RÉSUMÉ
Humoral immune responses depend on B cells encountering antigen (Ag) in lymph nodes (LNs) draining infection sites, getting activated, interacting with different cells, proliferating and differentiating into antibody (Ab)-secreting cells. Each of these events occurs in distinct LN sub-compartments, requiring the migration of B cells from niche to niche in a fast and tightly coordinated fashion. While some of the rules that characterize B cell behavior in secondary lymphoid organs have been elucidated at the population level, we have only limited knowledge of the precise dynamics of B cell interactions with different kinds of LN cells at the single-cell level. Here, we describe in detail an intravital microscopy technique that allows the analysis of B cell dynamic behavior in the popliteal lymph node of anesthetized mice at high spatial and temporal resolution. A detailed understanding of the spatiotemporal dynamics of B cells within secondary lymphoid organs may lead to novel, rational vaccine strategies aimed at inducing rapid and long-lived humoral immune responses.
Sujet(s)
Lymphocytes B/cytologie , Lymphocytes B/physiologie , Mouvement cellulaire , Suivi cellulaire/méthodes , Microscopie intravitale/méthodes , Noeuds lymphatiques/cytologie , Animaux , Souris , Souris de lignée C57BL , Souris transgéniques , Protéines proto-oncogènes c-bcr/physiologieRÉSUMÉ
Due to the apoptosis-prone nature of primary germinal center B (GCB) cells, it remains a huge challenge to dissect signals that guide their differentiation towards memory B cells and plasma cells in vitro. Here we show that the murine lymphoma cell line A20 resembles primary GCB cells in expression of GC-specific surface markers and the master transcription factor BCL6 and may serve as a useful system to model certain GCB cell behaviors in vitro. Using these cells, we found that both CD40 and B cell receptor (BCR) signaling are able to drive BCL6 downregulation, which is a prerequisite of post-GC B-cell differentiation. Under the steady state, BCL6 is constantly and rapidly degraded in A20 cells by the proteasome in a strictly FBXO11-dependent manner. This process can be further enhanced by signals downstream of the BCR. Both CD40 and BCR stimulation can upregulate IRF4, a transcription factor that suppresses BCL6 expression. However, only BCR signaling downregulate PAX5 and BACH2, two transcription factors that help maintain the GCB identity. Together, these results validate the A20 cell line as an experimental system suitable for studying regulation of BCL6 and potentially other transcription factors relevant to post-GC fate determination, and they support that combined signaling from BCR and CD40 receptors would drive termination of the GC program.
Sujet(s)
Centre germinatif/physiologie , Transduction du signal/physiologie , Facteurs de transcription/physiologie , Animaux , Antigènes CD40/physiologie , Lignée cellulaire tumorale , Cellules cultivées , Protéines F-box/physiologie , Facteurs de régulation d'interféron/physiologie , Lymphome malin non hodgkinien/anatomopathologie , Souris , Souris transgéniques , Proteasome endopeptidase complex/métabolisme , Protéines proto-oncogènes c-bcl-6/physiologie , Protéines proto-oncogènes c-bcr/physiologie , Organismes exempts d'organismes pathogènes spécifiquesRÉSUMÉ
Cortactin (CTTN) is a substrate of the Src kinase Lyn that is known to play an actin cytoskeletal regulatory role involved in cell migration and cancer progression following its phosphorylation at Y421. We recently demonstrated that Cortactin is overexpressed in patients with chronic lymphocytic leukaemia (CLL). This work was aimed at defining the functional role of Cortactin in these patients. We found that Cortactin is variably expressed in CLL patients both in the peripheral blood and lymph nodes and that its expression correlates with the release of matrix metalloproteinase 9 (MMP-9) and the motility of neoplastic cells. Cortactin knockdown, by siRNA, induced a reduction in MMP-9 release as well as a decrease of migration capability of leukaemic B cells in vitro, also after chemotactic stimulus. Furthermore, Cortactin phosphorylation was lowered by the Src kinase-inhibitor PP2 with a consequent decrease of MMP-9 release in culture medium. An impaired migration, as compared to control experiments without Cortactin knockdown, was observed following CXCL12 triggering. Reduced Cortactin expression and phosphorylation were also detected both in vivo and in vitro after treatment with Ibrutinib, a Btk inhibitor. Our results highlight the role of Cortactin in CLL as a check-point molecule between the BCR and CXCR4 signalling pathways.
