensitization towards Fas-mediated apoptosis was seen. In summary, we show that the anti-apoptotic factor TOSO is associated with progressive disease and enhanced in the proliferative CD38+ cell subset. Both association with unmutated IgVH and the specific induction of TOSO via the BCR suggest autoreactive BCR signaling as a key mediator of apoptosis resistance in CLL. Down-regulation of TOSO by CD40Ligand in the context of CD40Ligand-mediated Fas-sensitization of CLL reveal TOSO as a new anti-apoptotic factor in CLL. CLL-specific over-expression of the transmembrane protein might further offer new therapeutic strategies in CLL treatment.

scholarly journals Selection of stereotyped VH81X-μH chains via pre-B cell receptor early in ontogeny and their conservation in adults by marginal zone B cells

2005 ◽  
Vol 17 (7) ◽  
pp. 857-867 ◽  
Author(s):  
Yohei Kawano ◽  
Soichiro Yoshikawa ◽  
Yoshiyuki Minegishi ◽  
Hajime Karasuyama
Keyword(s):  
B Cells ◽  
B Cell ◽  
Marginal Zone ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Marginal Zone B Cells ◽  
Selection Of

A “Universal” Lectin-Mediated Interaction Drives B-Cell Receptor Signaling In Primary Follicular Lymphoma Cells

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4291-4291
Author(s):  
Graham Packham ◽  
Serge Krysov ◽  
Vania Coelho ◽  
Peter Johnson ◽  
Freda K. Stevenson
Keyword(s):  
Flow Cytometry ◽  
B Cells ◽  
Follicular Lymphoma ◽  
B Cell ◽  
Kinase Inhibitors ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Malignant Cells ◽  
Glycosylation Sites ◽  
Mannose Binding

Abstract B-cell receptor (BCR) signaling has been identified as a critical driver of B-cell malignancies and as a target for therapeutic attack. Clinical responses to novel inhibitors of BCR-associated kinases have been relatively modest in follicular lymphoma (FL) and a more detailed knowledge of BCR function in these cells is required. Surface Ig (sIg) is unusual in FL since variable regions contain N-linked glycosylation sites which are introduced by somatic mutation. These are rarely found in normal B cells, indicating strong positive selective pressure in malignant cells. Remarkably, added sugars terminate with high mannose suggesting a novel function for FL BCRs in binding to, and perhaps receiving stimulation via, microenvironmental mannose-binding lectins. In previous studies we demonstrated that candidate mannose-binding lectins, including DC-SIGN, promote intracellular calcium mobilization in primary FL cells, but not normal B cells. In this work, we characterized in more detail the response of FL cells to DC-SIGN and its inhibition by BCR-targeted kinase inhibitors. Initial studies using immunoblotting demonstrated that, like anti-Ig antibodies, DC-SIGN caused increased phosphorylation of the downstream kinases AKT and ERK in primary FL samples. DC-SIGN treatment also resulted in increased expression of the MYC oncoprotein in a subset of samples. In contrast to FL samples, DC-SIGN did not increase AKT or ERK phosphorylation in normal B cells although anti-IgM induced strong responses in these cells. Overall, responses to DC-SIGN were similar in IgM+ and IgG+ FL samples. Flow cytometry demonstrated that DC-SIGN also increased phosphorylation of proximal signaling kinases (SYK and BTK), as well as phosphorylation of PLCγ2 in FL samples, and that DC-SIGN-induced signaling occurred within the CD19+BCL2+ malignant cells. Flow cytometry also revealed intraclonal variation in responses to DC-SIGN and, like responses to anti-Ig, DC-SIGN responses were strongest in a sub-population of malignant cells with high CD20 expression. Finally, we demonstrated that tamatinib, the active form of the SYK inhibitor pro-drug fostamatinib, significantly inhibited phosphorylation of ERK and PLCγ2 induced by either anti-Ig or DC-SIGN. Overall our results are consistent with the hypothesis that N-linked glycosylation sites, introduced into BCRs by somatic mutation, are selected for in FL since they confer signaling responsiveness following binding of environmental lectins. Like canonical antigen signaling, lectin-mediated signaling via the BCR appears to be susceptible to therapeutic blockade using kinase inhibitors. However, further analysis of this novel lectin-mediated pathway may reveal novel targets for optimal therapeutic attack. Disclosures: No relevant conflicts of interest to declare.

