Project description:The B cell antigen receptor (BCR) is a signaling complex that mediates differentiation stage-specific cell fate decisions in B lymphocytes. While several studies have evidenced differences in signal transduction components as being key to contrasting phenotypic outcomes, little is known about the differential BCR-triggered gene transcription. Here we defined transcriptional changes underlying BCR-induced apoptosis and proliferation of immature and mature B cells, respectively. These findings provide the first insights into the early transcriptional events leading to deletion or clonal expansion of B cells upon antigen recognition. Keywords: cell type comparison, development or differentiation design, time course, compound treatment design, microarray experiment record
Project description:The B cell antigen receptor (BCR) is a signaling complex that mediates differentiation stage-specific cell fate decisions in B lymphocytes. While several studies have evidenced differences in signal transduction components as being key to contrasting phenotypic outcomes, little is known about the differential BCR-triggered gene transcription. Here we defined transcriptional changes underlying BCR-induced apoptosis and proliferation of immature and mature B cells, respectively. These findings provide the first insights into the early transcriptional events leading to deletion or clonal expansion of B cells upon antigen recognition. Keywords: cell type comparison, development or differentiation design, time course, compound treatment design, microarray experiment record Total RNA was extracted from immature (IM) and mature (M) B cells both freshly isolated (0h) and stimulated with (BCR) or without (ctrl) anti-μ F(ab’)2 for 2, 8 or 24h. Following RNA extraction, one round of mRNA amplification was applied using the MessageAmpTM Kit (Ambion). All samples, including the amplified Universal Mouse Reference RNA (Stratagene) which served as a reference at each hybridization, were reverse-transcribed and indirectly labeled with Cy3 and Cy5 (Amersham). Quality was controlled by performing 6 or 8 hybridizations of each sample to the cDNA arrays using a dye-swap strategy. 94 arrays were used for hybridization in total.
Project description:BCR-induced gene expression profile in wild-type and B cell-specific TAK1-deficient B cells; to elucidate how TAK1 regulates BCR-mediated proliferative response Keywords: ordered
Project description:IgG cytoplasmic tail interferes with the induction of antigen-response genes Experiment Overall Design: Comparing antigen-induced genes between B cells expressing anti-HEL IgM BCR and IgMG BCR (chimeric receptor where the extracellular spacer, transmembrane, and cytoplasmic domain of IgM are replaced with those of IgG1)
Project description:To investigate the mechanism of telomerase regulation in BCR-ABL positive cells due to its clinical value, we studied the catalytic component of telomerase, TERT. Our results suggest that BCR-ABL plays an important role in regulating hTERT in K562 (BCR-ABL positive human leukemia) cells. When Gleevec inhibited the tyrosine kinase activity of BCR-ABL, phosphorylation of hTERT was downregulated, therefore suggesting a positive correlation between BCR-ABL and hTERT. Gleevec treatment inhibited hTERT at the mRNA level and significantly reduced telomerase activity (TA) in K562 cells, but not in HL60 or Jurkat cells. TRAP assay also revealed that Gleevec treatment significantly reduced TA specifically in K562 cells. Furthermore, translocation of hTERT from nucleoli to nucleoplasm was observed in K562 cells induced by Gleevec. Although Gleevec down-regulated hTERT mRNA level, the protein level of hTERT remained unchanged. Therefore, Gleevec-induced-TA decrease is not due to the alteration in telomerase subunits expression. It could be presumably due to posttranslational modification of hTERT, possibly through multiple signaling pathways. We have found that Gleevec reduced the tyrosine phosphorylation of hTERT by BCR-ABL, which is associated with the nucleoplasm localization of hTERT from nucleoli sequesters. These findings reveal unknown functions and regulations of telomerase by BCR-ABL. Using cRNA microarray, gene expression of Gleevec-treated and non-treated K562 (BCR-ABL positive) cells were compared against Gleevec-treated and non-treated HL60 (BCR-ABL deficient) cells.
Project description:Compared with naïve B cells, the B cell receptor (BCR) signal in germinal center (GC) B cells is attenuated; however, the significance of this signaling attenuation has not been well defined. Here, to investigate the role of attenuation of BCR signaling, we employed a Csk mutant mouse model in which Csk-deficiency in GC B cells resulted in augmentation of net BCR signaling with no apparent effect on antigen presentation. We found that Csk is required for GC maintenance and efficient antibody affinity maturation. Mechanistically, ROS-induced apoptosis was exacerbated concomitantly with mitochondrial dysfunction in Csk-deficient GC B cells. Hence, our data suggest that attenuation of the BCR signal restrains hyper-ROS production, thereby protecting GC B cells from apoptosis and contributing to efficient affinity maturation.
Project description:The development, homeostasis and function of B lymphocytes involve multiple rounds of B cell receptor (BCR)-controlled proliferation and prolonged maintenance. We analyzed the role of transcription factor Zfx, a recently identified regulator of stem cell maintenance, in B cell development and homeostasis. Conditional Zfx deletion in the bone marrow blocked B cell development at the pre-BCR selection checkpoint. Zfx deficiency in peripheral B cells caused impaired generation of the B-1 cell lineage, accelerated B cell turnover, depletion of mature recirculating cells, and delayed T-dependent antibody responses. Zfx-deficient B cells showed normal proximal BCR signaling, but impaired BCR-induced proliferation and survival. This was accompanied by aberrantly enhanced and prolonged integrated stress response, and delayed induction of Cyclin D2 and Bcl-xL proteins. Thus, Zfx restrains the stress response and couples antigen receptor signaling to B cell expansion and maintenance during development and peripheral homeostasis. Keywords: Expression profiling by array
Project description:The role of mitochondrial homeostatic control in the metabolic remodeling associated with antigen receptor stimulation in B cells remains incompletely defined. Here we report that the mitochondrial antiviral signaling (MAVS) adaptor protein is involved in B cell receptor (BCR) initiated cellular proliferation and prolonged survival. MAVS is well known as a mitochondrial tethered signaling adaptor for sensing viral double-stranded RNA and type I interferon inducer. The role of MAVS downstream of BCR stimulation was recognized in absence of interferon, suggesting a new pathway for MAVS activation that is independent of viral infection. Mitochondria of BCR-activated MAVS-deficient B cells exhibited a damaged phenotype including disrupted mitochondrial morphology, excess mitophagy and the temporal progressive blunting of mitochondrial oxidative capacity with mitochondrial hyperpolarization and cell death. Co-stimulation with anti-CD40, in addition to BCR stimulation, corrected the proliferative gap between MAVS-deficient and -sufficient B cells; however, mitochondrial structural defects and functionality were only partially restored. Our data reveal a previously unrecognized role of MAVS in controlling mitochondrial homeostasis in response to BCR initiated B cell proliferation.
Project description:Reduced basal insulin secretion from islets isolated from tafazzin deficient mice is associated with reduced mitochondrial function