Sujet(s)
Points de contrôle du cycle cellulaire/physiologie , Cortactine/physiologie , Leucémie chronique lymphocytaire à cellules B/métabolisme , Protéines proto-oncogènes c-bcr/physiologie , Récepteurs CXCR4/physiologie , Adénine/analogues et dérivés , Adulte , Agammaglobulinaemia tyrosine kinase , Sujet âgé , Sujet âgé de 80 ans ou plus , Antinéoplasiques/pharmacologie , Antinéoplasiques/usage thérapeutique , Mouvement cellulaire/physiologie , Femelle , Humains , Leucémie chronique lymphocytaire à cellules B/traitement médicamenteux , Leucémie chronique lymphocytaire à cellules B/anatomopathologie , Mâle , Matrix metalloproteinase 9/métabolisme , Adulte d'âge moyen , Protéines tumorales/physiologie , Phosphorylation/effets des médicaments et des substances chimiques , Pipéridines , Protein-tyrosine kinases/antagonistes et inhibiteurs , Pyrazoles/pharmacologie , Pyrazoles/usage thérapeutique , Pyrimidines/pharmacologie , Pyrimidines/usage thérapeutique , Transduction du signal/physiologie , Cellules cancéreuses en culture , src-Family kinases/physiologieRÉSUMÉ
SOS is a key activator of the small GTPase Ras. In cells, SOS-Ras signaling is thought to be initiated predominantly by membrane recruitment of SOS via the adaptor Grb2 and balanced by rapidly reversible Grb2-SOS binding kinetics. However, SOS has multiple protein and lipid interactions that provide linkage to the membrane. In reconstituted-membrane experiments, these Grb2-independent interactions were sufficient to retain human SOS on the membrane for many minutes, during which a single SOS molecule could processively activate thousands of Ras molecules. These observations raised questions concerning how receptors maintain control of SOS in cells and how membrane-recruited SOS is ultimately released. We addressed these questions in quantitative assays of reconstituted SOS-deficient chicken B-cell signaling systems combined with single-molecule measurements in supported membranes. These studies revealed an essentially one-way trafficking process in which membrane-recruited SOS remains trapped on the membrane and continuously activates Ras until being actively removed via endocytosis.
Sujet(s)
Protéine SOS1/physiologie , Protéines G ras/métabolisme , Régulation allostérique , Animaux , Membrane cellulaire/métabolisme , Poulets , Endocytose , Activation enzymatique , Humains , Cellules Jurkat , Cinétique , Double couche lipidique/composition chimique , Système de signalisation des MAP kinases , Liaison aux protéines , Domaines protéiques , Transport des protéines , Protéines proto-oncogènes c-bcr/physiologie , Protéine SOS1/composition chimique , Protéines G ras/composition chimiqueRÉSUMÉ
B-cell receptor (BCR) signaling is crucial for the survival of normal and neoplastic B cells, and inhibitors targeting BCR signaling pathways have shown promising therapeutic outcomes for patients with B-cell lymphomas. In the current study, we analyzed de novo diffuse large B-cell lymphoma without BCR expression (DLBCL, BCR) in 25 cases to determine the BCR/phosphatidylinositol-3-kinase/AKT (BCR/PI3K/AKT) signaling status, clinicopathologic features, and underlying causes leading to the loss of BCR. On the basis of clinical features, 15 (60%) DLBCL, BCR patients were classified into the low-risk group, and 18 (86%) experienced complete remission. Morphologically and immunophenotypically, DLBCL, BCR demonstrated centroblastic cytology (21/25, 84%) and germinal center B-cell-like cell origin (18/25, 72%). Other components in BCR complexity remained intact, on the basis of immunohistochemical findings. Epstein-Barr virus infection, deficiency in B-lineage transcription factors (PAX5, Oct-2, and Bob.1), and oncogene rearrangement did not seem to be associated with BCR loss. The activated form of signaling proteins (pSYK and pAKT) involved in the BCR/PI3K/AKT pathway were expressed at low levels in DLBCL, BCR tissue. In vitro validation revealed that in DLBCL, BCR cell lines, the BCR/PI3K/AKT pathway did not respond to BCR stimulation or inhibition. Our findings suggest that DLBCL, BCR was characterized by a silent BCR/PI3K/AKT pathway, germinal center phenotype, and low risk and may not be a candidate for BCR-targeted therapies.