Overexpression of TOSO in CLL is triggered by B-cell receptor signaling and associated with progressive disease

Blood ◽  
2008 ◽  
Vol 112 (10) ◽  
pp. 4213-4219 ◽  
Author(s):  
Christian Philipp Pallasch ◽  
Alexandra Schulz ◽  
Nadine Kutsch ◽  
Janine Schwamb ◽  
Susanne Hagist ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Progressive Disease ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Apoptosis Resistance ◽  
B Cell Receptor Signaling ◽  
Microarray Datasets ◽  
Functional Mechanisms

Abstract Resistance toward apoptotic stimuli mediated by overexpression of antiapoptotic factors or extracellular survival signals is considered to be responsible for accumulation of malignant B cells in chronic lymphocytic leukemia (CLL). TOSO was identified as overexpressed candidate gene in CLL, applying unit-transformation assays of publicly available microarray datasets. Based on CLL samples from 106 patients, TOSO was identified to exhibit elevated relative expression (RE) of 6.8 compared with healthy donor B cells using quantitative real-time polymerase chain reaction (PCR; P = .004). High levels of TOSO expression in CLL correlated with high leukocyte count, advanced Binet stage, previous need for chemotherapy, and unmutated IgVH status. CD38+ CLL subsets harboring proliferative activity showed enhanced TOSO expression. We evaluated functional mechanisms of aberrant TOSO expression and identified TOSO expression significantly induced by B-cell receptor (BCR) stimulation compared with control cells (RE; 8.25 vs 4.86; P = .01). In contrast, CD40L signaling significantly reduced TOSO expression (RE, 2.60; P = .01). In summary, we show that the antiapoptotic factor TOSO is associated with progressive disease and enhanced in the proliferative CD38+ CLL subset. Both association with unmutated IgVH and the specific induction of TOSO via the BCR suggest autoreactive BCR signaling as a key mediator of apoptosis resistance in CLL.

scholarly journals Delivery of B Cell Receptor–internalized Antigen to Endosomes and Class II Vesicles

1997 ◽  
Vol 186 (8) ◽  
pp. 1299-1306 ◽  
Author(s):  
James R. Drake ◽  
Paul Webster ◽  
John C. Cambier ◽  
Ira Mellman
Keyword(s):  
B Cells ◽  
B Cell ◽  
Antigen Processing ◽  
Specific Antigen ◽  
Cell Receptor ◽  
Class Ii ◽  
Time Frame ◽  
B Cell Receptor ◽  
Subcellular Compartment ◽  
Endocytic Vesicles

B cell receptor (BCR)-mediated antigen processing is a mechanism that allows class II–restricted presentation of specific antigen by B cells at relatively low antigen concentrations. Although BCR-mediated antigen processing and class II peptide loading may occur within one or more endocytic compartments, the functions of these compartments and their relationships to endosomes and lysosomes remain uncertain. In murine B cells, at least one population of class II– containing endocytic vesicles (i.e., CIIV) has been identified and demonstrated to be distinct both physically and functionally from endosomes and lysosomes. We now demonstrate the delivery of BCR-internalized antigen to CIIV within the time frame during which BCR-mediated antigen processing and formation of peptide–class II complexes occurs. Only a fraction of the BCR-internalized antigen was delivered to CIIV, with the majority of internalized antigen being delivered to lysosomes that are largely class II negative. The extensive colocalization of BCR-internalized antigen and newly synthesized class II molecules in CIIV suggests that CIIV may represent a specialized subcellular compartment for BCR-mediated antigen processing. Additionally, we have identified a putative CIIV-marker protein, immunologically related to the Igα subunit of the BCR, which further illustrates the unique nature of these endocytic vesicles.

scholarly journals Targeted inhibition of eIF4A suppresses B-cell receptor-induced translation and expression of MYC and MCL1 in chronic lymphocytic leukemia cells

2021 ◽  
Author(s):  
Sarah Wilmore ◽  
Karly-Rai Rogers-Broadway ◽  
Joe Taylor ◽  
Elizabeth Lemm ◽  
Rachel Fell ◽  
...  
Keyword(s):  
B Cells ◽  
Chronic Lymphocytic Leukemia ◽  
B Cell ◽  
Mrna Translation ◽  
Cell Receptor ◽  
Memory B Cells ◽  
B Cell Receptor ◽  
Lymphocytic Leukemia ◽  
Mrna Levels ◽  
Mrna Stabilization

