Molecular Pathogenesis of Non-Hodgkin's Lymphoma: the Role of Bcl-6 (original) (raw)
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Scientific Reports
Multiple genetic aberrations in the regulation of BCL6, including in acetyltransferase genes, occur in clinically aggressive B-cell lymphomas and lead to higher expression levels and activity of this transcriptional repressor. BCL6 is, therefore, an attractive target for therapy in aggressive lymphomas. In this study romidepsin, a potent histone deacetylase inhibitor (HDACi), induced apoptosis and cell cycle arrest in Burkitt and diffuse large B-cell lymphoma cell lines, which are model cells for studying the mechanism of action of BCL6. Romidepsin caused BCL6 acetylation at early timepoints inhibiting its function, while at later timepoints BCL6 expression was reduced and target gene expression increased due to chromatin modification. MYC contributes to poor prognosis in aggressive lymphoma. MYC function is reduced by inhibition of chromatin readers of the bromodomain and extra-terminal repeat (BET) family, which includes BRD4. The novel combination of romidepsin and JQ1, a BRD4 in...
Epigenetic Programing of B-Cell Lymphoma by BCL6 and Its Genetic Deregulation
Frontiers in Cell and Developmental Biology, 2019
B cell lymphoma is a clinically heterogeneous and pathologically diverse group of diseases with a strong epigenetic component. The B cell lymphoma 6 (BCL6) gene encodes a transcription factor that is critical for normal germinal center reaction B cell development by maintaining an epigenetic and transcriptional state that is permissive for cellular proliferation and DNA damage. The activity of BCL6 can be deregulated by a variety of mechanisms and contributes to the development of B-cell lymphoma. Here we review the direct and indirect mechanisms BCL6 dysregulation in B cell lymphoma, including transcriptional and post-translational regulation of BCL6 expression and activity, and the perturbation of BCL6-regulated epigenetic programs by cooperating chromatin modifying gene mutations. We underscore the critical importance of BCL6 and its associated epigenetic programs in the development of B-cell lymphoma, and discuss avenues for the therapeutic targeting of BCL6 in this context.
Inducible knock-out of BCL6 in lymphoma cells results in tumor stasis
Oncotarget, 2020
Diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphomas worldwide and is characterized by a high diversity of genetic and molecular alterations. Chromosomal translocations and mutations leading to deregulated expression of the transcriptional repressor BCL6 occur in a significant fraction of DLBCL patients. An oncogenic role of BCL6 in the initiation of DLBCL has been shown as the constitutive expression of BCL6 in mice recapitulates the pathogenesis of human DLBCL. However, the role of BCL6 in tumor maintenance remains poorly investigated due to the absence of suitable genetic models and limitations of pharmacological inhibitors. Here, we have utilized tetracycline-inducible CRISPR/Cas9 mutagenesis to study the consequences of BCL6 deletion in established DLBCL models in culture and in vivo. We show that BCL6 knockout in SU-DHL-4 cells in vitro results in an anti-proliferative response 4-7 days after Cas9 induction that was characterized by cell cycle (G1) arrest. Conditional BCL6 deletion in established DLBCL tumors in vivo induced a significant tumor growth inhibition with initial tumor stasis followed by slow tumor growth kinetics. Our findings support a role of BCL6 in the maintenance of lymphoma growth and showcase the utility of inducible CRISPR/ Cas9 systems for probing oncogene addiction. Recent comprehensive sequencing studies in a large cohort of DLBCL patients highlight the heterogeneity of alterations including somatic mutations, copy number alterations, and structural variants [2-4]. Among the most frequently rearranged genes are IGH, BCL2, BCL6, and MYC, with 40%, 21%, 19%, and 8% of cases affected, respectively [5-8]. BCL6 is a DNA-binding protein that represses gene transcription in Germinal Center (GC) B-cells through the recruitment of corepressor proteins. In GCs, BCL6 inhibits DNA damage response pathways and thereby prevents cell cycle arrest and apoptosis during class switch recombination and somatic hypermutation required for antibody maturation in B-cells. Subsequent BCL6 downregulation is crucial
Blood, 2003
The BCL6 proto-oncogene encodes a transcriptional repressor whose expression is deregulated by chromosomal translocations in approximately 40% of diffuse large B-cell lymphomas (DLBCLs). The BCL6 regulatory sequences are also targeted by somatic hypermutation in germinal center (GC) B cells and in a fraction of all GC-derived lymphomas. However, the functional consequences of these mutations are unknown. Here we report that a subset of mutations specifically associated with DLBCL causes deregulated BCL6 transcription. These mutations affect 2 adjacent BCL6 binding sites located within the first noncoding exon of the gene, and they prevent BCL6 from binding its own promoter, thereby disrupting its negative autoregulatory circuit. These alterations were found in approximately 16% of DLBCLs devoid of chromosomal translocations involving the BCL6 locus, but they were not found in normal GC B cells. This study establishes a novel mechanism for BCL6 deregulation and reveals a broader invo...