Sujet(s)
Centre germinatif , Lymphome B diffus à grandes cellules/classification , Lymphome B diffus à grandes cellules/anatomopathologie , Protéine oncogène v-akt/physiologie , Phosphatidylinositol 3-kinases/physiologie , Protéines proto-oncogènes c-bcr/physiologie , Transduction du signal , Adulte , Sujet âgé , Sujet âgé de 80 ans ou plus , Femelle , Régulation de l'expression des gènes tumoraux , Humains , Lymphome B diffus à grandes cellules/génétique , Mâle , Adulte d'âge moyen , Phénotype , Appréciation des risques , Jeune adulteRÉSUMÉ
We previously demonstrated that c-Jun N-terminal kinase (JNK) phosphorylates serine 62 of Bcl-xL to induce the degradation of Bcl-xL and apoptosis in WEHI-231 cells upon BCR crosslinking. In order to elucidate the regulatory mechanisms underlying the phosphorylation of Bcl-xL, we prepared an assay system in which JNK phosphorylated Bcl-xL in HEK293T cells. Consequently, we found that a signal transduction molecule, alpha4, enhanced the phosphorylation of Bcl-xL by JNK, while the co-expression of C-terminal alpha4 (220-340) diminished the phosphorylation of Bcl-xL induced by JNK. Furthermore, full-length alpha4 associated with both JNK and Bcl-xL, whereas C-terminal alpha4 (220-340) associated only with Bcl-xL, not JNK. In addition, WEHI-231 cells transfected with the cDNA of C-terminal alpha4 (220-340) exhibited decreased phosphorylation of Bcl-xL and stronger resistance to apoptosis induced by BCR crosslinking. These results indicate that alpha4 is an important regulatory molecule of apoptosis induced by BCR crosslinking in WEHI-231 cells and that C-terminal alpha4 (220-340) functions as a dominant negative form.
Sujet(s)
Protéines et peptides de signalisation intracellulaire/physiologie , Précurseurs lymphoïdes B/physiologie , Protéines proto-oncogènes c-bcr/physiologie , Protéine bcl-X/métabolisme , Protéines adaptatrices de la transduction du signal , Apoptose , Cellules HEK293 , Humains , Chaperons moléculaires , Phosphorylation , Maturation post-traductionnelle des protéinesRÉSUMÉ
OBJECTIVE: B cells play an important role in the pathogenesis of autoimmune diseases. The role of Bruton's tyrosine kinase (Btk) in cytokine-induced human B cell differentiation and class-switch recombination remains incompletely defined. This study analysed the effect of Btk on human activated B cells. METHODS: Purified B cells from healthy subjects were stimulated with B cell receptor (BCR) and other stimuli with or without a Btk inhibitor and gene expression was measured. The B cell line BJAB was used to assess Btk-associated signalling cascades. Phosphorylated Btk (p-Btk) in peripheral blood B cells obtained from 10 healthy subjects and 41 patients with RA was measured by flow cytometry and compared with patient backgrounds. RESULTS: IL-21 signalling, in concert with BCR, CD40 and BAFF signals, led to robust expression of differentiation- and class-switch DNA recombination-related genes and IgG production in human B cells, all of which were significantly suppressed by the Btk inhibitor. Although phosphorylation of STAT1 and STAT3 was induced by co-stimulation with IL-21, BCR and CD40, STAT1 phosphorylation in the nucleus, but not in the cytoplasm, was exclusively impaired by Btk blockade. High levels of p-Btk were noted in B cells of RA patients compared with controls and they correlated significantly with titres of RF among RF-positive patients. CONCLUSION: The findings elucidate a model in which Btk not only plays a fundamental role in the regulation of BCR signalling, but may also mediate crosstalk with cytokine signalling pathways through regulation of IL-21-induced phosphorylation of STAT1 in the nuclei of human B cells. Btk appears to have pathological relevance in RA.
Sujet(s)
Facteur d'activation des lymphocytes B/physiologie , Lymphocytes B/physiologie , Antigènes CD40/physiologie , Interleukines/physiologie , Protein-tyrosine kinases/physiologie , Protéines proto-oncogènes c-bcr/physiologie , Transduction du signal/physiologie , Agammaglobulinaemia tyrosine kinase , Sujet âgé , Polyarthrite rhumatoïde/métabolisme , Polyarthrite rhumatoïde/anatomopathologie , Polyarthrite rhumatoïde/physiopathologie , Lymphocytes B/effets des médicaments et des substances chimiques , Lymphocytes B/anatomopathologie , Antigènes CD40/pharmacologie , Études cas-témoins , Différenciation cellulaire/effets des médicaments et des substances chimiques , Lignée cellulaire , Cellules cultivées , Femelle , Techniques de knock-down de gènes , Humains , Techniques in vitro , Interleukines/pharmacologie , Mâle , Adulte d'âge moyen , Phosphorylation , Protein-tyrosine kinases/génétique , Protéines proto-oncogènes c-bcr/pharmacologie , Facteur rhumatoïde/métabolisme , Facteur de transcription STAT-1/métabolismeRÉSUMÉ
BCR-JAK2 is an infrequent gene fusion found in chronic/acute, myeloid/lymphoid Philadelphia chromosome-negative leukaemia. In this study, we demonstrated that in vivo expression of BCR-JAK2 in mice induces neoplasia, with fatal consequences. Transplantation of BCR-JAK2 bone marrow progenitors promoted splenomegaly, with megakaryocyte infiltration and elevated leukocytosis of myeloid origin. Analysis of peripheral blood revealed the presence of immature myeloid cells, platelet aggregates and ineffective erythropoiesis. A possible molecular mechanism for these observations involved inhibition of apoptosis by deregulated expression of the anti-apoptotic mediator Bcl-xL and the serine/threonine kinase Pim1. Together, these data provide a suitable in vivo molecular mechanism for leukaemia induction by BCR-JAK2 that validates the use of this model as a relevant preclinical tool for the design of new targeted therapies in Philadelphia chromosome-negative leukaemia involving BCR-JAK2-driven activation of the JAK2 pathway.
