High Myc activity is an independent negative prognostic factor for diffuse large B cell lymphomas
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ABSTRACT: In this study we investigated to which extend aberrant c Myc activity plays a functional role in other aNHL and whether it is independent from MYC translocations. Based on a combined microarray analysis of human germinal centre (GC) B cells transfected with c Myc and 220 aNHLs cases, we developed a ‘c Myc index’. This data set contains expresssion profiles from Germinal center B-cells transfected with control or Myc expression vectors. The corresponding lymphoma gene expression profiles are included in the series with GEO accession GSE4475. Expression profiling of germinal center B-cells extracted from human tonsils with and without ectopic expression of Myc.
Project description:In order to assess the miRNA signature of B cells undergoing the germinal center reaction, we isolated three subsets of B cells from tonsils--naive (N), germinal center (GC), and memory (M) and looked at their miRNA profile. We looked at three subsets of B cells, isolated from tonsils obtained from four different donors. Thus, we profiled a total of 12 samples, and each tonsillar subset--N, GC, and M, had four biological replicates.
Project description:The transcription factor Bcl6 orchestrates the germinal center reaction through its actions in B and T cells, and regulates inflammatory signaling in macrophages. We report that genetic replacement by mutant Bcl6, which cannot bind corepressors to its BTB domain, disrupted  germinal center formation and immunoglobulin affinity maturation, due to a defect in B cell  proliferation and survival. In contrast, BTB loss of function had no effect on T follicular helper cell differentiation and function, nor other T helper subsets. Bcl6 null mice displayed a lethal inflammatory phenotype, whereas BTB mutant mice experienced normal healthy lives with no inflammation. Bcl6 repression of inflammatory responses in macrophages was accordingly independent of the BTB domain repressor function. Bcl6 thus mediates its actions through lineage-specific biochemical functions. ChIP-seq for Bcl6, SMRT and BCOR in germinal center B cells
Project description:Given the tumor suppressing function of miR-15a/16-1 cluster, we studied its role in the germinal center B-cells that give rise to most lymphoid malignancies.
Project description:PI3K signaling and FOXO transcription factors play opposing roles at several B cell developmental stages. We show here abundant nuclear FOXO1 expression in the proliferative compartment of the germinal center (GC), its dark zone (DZ), and PI3K activity, downregulating FOXO1, in the GC light zone (LZ), where cells are selected for further differentiation. However, here FOXO1 is expressed in c-Myc+ cells destined for DZ reentry. Upon FOXO1 ablation by genetic means or induction of PI3K activity GCs become devoid of their DZ, due at least partly to the downregulation of the chemokine receptor CXCR4. While this is known to prevent proper cyclic selection of cells expressing high-affinity antibodies, the initiation of immunoglobulin switching is essentially dependent on FOXO1 activity. All samples were obtained from mouse germinal center (GC) B cells (Cgamma1-cre; YFP, P110*, FOXO1 f/f or PTEN f/f animals). Cells used for microarray analysis were FACS sorted cells.
Project description:Upon immunization with a T cell dependent antigen naive follicular B cells (Fo) are activated and a germinal center reaction is induced. Within the next 2 weeks large germinal centers develop where the process of affinity maturation takes place. To analyze the gene expression profile of resting and activated B cells, follicular B cells (Fo), B cells from early (GC1) and late germinal centers (GC2) were isolated and their gene expression profile compared. Gene expression profiles of Fo versus GC1 and GC2 B cells, respectively. Naïve Fo B cells were isolated from non-immunized BALB/c mice. Germinal center B cells sorted from spleen cell suspensions of BALB/-c mice immunized with the T cell dependent antigen 2-phenyl Oxazolone. GC1 B cells were isolated 7 days after primary immunization. GC2 cells were isolated 15 days after primary immunization. After total RNA extraction, reverse transcription, cDNA extraction, the biotinylated cRNA was transcribed, fragmented, and 15 µg cRNA hybridized in duplicates for each of the three groups to the GeneChip arrays. Group1: Fo, Group2: GC1, Group3: GC2. Lists of differentially regulated genes were created using High Performance Chip Data Analysis (HPCDA) with Bioretis database (http://www.bioretis-analysis.de). Worldwide data sharing is possible via Bioretis, please ask the authors.