Immunity, 2000
expressed in plasma cells (Falini et al., 2000). These BCL-6-negative GC B cells include cells with plas-BCL-6, a transcriptional repressor frequently translomacytic morphology and may represent cells that are cated in lymphomas, regulates germinal center B cell terminally differentiating as they exit the GC (Falini et differentiation and inflammation. DNA microarray screenal., 2000). Thus, plasmacytic differentiation, both preing identified genes repressed by BCL-6, including and post-GC, occurs only in the absence of BCL-6 exmany lymphocyte activation genes, suggesting that pression. BCL-6 modulates B cell receptor signals. BCL-6 re-Roughly one-sixth of all B cell non-Hodgkin's lymphopression of two chemokine genes, MIP-1␣ and IP-10, mas have translocations of the BCL-6 gene, making may also attenuate inflammatory responses. Blimp-1, BCL-6 one of the most frequently translocated genes another BCL-6 target, is important for plasmacytic difin these cancers (reviewed in Dalla-Favera et al., 1999; ferentiation. Since BCL-6 expression is silenced in Staudt et al., 1999). An attractive hypothesis is that plasma cells, repression of blimp-1 by BCL-6 may con-BCL-6 translocations cause non-Hodgkin's lymphomas trol plasmacytic differentiation. Indeed, inhibition of by coopting BCL-6's regulatory functions during B cell BCL-6 function initiated changes indicative of plasdifferentiation. In keeping with this idea, the BCL-6 macytic differentiation, including decreased exprestranslocations do not disrupt the BCL-6 coding region sion of c-Myc and increased expression of the cell but invariably substitute the BCL-6 promoter with a varicycle inhibitor p27kip1. These data suggest that maligety of other promoters. Thus, BCL-6 translocations likely cause transformation of B cells by deregulating the exnant transformation by BCL-6 involves inhibition of pression of normal BCL-6 protein. differentiation and enhanced proliferation. The aim of the present study was to discover the molecular pathways regulated by BCL-6 in order to un-Introduction derstand how BCL-6 controls immune responses and promotes the formation of lymphomas. Although the The transcriptional repressor BCL-6 has emerged as a consensus DNA binding site of BCL-6 has been defined multifunctional regulator of lymphocyte differentiation (Baron et al., 1995; Seyfert et al., 1996), the genomic and immune responses (reviewed in Dalla-Favera et al., targets of BCL-6 repression remain largely unknown. 1999; Staudt et al., 1999). BCL-6 mutant mice display The BCL-6 consensus binding site resembles the "GAS" two prominent phenotypes: a failure to form germinal motif recognized by the STAT family of transcription centers during a T cell-dependent immune response factors, raising speculation that BCL-6 may repress and a fatal inflammatory disease distinguished by the some cytokine response genes (Dent et al., 1997; Gupta presence of T helper type 2 (TH2) cells (Dent et al., et al., 1999; Harris et al., 1999). To comprehensively 1997; Fukuda et al., 1997; Ye et al., 1997). Although the identify BCL-6 target genes, we used Lymphochip mimolecular mechanisms underlying these phenotypes croarrays (Alizadeh et al., 1999, 2000). BCL-6 was found are largely unknown, clues to the function of BCL-6 to repress a number of genes involved in B cell activation come from its expression pattern. Although BCL-6 and terminal differentiation, inflammation, and cell cycle mRNA can be detected in many tissues (Allman et al., regulation, providing rich insights into the roles BCL-6 1996), its protein expression is limited mainly to lymphoplays in the immune system and in human lymphomas. cytes (Cattoretti et al., 1995; Onizuka et al., 1995; Allman et al., 1996), with the highest level of BCL-6 protein Results expressed in germinal center (GC) B cells. BCL-6 protein expression in B cells is exquisitely regulated following BCL-6 Expression Systems antigen encounter. Nascent pre-GC B cells upregulate To screen for BCL-6 target genes, we designed indepen-BCL-6 protein, migrate to the follicular area, and initiate dent and complementary cellular systems in which BCL-6 GC formation (Fukuda et al., 1997). In contrast, antigenfunction could be positively or negatively modulated. specific B cells that do not upregulate BCL-6 protein Gain-of-function systems were created by introducing expression differentiate in the periarteriolar lymphoid full-length (FL) BCL-6 into cell lines lacking endogenous sheath (PALS) into plasmablasts and provide an initial BCL-6 protein expression (Figure 1A). Loss-of-function systems were created by expressing a dominant-negative form of BCL-6, consisting solely of the BCL-6 zinc * To whom correspondence should be addressed (e-mail: lstaudt@ box-l.nih.gov).