Sujet(s)
Kinase Janus-2/physiologie , Leucémie myéloïde chronique atypique BCR-ABL négative/génétique , Protéines proto-oncogènes c-bcr/physiologie , Animaux , Femelle , Réarrangement des gènes , Transplantation de cellules souches hématopoïétiques/méthodes , Cellules souches hématopoïétiques/physiologie , Kinase Janus-2/génétique , Leucémie myéloïde chronique atypique BCR-ABL négative/mortalité , Hyperleucocytose/étiologie , Mâle , Souris de lignée BALB C , Transplantation tumorale , Protéines proto-oncogènes c-bcr/génétique , Retroviridae , Facteur de transcription STAT-5/métabolisme , Splénomégalie/étiologie , Transduction génétique/méthodes , TransgènesRÉSUMÉ
The activity of protein kinases is regulated by multiple molecular mechanisms, and their disruption is a common driver of oncogenesis. A central and almost universal control element of protein kinase activity is the activation loop that utilizes both conformation and phosphorylation status to determine substrate access. In this study, we use recombinant Abl tyrosine kinases and conformation-specific kinase inhibitors to quantitatively analyse structural changes that occur after Abl activation. Allosteric SH2-kinase domain interactions were previously shown to be essential for the leukemogenesis caused by the Bcr-Abl oncoprotein. We find that these allosteric interactions switch the Abl activation loop from a closed to a fully open conformation. This enables the trans-autophosphorylation of the activation loop and requires prior phosphorylation of the SH2-kinase linker. Disruption of the SH2-kinase interaction abolishes activation loop phosphorylation. Our analysis provides a molecular mechanism for the SH2 domain-dependent activation of Abl that may also regulate other tyrosine kinases.
Sujet(s)
Protéines oncogènes v-abl/physiologie , Domaine d'homologie SRC/physiologie , Activation enzymatique/physiologie , Protéines de fusion bcr-abl/physiologie , Protéines oncogènes v-abl/métabolisme , Phosphorylation , Structure tertiaire des protéines , Protéines proto-oncogènes c-bcr/physiologieRÉSUMÉ
The small GTPase Rac1 orchestrates actin-dependent remodeling essential for numerous cellular processes including synapse development. While precise spatiotemporal regulation of Rac1 is necessary for its function, little is known about the mechanisms that enable Rac1 activators (GEFs) and inhibitors (GAPs) to act in concert to regulate Rac1 signaling. Here, we identify a regulatory complex composed of a Rac-GEF (Tiam1) and a Rac-GAP (Bcr) that cooperate to control excitatory synapse development. Disruption of Bcr function within this complex increases Rac1 activity and dendritic spine remodeling, resulting in excessive synaptic growth that is rescued by Tiam1 inhibition. Notably, EphB receptors utilize the Tiam1-Bcr complex to control synaptogenesis. Following EphB activation, Tiam1 induces Rac1-dependent spine formation, whereas Bcr prevents Rac1-mediated receptor internalization, promoting spine growth over retraction. The finding that a Rac-specific GEF/GAP complex is required to maintain optimal levels of Rac1 signaling provides an important insight into the regulation of small GTPases.