Project description:This SuperSeries is composed of the following subset Series: GSE38696: Gene expression in mouse germinal center light zone and dark zone B cells GSE38697: Gene expression in human germinal center light zone and dark zone B cells Keywords: Expression profiling by array Refer to individual Series
Project description:The germinal center (GC) is a microanatomical compartment wherein high-affinity antibody-producing B cells are selectively expanded. B cells proliferate and mutate their antibody genes in the dark zone (DZ) of the GC and are then selected by T cells in the light zone (LZ) on the basis of affinity. Here, we show that T cell help regulates the speed of cell cycle phase transitions and DNA replication of GC B cells. Genome sequencing and single-molecule analyses revealed that T cell help shortens S phase by regulating replication fork progression while preserving the relative order of replication origin activation. Thus, high-affinity GC B cells are selected by a mechanism that involves prolonged dwell time in the DZ where selected cells undergo accelerated cell cycles. To determine whether GC B cells receiving high levels of T cell help show a specific change in gene expression, we compared DZ cells in the G1 phase of the cell cycle from αDEC-OVA and control αDEC-CS treated GCs using a fluorescent ubiquitination-based cell cycle indicator (Fucci-tg). RNA sequencing revealed that T cell-mediated selection produced an increase in gene expression programs associated with the cell cycle, metabolism, including the metabolism of nucleotides, and genes downstream of c-Myc and the E2F transcription factors.
Project description:B cells diversify and affinity-mature their antigen receptor repertoire in germinal centers (GC). GC B cells receive help signals during transient interaction with T cells, yet it remains unknown how these transient T-B interactions sustain the subsequent proliferative program of selected B cells. Here we show that the transcription factor AP4 is required for sustained GC B cell proliferation and subsequent establishment of a diverse and protective antibody repertoire. AP4 is induced by c-MYC during the T-B interactions and maintained by T cell-derived IL-21. B cell-specific deletion of AP4 resulted in reduced GC sizes and somatic hypermutation and a failure to control chronic viral infection. These results indicate that AP4 integrates T cell-mediated selection and sustained expansion of GC B cells for humoral immunity. Splenic B cells activated with anti-IgM and CD40L in vitro and germinal center B cells sorted from C57BL/6 mice eight days after immunization with sheep red blood cells (SRBCs) were obtained and genome-wide occupancy of c-MYC, AP4 and active histone marks was profiled by chromatin immunoprecipitation and high-throughput sequencing. Gene expression in MYCâAP4â LZ, MYC+AP4+ LZ, AP4+ DZ, and APâ DZ GC B cells from SRBC-immunized AP4-mCherry / c-MYC-GFP dual reporter mice was profiled by RNA-seq.
Project description:MYC translocations are the biologic hallmark of Burkitt lymphomas but also occur in other mature B-cell lymphomas. If accompanied by chromosomal breaks targeting the BCL2 and/or BCL6 oncogenes, these MYC translocation-positive (MYC+) lymphomas are called double-hit lymphomas (DHLs); otherwise, the term single-hit lymphoma (SHL) is applied. In order to characterize the biologic features of these MYC+ lymphomas other than Burkitt lymphomas, we explored, after exclusion of molecular Burkitt lymphoma (mBL) as defined by gene expression profiling (GEP), the molecular, pathological and clinical aspects of 80 MYC translocation (MYC+) lymphomas (31 SHL, 26 BCL2+/MYC+, 14 BCL6+/MYC+, 6 BCL2+/BCL6+/MYC+ and 3 MYC+ lymphomas with unknown BCL6 status). Comparison of SHL and DHL revealed no difference in frequency of MYC partner (IG/non-IG), genomic complexity or MYC expression and no differences in GEP. DHL showed a more frequent GCB-like GEP and higher IGH and MYC mutation rates. GEP revealed 130 differentially expressed genes between BCL6+/MYC+ and BCL2+/MYC+ DHL. BCL2+/MYC+ DHL showed a more frequent GCB-like GEP. Analysis of all lymphomas according to MYC partner (IG/non-IG) revealed no substantial differences. In contrast to mBL and lymphomas without MYC break, SHL and DHL patients had similar poor outcome. Our data suggest that after excluding mBL, MYC+ lymphomas could be biologically widely lumped without further need for subclassification. 32 diffuse large B-cell lymphoma samples were hybridized to HG-U133A Affymetrix GeneChips. In addition, this study contains 30 already published samples, which contribute to GSE4475 (Hummel et al. 2006 (PMID 16760442)), as well as 18 already published samples from GSE22470 (Salaverria et al. 2011 (PMID 21487109)). No re-normalisation of the published samples was performed. The complete dataset representing: (1) the 32 diffuse large B-cell lymphoma Samples, (2) the 30 Samples from GSE4475 and (3) the 18 Samples from GSE22470, is linked below as a supplementary file
Project description:Microarrays of gene expression in human germinal center light zone and dark zone B cells sorted according to the expression of cell surface molecules CD83 and CXCR4 We used microarray data to identify genes differentially expressed by B cells in the light and dark zones of germinal centers from human tonsil specimens. Germinal center B cells (CD19+CD38+IgD-) were sorted from pediatric tonsil discard specimens into light zone (CXCR4hi/CD83lo) and dark zone (CXCR4lo/CD83hi) populations, from which RNA was extracted