BCL6 suppression of BCL2 via Miz1 and its disruption in diffuse large B cell lymphoma
Proceedings of the National Academy of Sciences, 2009
The BCL6 proto-oncogene encodes a transcriptional repressor that is required for germinal center (GC) formation and whose deregulation by genomic lesions is implicated in the pathogenesis of GC-derived diffuse large B cell lymphoma (DLBCL) and, less frequently, follicular lymphoma (FL). The biological function of BCL6 is only partially understood because no more than a few genes have been functionally characterized as direct targets of BCL6 transrepression activity. Here we report that the anti-apoptotic proto-oncogene BCL2 is a direct target of BCL6 in GC B cells. BCL6 binds to the BCL2 promoter region by interacting with the transcriptional activator Miz1 and suppresses Miz1-induced activation of BCL2 expression. BCL6-mediated suppression of BCL2 is lost in FL and DLBCL, where the 2 proteins are pathologically coexpressed, because of BCL2 chromosomal translocations and other mechanisms, including Miz1 deregulation and somatic mutations in the BCL2 promoter region. These results identify an important function for BCL6 in facilitating apoptosis of GC B cells via suppression of BCL2, and suggest that blocking this pathway is critical for lymphomagenesis.
Bcl-6 protein expression in normal and neoplastic lymphoid tissue
Annals of Oncology, 1997
The human bcl-6 gene, which is rearranged in about 30% of diffuse large B-cell lymphomas (DLCL-B), encodes for a Kruppel-type zinc finger protein of 706 amino acids. In order to investigate the expression of the bcl-6 gene at the protein level, two monoclonal antibodies (PG-B6a and PG-B6p) directed against the human bcl-6 protein were generated by immunizing Balb/c mice with a recombinant protein corresponding to the amino-terminal region (amino acids 3^484) of bcl-6. PG-B6a (a = avian) recognized the most conserved bcl-6 epitope (expressed in many animal species, including avian). PG-B6p (p = paraffin) reacted with an epitope of bcl-6 partially resistant to fixatives and detectable on microwave-heated paraffin sections. At immunocytochemistry, bcl-6 localized in the nucleus with a microgranular or diffuse pattern. Strong nuclear expression of bcl-6 was mainly detected in normal germinal-center B-cells, whereas mantle-and marginal-zone B cells, as well as plasma cells and marrow B-cell precursors, did not express bcl-6. These immunohistological findings strongly suggest that bcl-6 may play a role as a regulator of germinal-center related functions. All MoAbs stained neoplastic cells of follicular lymphomas, DLCL-B, and Burkitt's lymphomas. In DLCL-B, bcl-6 expression was independent of bcl-6 gene rearrangements and did not correlate with expression of other markers or the proliferation index. Among low-grade B-cell lymphomas, immunostaining for bcl-6 proved useful for differentiating proliferation centers in B-CLL (bcl-2+/bcl-6-) from trapped germinal centers in mantle-cell lymphomas (bcl-2-lbcl-6+). Strong nuclear positivity for bcl-6 was consistently detected in tumor (L&H) cells of nodular, lymphocyte-predominant Hodgkin's disease (NLPHD). These results further support the concept that NLPHD is a histogenetically distinct (germinal-center derived) subtype of HD. Notably, the nuclei of reactive CD3+/CD4+ T cells near to and rosetting around L&H cells in NLPHD were also strongly bcl-6+, but lacked CD40 ligand (CD40L) expression. This staining pattern clearly differed from that of classic HD, whose cellular background was made up of CD3+/ CD4+ T cells showing the Z>c/-6-/CD40L+ phenotype. The above immunohistological findings suggest that (a) bcl-6 may play a role in regulating B-cell differentiation step(s) occurring within germinal centers; (b) deregulated bcl-6 expression caused by rearrangements may contribute to B-lymphomagenesis; (c) bcl-6 is possibly involved in the pathogenesis of NLPHD.
Blood, 2009
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 a...