The ansamycin antibiotic, rifamycin SV, inhibits BCL6 transcriptional repression and forms a complex with the BCL6-BTB/POZ domain

BCL6 is a transcriptional repressor that is over-expressed due to chromosomal translocations, or other abnormalities, in ~40% of diffuse large B-cell lymphoma. BCL6 interacts with co-repressor, SMRT, and this is essential for its role in lymphomas. Peptide or small molecule inhibitors, which prevent the association of SMRT with BCL6, inhibit transcriptional repression and cause apoptosis of lymphoma cells in vitro and in vivo. In order to discover compounds, which have the potential to be developed into BCL6 inhibitors, we screened a natural product library. The ansamycin antibiotic, rifamycin SV, inhibited BCL6 transcriptional repression and NMR spectroscopy confirmed a direct interaction between rifamycin SV and BCL6. To further determine the characteristics of compounds binding to BCL6-POZ we analyzed four other members of this family and showed that rifabutin, bound most strongly. An X-ray crystal structure of the rifabutin-BCL6 complex revealed that rifabutin occupies a partly non-polar pocket making interactions with tyrosine58, asparagine21 and arginine24 of the BCL6-POZ domain. Importantly these residues are also important for the interaction of BLC6 with SMRT. This work demonstrates a unique approach to developing a structure activity relationship for a compound that will form the basis of a therapeutically useful BCL6 inhibitor

An Oncogenic Role for Alternative NF-κB Signaling in DLBCL Revealed upon Deregulated BCL6 Expression

SummaryDiffuse large B cell lymphoma (DLBCL) is a complex disease comprising diverse subtypes and genetic profiles. Possibly because of the prevalence of genetic alterations activating canonical NF-κB activity, a role for oncogenic lesions that activate the alternative NF-κB pathway in DLBCL has remained elusive. Here, we show that deletion/mutation of TRAF3, a negative regulator of the alternative NF-κB pathway, occurs in ∼15% of DLBCLs and that it often coexists with BCL6 translocation, which prevents terminal B cell differentiation. Accordingly, in a mouse model constitutive activation of the alternative NF-κB pathway cooperates with BCL6 deregulation in DLBCL development. This work demonstrates a key oncogenic role for the alternative NF-κB pathway in DLBCL development

Глобализация как фактор радикализации трансформации приоритетов социально-экономического развития в посттранзитивних экономиках

Expression of the for khead transcription factor FOXP1 is essential for early B-cell development, whereas down regulation ofFOXP1at the germinal center (GC) stage is required for GC B-cell function. Aberrantly high FOXP1 expression is frequently observed in diffuse large B-cell lymphoma and mucosa-associated lymphoid tissue lymphoma, being associated with poor prognosis. Here, by gene expression analysis upon ectopic over expression of FOXP1 in primary

Epitope Density Influences CD8+ Memory T Cell Differentiation

The generation of long-lived memory T cells is critical for successful vaccination but the factors controlling their differentiation are still poorly defined. We tested the hypothesis that the strength of T cell receptor (TCR) signaling contributed to memory CD8(+) T cell generation.We manipulated the density of antigenic epitope presented by dendritic cells to mouse naïve CD8(+) T cells, without varying TCR affinity. Our results show that a two-fold decrease in antigen dose selectively affects memory CD8(+) T cell generation without influencing T cell expansion and acquisition of effector functions. Moreover, we show that low antigen dose alters the duration of the interaction between T cells and dendritic cells and finely tunes the expression level of the transcription factors Eomes and Bcl6. Furthermore, we demonstrate that priming with higher epitope density results in a 2-fold decrease in the expression of Neuron-derived orphan nuclear receptor 1 (Nor-1) and this correlates with a lower level of conversion of Bcl-2 into a pro-apoptotic molecule and an increased number of memory T cells.Our results show that the amount of antigen encountered by naïve CD8(+) T cells following immunization with dendritic cells does not influence the generation of functional effector CD8(+) T cells but rather the number of CD8(+) memory T cells that persist in the host. Our data support a model where antigenic epitope density sensed by CD8(+) T cells at priming influences memory generation by modulating Bcl6, Eomes and Nor-1 expression