Sujet(s)
Épines dendritiques/physiologie , Protéines d'activation de la GTPase/physiologie , Facteurs d'échange de nucléotides guanyliques/métabolisme , Protéines proto-oncogènes c-bcr/physiologie , Famille des récepteurs Eph/métabolisme , Synapses/physiologie , Protéine G rac1/métabolisme , Animaux , Technique de Western , Électrophysiologie , Endocytose , Facteurs d'échange de nucléotides guanyliques/antagonistes et inhibiteurs , Facteurs d'échange de nucléotides guanyliques/génétique , Techniques immunoenzymatiques , Immunoprécipitation , Souris , Souris knockout , Neurites/métabolisme , Petit ARN interférent/génétique , Transduction du signal , Protéine-1 de lymphome-T induisant l'invasion et les metastasesRÉSUMÉ
We have reported Metadherin (MTDH) was proven to be overexpression and involved in malignance of chronic lymphocytic leukemia (CLL) via Wnt signaling pathway. In this study, we further investigate the role of MTDH in regulation of BCR signaling pathway in CLL. Six CLL samples whose cells were proliferation after BCR activation were chosen from patients with unmutated IgVH. CCK-8 method used to evaluate the proliferation rate. MTDH expression was measured by quantitative PCR and Western blot. After BCR activation, there exist upregulation of MTDH expression in mRNA and protein level in all six CLL patients (P<0.05). In cell line MEC-1, we observed the same pro-proliferation effect accompanying with elevated MTDH expression. The proliferation effects of BCR activation to MEC-1 can be inhibited by MTDH interference. The results of this study indicate that MTDH involved in the pro-proliferation effect of BCR activation in CLL. And the results imply that MTDH can be a potential therapy target of CLL.
Sujet(s)
Molécules d'adhérence cellulaire/physiologie , Leucémie chronique lymphocytaire à cellules B/physiopathologie , Protéines proto-oncogènes c-bcr/physiologie , Transduction du signal/physiologie , Marqueurs biologiques tumoraux/physiologie , Molécules d'adhérence cellulaire/génétique , Lignée cellulaire tumorale , Prolifération cellulaire/physiologie , Régulation de l'expression des gènes tumoraux/physiologie , Humains , Protéines membranaires , Protéines de liaison à l'ARN , Régulation positive/physiologieRÉSUMÉ
Chronic lymphocytic leukemia (CLL) represents 30% of adult leukemia. TCL1 is expressed in ~ 90% of human CLL. Transgenic expression of TCL1 in murine B cells (Eµ-TCL1) results in mouse CLL. Here we show for the first time that the previously unexplored endoplasmic reticulum (ER) stress response is aberrantly activated in Eµ-TCL1 mouse and human CLL. This includes activation of the IRE-1/XBP-1 pathway and the transcriptionally up-regulated expression of Derlin-1, Derlin-2, BiP, GRP94, and PDI. TCL1 associates with the XBP-1 transcription factor, and causes the dysregulated expression of the transcription factors, Pax5, IRF4, and Blimp-1, and of the activation-induced cytidine deaminase. In addition, TCL1-overexpressing CLL cells manufacture a distinctly different BCR, as we detected increased expression of membrane-bound IgM and altered N-linked glycosylation of Igα and Igß, which account for the hyperactive BCR in malignant CLL. To demonstrate that the ER stress-response pathway is a novel molecular target for the treatment of CLL, we blocked the IRE-1/XBP-1 pathway using a novel inhibitor, and observed apoptosis and significantly stalled growth of CLL cells in vitro and in mice. These studies reveal an important role of TCL1 in activating the ER stress response in support for malignant progression of CLL.
Sujet(s)
Stress du réticulum endoplasmique/génétique , Leucémie myéloïde chronique BCR-ABL positive/génétique , Protéines proto-oncogènes/physiologie , Animaux , Lymphocytes B/métabolisme , Lymphocytes B/anatomopathologie , Cellules cultivées , Modèles animaux de maladie humaine , Évolution de la maladie , Régulation de l'expression des gènes dans la leucémie , Humains , Leucémie myéloïde chronique BCR-ABL positive/anatomopathologie , Souris , Souris transgéniques , Protéines proto-oncogènes/génétique , Protéines proto-oncogènes c-bcr/génétique , Protéines proto-oncogènes c-bcr/métabolisme , Protéines proto-oncogènes c-bcr/physiologie , Facteurs temps , Régulation positive/génétique , Régulation positive/physiologieRÉSUMÉ
The spleen tyrosine kinase family members Syk and Zap-70 are pivotal signal transducers downstream of antigen receptors and exhibit overlapping expression patterns at early lymphocytic developmental stages. To assess their differential kinase fitness in vivo, we generated mice, which carry a Zap-70 cDNA knock-in controlled by intrinsic Syk promoter elements that disrupts wild-type Syk expression. Kinase replacement severely compromised Erk1/2-mediated survival and proper selection of developing B cells at central and peripheral checkpoints, demonstrating critical dependence on BCR signalling quality. Furthermore, ITAM- and hemITAM-mediated activation of platelets and neutrophils was completely blunted, while surprisingly FcγR-mediated phagocytosis in macrophages was retained. The alteration in BCR signalling quality resulted in preferential development and survival of marginal zone B cells and prominent autoreactivity, causing the generation of anti-insulin antibodies and age-related glomerulonephritis. Development of concomitant fasting glucose intolerance in knock-in mice highlights aberrant B cell selection as a potential risk factor for type 1 diabetes, and suggests altered BCR signalling as a mechanism to cause biased cellular and Ig repertoire selection, ultimately contributing to B cell-mediated autoimmune predisposition.