AbstractSignaling via the B-cell receptor (BCR) is a key driver and therapeutic target in chronic lymphocytic leukemia (CLL). BCR stimulation of CLL cells induces expression of eIF4A, an initiation factor important for translation of multiple oncoproteins, and reduces expression of PDCD4, a natural inhibitor of eIF4A, suggesting that eIF4A may be a critical nexus controlling protein expression downstream of the BCR in these cells. We, therefore, investigated the effect of eIF4A inhibitors (eIF4Ai) on BCR-induced responses. We demonstrated that eIF4Ai (silvestrol and rocaglamide A) reduced anti-IgM-induced global mRNA translation in CLL cells and also inhibited accumulation of MYC and MCL1, key drivers of proliferation and survival, respectively, without effects on upstream signaling responses (ERK1/2 and AKT phosphorylation). Analysis of normal naïve and non-switched memory B cells, likely counterparts of the two main subsets of CLL, demonstrated that basal RNA translation was higher in memory B cells, but was similarly increased and susceptible to eIF4Ai-mediated inhibition in both. We probed the fate of MYC mRNA in eIF4Ai-treated CLL cells and found that eIF4Ai caused a profound accumulation of MYC mRNA in anti-IgM treated cells. This was mediated by MYC mRNA stabilization and was not observed for MCL1 mRNA. Following drug wash-out, MYC mRNA levels declined but without substantial MYC protein accumulation, indicating that stabilized MYC mRNA remained blocked from translation. In conclusion, BCR-induced regulation of eIF4A may be a critical signal-dependent nexus for therapeutic attack in CLL and other B-cell malignancies, especially those dependent on MYC and/or MCL1.

scholarly journals Characterization of B-cell Receptor Heavy Chain Complementarity-determining Region 3 Repertoire Based on the Differences of Symbiotic Microorganisms in the Gut of Mice

2021 ◽  
Author(s):  
Jun Li ◽  
Yurong Pan ◽  
Qingqing Ma ◽  
Long Ma ◽  
Bin Shi ◽  
...  
Keyword(s):  
Small Intestine ◽  
B Cells ◽  
B Cell ◽  
Heavy Chain ◽  
Intestinal Microflora ◽  
Cell Receptor ◽  
Memory B Cells ◽  
B Cell Receptor ◽  
Positive Effects ◽  
Significant Difference

Abstract Background Colonization of gut microorganism is related to maturation of B cells in peripheral immune organs. This study aims to investigate the effect of intestinal microflora in Germ-free (GF), Specific Pathogen-free (SPF) and Clean (CL) BALB/C mice to small intestine total B-cell and memory B-cell receptor (BCR) complementary-determining region 3 (CDR3) repertoire. Results The composition and characteristics of intestinal microflora were analyzed by 16S rDNA sequencing. Genomic DNA extracted from small intestine tissue and memory B-cells of GF, SPF and CL mice were conducted via high-throughput DNA sequencing methods. As expected, significant differences of gut microflora diversity were observed in the three mice groups. CL group showed the most diversity, followed by SPF group, and GF group had the lowest diversity. Moreover, anormogenesis of intestinal lymphoid tissue were observed in GF mice. Diversity of the BCR heavy chain CDR3 repertoire in memory B cells were significant difference among three groups, but not in total B cells. The nucleotide polymorphism, usage frequency of gene segments (V, D, J, V–J gene segments) and amino acid of total B cells and memory B cells CDR3 were comparable among three mice groups, and there was significant difference between CL and GF mice groups. Conclusions The results of this study advocate that the colonization of intestinal microorganisms affect the diversity of B cells CDR3 repertoire. Elucidating mechanism of microbiome participated in the function of intestinal mucosal immune system may have positive effects on human health, and it requires further investigation.

scholarly journals Signatures of B Cell Receptor Repertoire Following Pneumocystis Infection

2021 ◽  
Vol 12 ◽  
Author(s):  
Han Sun ◽  
Hu-Qin Yang ◽  
Kan Zhai ◽  
Zhao-Hui Tong
Keyword(s):  
B Cells ◽  
B Cell ◽  
Single Cell ◽  
Plasma Cells ◽  
Somatic Hypermutation ◽  
Cell Receptor ◽  
B Cell Receptor ◽  
Receptor Repertoire ◽  
Elevated Expression ◽  
Pneumocystis Infection

B cells play vital roles in host defense against Pneumocystis infection. However, the features of the B cell receptor (BCR) repertoire in disease progression remain unclear. Here, we integrated single-cell RNA sequencing and single-cell BCR sequencing of immune cells from mouse lungs in an uninfected state and 1–4 weeks post-infection in order to illustrate the dynamic nature of B cell responses during Pneumocystis infection. We identified continuously increased plasma cells and an elevated ratio of (IgA + IgG) to (IgD + IgM) after infection. Moreover, Pneumocystis infection was associated with an increasing naïve B subset characterized by elevated expression of the transcription factor ATF3. The proportion of clonal expanded cells progressively increased, while BCR diversity decreased. Plasma cells exhibited higher levels of somatic hypermutation than naïve B cells. Biased usage of V(D)J genes was observed, and the usage frequency of IGHV9-3 rose. Overall, these results present a detailed atlas of B cell transcriptional changes and BCR repertoire features in the context of Pneumocystis infection, which provides valuable information for finding diagnostic biomarkers and developing potential immunotherapeutic targets.

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