An extended set of PRDM1/BLIMP1 target genes links binding motif type to dynamic repression

The transcriptional repressor B lymphocyte-induced maturation protein-1 (BLIMP1) regulates gene expression and cell fate. The DNA motif bound by BLIMP1 in vitro overlaps with that of interferon regulatory factors (IRFs), which respond to inflammatory/immune signals. At such sites, BLIMP1 and IRFs can antagonistically regulate promoter activity. In vitro motif selection predicts that only a subset of BLIMP1 or IRF sites is subject to antagonistic regulation, but the extent to which antagonism occurs is unknown, since an unbiased assessment of BLIMP1 occupancy in vivo is lacking. To address this, we identified an extended set of promoters occupied by BLIMP1. Motif discovery and enrichment analysis demonstrate that multiple motif variants are required to capture BLIMP1 binding specificity. These are differentially associated with CpG content, leading to the observation that BLIMP1 DNA-binding is methylation sensitive. In occupied promoters, only a subset of BLIMP1 motifs overlap with IRF motifs. Conversely, a distinct subset of IRF motifs is not enriched amongst occupied promoters. Genes linked to occupied promoters containing overlapping BLIMP1/IRF motifs (e.g. AIM2, SP110, BTN3A3) are shown to constitute a dynamic target set which is preferentially activated by BLIMP1 knock-down. These data confirm and extend the competitive model of BLIMP1 and IRF interaction

MicroRNA signatures in B-cell lymphomas

Accurate lymphoma diagnosis, prognosis and therapy still require additional markers. We explore the potential relevance of microRNA (miRNA) expression in a large series that included all major B-cell non-Hodgkin lymphoma (NHL) types. The data generated were also used to identify miRNAs differentially expressed in Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL) samples. A series of 147 NHL samples and 15 controls were hybridized on a human miRNA one-color platform containing probes for 470 human miRNAs. Each lymphoma type was compared against the entire set of NHLs. BL was also directly compared with DLBCL, and 43 preselected miRNAs were analyzed in a new series of routinely processed samples of 28 BLs and 43 DLBCLs using quantitative reverse transcription-polymerase chain reaction. A signature of 128 miRNAs enabled the characterization of lymphoma neoplasms, reflecting the lymphoma type, cell of origin and/or discrete oncogene alterations. Comparative analysis of BL and DLBCL yielded 19 differentially expressed miRNAs, which were confirmed in a second confirmation series of 71 paraffin-embedded samples. The set of differentially expressed miRNAs found here expands the range of potential diagnostic markers for lymphoma diagnosis, especially when differential diagnosis of BL and DLBCL is required

Blimp1 Activation by AP-1 in Human Lung Cancer Cells Promotes a Migratory Phenotype and Is Inhibited by the Lysyl Oxidase Propeptide

B lymphocyte-induced maturation protein 1 (Blimp1) is a master regulator of B cell differentiation, and controls migration of primordial germ cells. Recently we observed aberrant Blimp1 expression in breast cancer cells resulting from an NF-κB RelB to Ras signaling pathway. In order to address the question of whether the unexpected expression of Blimp1 is seen in other epithelial-derived tumors, we selected lung cancers as they are frequently driven by Ras signaling. Blimp1 was detected in all five lung cancer cell lines examined and shown to promote lung cancer cell migration and invasion. Interrogation of microarray datasets demonstrated elevated BLIMP1 RNA expression in lung adenocarcinoma, pancreatic ductal carcinomas, head and neck tumors as well as in glioblastomas. Involvement of Ras and its downstream kinase c-Raf was confirmed using mutant and siRNA strategies. We next addressed the issue of mechanism of Blimp1 activation in lung cancer. Using knockdown and ectopic expression, the role of the Activator Protein (AP)-1 family of transcription factors was demonstrated. Further, chromatin immunoprecipitation assays confirmed binding to identified AP-1 elements in the BLIMP1 promoter of ectopically expressed c-Jun and of endogenous AP-1 subunits following serum stimulation. The propeptide domain of lysyl oxidase (LOX-PP) was identified as a tumor suppressor, with ability to reduce Ras signaling in lung cancer cells. LOX-PP reduced expression of Blimp1 by binding to c-Raf and inhibiting activation of AP-1, thereby attenuating the migratory phenotype of lung cancer cells. Thus, Blimp1 is a mediator of Ras/Raf/AP-1 signaling that promotes cell migration, and is repressed by LOX-PP in lung cancer