Sujet(s)
Maladies auto-immunes/génétique , État prédiabétique/génétique , Protéines proto-oncogènes c-bcr/physiologie , Animaux , Lymphocytes B/immunologie , Lymphocytes B/métabolisme , Lymphocytes B/physiologie , Cellules cultivées , Techniques de knock-in de gènes , Réarrangement des gènes des lymphocytes B/génétique , Prédisposition génétique à une maladie , Humains , Protéines et peptides de signalisation intracellulaire/génétique , Souris , Souris de lignée C57BL , Souris transgéniques , Protein-tyrosine kinases/génétique , Protéines proto-oncogènes c-bcr/génétique , Protéines proto-oncogènes c-bcr/métabolisme , Transduction du signal/génétique , Syk kinase , ZAP-70 Protein-tyrosine kinase/génétiqueSujet(s)
Lymphocytes B/métabolisme , Acide N-acétyl-neuraminique/métabolisme , Acétylation , Animaux , Humains , Ligands , Souris , Acide N-acétyl-neuraminique/physiologie , Liaison aux protéines , Protéines proto-oncogènes c-bcr/physiologie , Lectine-2 de type Ig liant l'acide sialique/physiologie , Transduction du signal/physiologieRÉSUMÉ
Generation of mature B lymphocytes from early (T1) and late transitional (T2) precursors requires cooperative signaling through BCR and B cell-activating factor receptor 3 (BR3). Recent studies have shown that BCR signaling positively regulates NF-kappaB2, suggesting BCR regulation of BR3 signaling. To investigate the significance of signal integration from BCR and BR3 in B cell development and function, we crossed Btk-deficient mice (btk(-/-)), which are developmentally blocked between the T2 and the mature follicular B cell stage as a result of a partial defect in BCR signaling, and A/WySnJ mice, which possess a mutant BR3 defective in propagating intracellular signals that results in a severely reduced peripheral B cell compartment, although all B cell subsets are present in relatively normal ratios. A/WySnJ x btk(-/-) mice display a B cell-autonomous defect, resulting in a developmental block at an earlier stage (T1) than either mutation alone, leading to the loss of mature splenic follicular and marginal zone B cells, as well as the loss of peritoneal B1 and B2 cell populations. The competence of the double mutant T1 B cells to respond to TLR4 and CD40 survival and activation signals is further attenuated compared with single mutations as evidenced by severely reduced humoral immune responses in vivo and proliferation in response to anti-IgM, LPS, and anti-CD40 stimulation in vitro. Thus, BCR and BR3 independently and in concert regulate the survival, differentiation, and function of all B cell populations at and beyond T1, earliest transitional stage.
Sujet(s)
Sous-populations de lymphocytes B/immunologie , Sous-populations de lymphocytes B/anatomopathologie , Lymphopénie/immunologie , Lymphopénie/anatomopathologie , Récepteurs pour l'antigène des lymphocytes B/déficit , Transduction du signal/immunologie , Agammaglobulinaemia tyrosine kinase , Animaux , Récepteur du BAFF/déficit , Récepteur du BAFF/physiologie , Sous-populations de lymphocytes B/métabolisme , Différenciation cellulaire/génétique , Différenciation cellulaire/immunologie , Survie cellulaire/génétique , Survie cellulaire/immunologie , Cellules cultivées , Lymphopénie/génétique , Souris , Souris de lignée A , Souris de lignée C57BL , Souris knockout , Protein-tyrosine kinases/déficit , Protein-tyrosine kinases/génétique , Protéines proto-oncogènes c-bcr/physiologie , Récepteurs pour l'antigène des lymphocytes B/physiologie , Transduction du signal/génétiqueRÉSUMÉ
PURPOSE: Few biological prognosticators are useful for prediction of Richter syndrome (RS), representing the transformation of chronic lymphocytic leukemia (CLL) to aggressive lymphoma. Stereotyped B-cell receptors (BCR) may have prognostic effect in CLL progression. We tested the prognostic effect of stereotyped BCR for predicting RS transformation. EXPERIMENTAL DESIGN: The prevalence of stereotyped BCR was compared in RS (n = 69) versus nontransformed CLL (n = 714) by a case-control analysis. Subsequently, the effect of stereotyped BCR at CLL diagnosis on risk of RS transformation was actuarially assessed in a consecutive CLL series (n = 753). RESULTS: RS (n = 69) displayed a higher prevalence of stereotyped BCR (P < 0.001) compared with nontransformed CLL. The actuarial risk of RS transformation was significantly higher in CLL carrying stereotyped BCR (P < 0.001). Among BCR subsets most represented in CLL, subset 8 using IGHV4-39/IGHD6-13/IGHJ5 carried the highest risk of RS transformation [hazard ratio (HR), 24.50; P < 0.001]. Multivariate analysis selected stereotyped BCR (HR, 3.33; P = 0.001) and IGHV4-39 usage (HR, 4.03; P = 0.004) as independent predictors of RS transformation. The combination of IGHV4-39 usage and stereotyped BCR in the same patient identified CLL with a very high risk of RS transformation (5-year risk, 68.7%). The risk carried by stereotyped BCR and IGHV4-39 usage was specific for RS transformation and had no effect on CLL progression without transformation. CONCLUSIONS: Analysis of BCR features may help identify CLL patients at risk of RS. A close monitoring and a careful biopsy policy may help early recognition of RS in CLL patients using stereotyped BCR, particularly if combined with IGHV4-39.
Sujet(s)
Leucémie chronique lymphocytaire à cellules B/génétique , Lymphomes/génétique , Protéines proto-oncogènes c-bcr/physiologie , Sujet âgé , Études de cohortes , Évolution de la maladie , Femelle , Fréquence d'allèle , Prédisposition génétique à une maladie , Humains , Leucémie chronique lymphocytaire à cellules B/complications , Leucémie chronique lymphocytaire à cellules B/anatomopathologie , Lymphomes/anatomopathologie , Mâle , Adulte d'âge moyen , Invasion tumorale , Polymorphisme génétique/physiologie , Protéines proto-oncogènes c-bcr/génétique , Facteurs de risque , SyndromeRÉSUMÉ
The BCL6 transcriptional repressor is required for development of germinal center (GC) B cells and when expressed constitutively causes diffuse large B-cell lymphomas (DLBCLs). We examined genome-wide BCL6 promoter binding in GC B cells versus DLBCLs to better understand its function in these settings. BCL6 bound to both distinct and common sets of functionally related gene in normal GC cells versus DLBCL cells. Certain BCL6 target genes were preferentially repressed in GC B cells, but not DLBCL cells. Several such genes have prominent oncogenic functions, such as BCL2, MYC, BMI1, EIF4E, JUNB, and CCND1. BCL6 and BCL2 expression was negatively correlated in primary DLBCLs except in the presence of BCL2 translocations. The specific BCL6 inhibitor retro-inverso BCL6 peptidomimetic inhibitor-induced expression of BCL2 and other oncogenes, consistent with direct repression effects by BCL6. These data are consistent with a model whereby BCL6 can directly silence oncogenes in GC B cells and counterbalance its own tumorigenic potential. Finally, a BCL6 consensus sequence and binding sites for other physiologically relevant transcription factors were highly enriched among target genes and distributed in a pathway-dependent manner, suggesting that BCL6 forms specific regulatory circuits with other B-cell transcriptional factors.
Sujet(s)
Lymphocytes B/métabolisme , Lymphome B diffus à grandes cellules/génétique , Oncogènes/génétique , Protéines proto-oncogènes c-bcl-6/physiologie , Sites de fixation , Techniques de culture cellulaire , Cellules cultivées , Régulation négative , Analyse de profil d'expression de gènes , Régulation de l'expression des gènes tumoraux , Réseaux de régulation génique , Humains , Lymphome B diffus à grandes cellules/métabolisme , Séquençage par oligonucléotides en batterie , Régions promotrices (génétique) , Liaison aux protéines , Protéines proto-oncogènes c-bcl-6/métabolisme , Protéines proto-oncogènes c-bcr/physiologie , Transcription génétique/physiologieRÉSUMÉ
Complement receptor proteins CR2 (CD21) and CR1 (CD35) have been identified as components of the murine B cell co-receptor complex. Gene expression profiles between naïve WT, C3-/-, and CD21/35-/- B cells demonstrate enhanced expression of a Ca(2+)-modulating gene, Pcp4, in WT mice compared to the complement-deficient animals. Increased expression of Pcp4 is also coincident with B cell maturation into end stage phenotypes. Prolonged activation of B cells via cross-linking of the BCR (but not CR1/CR2 alone) leads to increased expression of Pcp4 and suppressed Ca(2+) release. In total these data demonstrate that the expression of Pcp4 in naïve resting mature B cells is dependent upon tonic stimulation from the CR1/CR2 proteins via a C3 ligand, and that antigen specific B cell activation can also elevate Pcp4 expression that is coincident with suppression of calcium-dependent responses.