Functionally Distinct Subpopulations of CpG-Activated Memory B Cells

During the human B cell (Bc) recall response, rapid cell division results in multiple Bc subpopulations. The TLR-9 agonist CpG oligodeoxynucleotide, combined with cytokines, causes Bc activation and division in vitro and increased CD27 surface expression in a sub-population of Bc. We hypothesized that the proliferating CD27lo subpopulation, which has a lower frequency of antibody-secreting cells (ASC) than CD27hi plasmablasts, provides alternative functions such as cytokine secretion, costimulation, or antigen presentation. We performed genome-wide transcriptional analysis of CpG activated Bc sorted into undivided, proliferating CD27lo and proliferating CD27hi subpopulations. Our data supported an alternative hypothesis, that CD27lo cells are a transient pre-plasmablast population, expressing genes associated with Bc receptor editing. Undivided cells had an active transcriptional program of non-ASC B cell functions, including cytokine secretion and costimulation, suggesting a link between innate and adaptive Bc responses. Transcriptome analysis suggested a gene regulatory network for CD27lo and CD27hi Bc differentiation

Stochastic Modeling of B Lymphocyte Terminal Differentiation and Its Suppression by Dioxin

<p>Abstract</p> <p>Background</p> <p>Upon antigen encounter, naïve B lymphocytes differentiate into antibody-secreting plasma cells. This humoral immune response is suppressed by the environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and other dioxin-like compounds, which belong to the family of aryl hydrocarbon receptor (AhR) agonists.</p> <p>Results</p> <p>To achieve a better understanding of the immunotoxicity of AhR agonists and their associated health risks, we have used computer simulations to study the behavior of the gene regulatory network underlying B cell terminal differentiation. The core of this network consists of two coupled double-negative feedback loops involving transcriptional repressors Bcl-6, Blimp-1, and Pax5. Bifurcation analysis indicates that the feedback network can constitute a bistable system with two mutually exclusive transcriptional profiles corresponding to naïve B cells and plasma cells. Although individual B cells switch to the plasma cell state in an all-or-none fashion when stimulated by the polyclonal activator lipopolysaccharide (LPS), stochastic fluctuations in gene expression make the switching event probabilistic, leading to heterogeneous differentiation response among individual B cells. Moreover, stochastic gene expression renders the dose-response behavior of a population of B cells substantially graded, a result that is consistent with experimental observations. The steepness of the dose response curve for the number of plasma cells formed vs. LPS dose, as evaluated by the apparent Hill coefficient, is found to be inversely correlated to the noise level in Blimp-1 gene expression. Simulations illustrate how, through AhR-mediated repression of the AP-1 protein, TCDD reduces the probability of LPS-stimulated B cell differentiation. Interestingly, stochastic simulations predict that TCDD may destabilize the plasma cell state, possibly leading to a reversal to the B cell phenotype.</p> <p>Conclusion</p> <p>Our results suggest that stochasticity in gene expression, which renders a graded response at the cell population level, may have been exploited by the immune system to launch humoral immune response of a magnitude appropriately tuned to the antigen dose. In addition to suppressing the initiation of the humoral immune response, dioxin-like compounds may also disrupt the maintenance of the acquired immunity.</p