Sujet(s)
Complément C3/physiologie , Régulation de l'expression des gènes , Protéines de tissu nerveux/génétique , Récepteurs au C3d du complément/physiologie , Animaux , Lymphocytes B/métabolisme , Lymphocytes B/physiologie , Différenciation cellulaire/génétique , Cellules cultivées , Complément C3/génétique , Complément C3/métabolisme , Réactifs réticulants/pharmacologie , Femelle , Souris , Souris de lignée BALB C , Souris de lignée C57BL , Souris knockout , Protéines de tissu nerveux/métabolisme , Protéines proto-oncogènes c-bcr/métabolisme , Protéines proto-oncogènes c-bcr/physiologie , Récepteurs au C3b du complément/génétique , Récepteurs au C3b du complément/métabolisme , Récepteurs au C3d du complément/génétique , Récepteurs au C3d du complément/métabolisme , Transduction du signal/génétique , Transduction du signal/physiologie , Rate/métabolismeSujet(s)
Angiotensine-II/physiologie , Muscles lisses vasculaires/effets des médicaments et des substances chimiques , Facteur de transcription NF-kappa B/physiologie , Récepteur PPAR gamma/antagonistes et inhibiteurs , Protein-Serine-Threonine Kinases/physiologie , Protéines proto-oncogènes c-bcr/physiologie , Protéines adaptatrices de la transduction du signal , Angiotensine-II/pharmacologie , Animaux , Protéines de transport/physiologie , Protéines du cycle cellulaire , Facteurs d'initiation eucaryotes , Protéines d'activation de la GTPase , Inositol polyphosphate 5-phosphatases , Protéines et peptides de signalisation intracellulaire , Souris , Muscles lisses vasculaires/enzymologie , Récepteur PPAR gamma/agonistes , Récepteur PPAR gamma/physiologie , Phosphoprotéines/physiologie , Phosphoric monoester hydrolases/physiologie , Phosphorylation , Maturation post-traductionnelle des protéines , Protein-Serine-Threonine Kinases/antagonistes et inhibiteurs , Protéines proto-oncogènes c-bcr/antagonistes et inhibiteurs , RatsRÉSUMÉ
Bcr is a serine/threonine kinase activated by platelet-derived growth factor that is highly expressed in the neointima after vascular injury. Here, we demonstrate that Bcr is an important mediator of angiotensin (Ang) II and platelet-derived growth factor-mediated inflammatory responses in vascular smooth muscle cells (VSMCs). Among transcription factors that might regulate Ang II-mediated inflammatory responses we found that ligand-mediated peroxisome proliferator-activated receptor (PPAR)gamma transcriptional activity was significantly decreased by Ang II. Ang II increased Bcr expression and kinase activity. Overexpression of Bcr significantly inhibited PPARgamma activity. In contrast, knockdown of Bcr using Bcr small interfering RNA and a dominant-negative form of Bcr (DN-Bcr) reversed Ang II-mediated inhibition of PPARgamma activity significantly, suggesting the critical role of Bcr in Ang II-mediated inhibition of PPARgamma activity. Point-mutation and in vitro kinase analyses showed that PPARgamma was phosphorylated by Bcr at serine 82. Overexpression of wild-type Bcr kinase did not inhibit ligand-mediated PPARgamma1 S82A mutant transcriptional activity, indicating that Bcr regulates PPARgamma activity via S82 phosphorylation. DN-Bcr and Bcr small interfering RNA inhibited Ang II-mediated nuclear factor kappaB activation in VSMCs. DN-PPARgamma reversed DN-Bcr-mediated inhibition of nuclear factor kappaB activation, suggesting that PPARgamma is downstream from Bcr. Intimal proliferation in low-flow carotid arteries was decreased in Bcr knockout mice compared with wild-type mice, suggesting the critical role of Bcr kinase in VSMC proliferation in vivo, at least in part, via regulating PPARgamma/nuclear factor kappaB transcriptional activity.