Suppression of Viral Gene Expression in Bovine Leukemia Virus-Associated B-Cell Malignancy: Interplay of Epigenetic Modifications Leading to Chromatin with a Repressive Histone Code

ABSTRACT Ovine leukemia/lymphoma resulting from bovine leukemia virus infection of sheep offers a large animal model for studying mechanisms underlying leukemogenesis. Silencing of viral information including Tax, the major contributor to the oncogenic potential of the virus, is critical if not mandatory for tumor progression. In this study, we have identified epigenetic mechanisms that govern the complete suppression of viral expression, using a lymphoma-derived B-cell clone carrying a silent provirus. Silencing was not relieved by injection of the malignant B cells into sheep. However, exogenous expression of Tax or treatment with either the DNA methyltransferase inhibitor 5′azacytidine or the histone deacetylase (HDAC) inhibitor trichostatin A rescued viral expression, as demonstrated by in vivo infectivity trials. Comparing silent and reactivated provirus, we found mechanistic connections between chromatin conformation and tumor-associated transcriptional repression. Silencing is associated with DNA methylation and decreased accessibility of promoter sequences. HDAC1 and the transcriptional corepressor mSin3A are associated with the inactive but not the reactivated promoter. Silencing correlates with a repressed chromatin structure marked by histone H3 and H4 hypoacetylation, a loss of methylation at H3 lysine 4, and an increase of H3 lysine 9 methylation. These observations point to the critical role of epigenetic mechanisms in tumor-specific virus/oncogene silencing, a potential strategy to evade immune response and favor the propagation of the transformed cell.

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Complete suppression of viral gene expression is associated with the onset and progression of lymphoid malignancy: observations in Bovine Leukemia Virus-infected sheep

Retrovirology ◽

10.1186/1742-4690-4-51 ◽

2007 ◽

Vol 4 (1) ◽

pp. 51 ◽

Cited By ~ 16

Author(s):

Makram Merimi ◽

Pavel Klener ◽

Maud Szynal ◽

Yvette Cleuter ◽

Claude Bagnis ◽

...

Keyword(s):

Gene Expression ◽

Bovine Leukemia Virus ◽

Leukemia Virus ◽

Viral Gene Expression ◽

Infected Sheep ◽

Viral Gene ◽

Complete Suppression ◽

Lymphoid Malignancy

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Inhibition of Histone Deacetylases Induces Bovine Leukemia Virus Expression In Vitro and In Vivo

Journal of Virology ◽

10.1128/jvi.76.10.5034-5042.2002 ◽

2002 ◽

Vol 76 (10) ◽

pp. 5034-5042 ◽

Cited By ~ 25

Author(s):

C. Merezak ◽

M. Reichert ◽

C. Van Lint ◽

P. Kerkhofs ◽

D. Portetelle ◽

...

Keyword(s):

Gene Expression ◽

Bovine Leukemia Virus ◽

Histone Deacetylases ◽

Mononuclear Cells ◽

Ex Vivo ◽

Trichostatin A ◽

Leukemia Virus ◽

Viral Expression ◽

Cellular Genes

ABSTRACT Packaging into nucleosomes results in a global transcriptional repression as a consequence of exclusion of sequence-specific factors. This inhibition can be relieved by using inhibitors of histone deacetylases, acetylation being a major characteristic of transcriptionally active chromatin. Paradoxically, the expression of only ∼2% of the total cellular genes is modulated by histone hyperacetylation. To unravel the potential role of this transcriptional control on BLV expression, we tested the effect of two highly specific inhibitors of deacetylases, trichostatin A (TSA) and trapoxin (TPX). Our results demonstrate that treatment with TSA efficiently enhanced long terminal repeat-directed gene expression of integrated reporter constructs in heterologous D17 stable cell lines. To further examine the biological relevance of these observations made in vitro, we analyzed ex vivo-isolated peripheral blood mononuclear cells (PBMCs) from bovine leukemia virus (BLV)-infected sheep. TSA deacetylase inhibitor induced a drastic increase in viral expression at levels comparable to those induced by treatment with phorbol-12-myristate 13-acetate and ionomycin, the most efficient activators of BLV expression known to date. TSA acted directly on BLV-infected B lymphocytes to increase viral expression and does not seem to require T-cell cooperation. Inhibition of deacetylation after treatment with TSA or TPX also significantly increased viral expression in PBMCs from cattle, the natural host for BLV. Together, our results show that BLV gene expression is, like that of a very small fraction of cellular genes, also regulated by deacetylation.

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Insights into Gene Expression Changes Impacting B-Cell Transformation: Cross-Species Microarray Analysis of Bovine Leukemia Virus Tax-Responsive Genes in Ovine B Cells

Journal of Virology ◽

10.1128/jvi.80.4.1922-1938.2006 ◽

2006 ◽

Vol 80 (4) ◽

pp. 1922-1938 ◽

Cited By ~ 28

Author(s):

Pavel Klener ◽

Maud Szynal ◽

Yvette Cleuter ◽

Makram Merimi ◽

Hugues Duvillier ◽

...

Keyword(s):

Gene Expression ◽

B Cells ◽

Animal Models ◽

B Cell ◽

Bovine Leukemia Virus ◽

Cell Transformation ◽

Leukemia Virus ◽

Large Animal ◽

Cell Leukemia ◽

Cell Leukemia Virus Type

ABSTRACT Large-animal models for leukemia have the potential to aid in the understanding of networks that contribute to oncogenesis. Infection of cattle and sheep with bovine leukemia virus (BLV), a complex retrovirus related to human T-cell leukemia virus type 1 (HTLV-1), is associated with the development of B-cell leukemia. Whereas the natural disease in cattle is characterized by a low tumor incidence, experimental infection of sheep leads to overt leukemia in the majority of infected animals, providing a model for studying the pathogenesis associated with BLV and HTLV-1. TaxBLV, the major oncoprotein, initiates a cascade of events leading toward malignancy, although the basis of transformation is not fully understood. We have taken a cross-species ovine-to-human microarray approach to identify TaxBLV-responsive transcriptional changes in two sets of cultured ovine B cells following retroviral vector-mediated delivery of TaxBLV. Using cDNA-spotted microarrays comprising 10,336 human genes/expressed sequence tags, we identified a cohort of differentially expressed genes, including genes related to apoptosis, DNA transcription, and repair; proto-oncogenes; cell cycle regulators; transcription factors; small Rho GTPases/GTPase-binding proteins; and previously reported TaxHTLV-1-responsive genes. Interestingly, genes known to be associated with human neoplasia, especially B-cell malignancies, were extensively represented. Others were novel or unexpected. The results suggest that TaxBLV deregulates a broad network of interrelated pathways rather than a single B-lineage-specific regulatory process. Although cross-species approaches do not permit a comprehensive analysis of gene expression patterns, they can provide initial clues for the functional roles of genes that participate in B-cell transformation and pinpoint molecular targets not identified using other methods in animal models.

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DNA Cytosine Methylation in the Bovine Leukemia Virus Promoter Is Associated with Latency in a Lymphoma-derived B-cell Line

Journal of Biological Chemistry ◽

10.1074/jbc.m110.107607 ◽

2010 ◽

Vol 285 (25) ◽

pp. 19434-19449 ◽

Cited By ~ 15

Author(s):

Valérie Pierard ◽

Allan Guiguen ◽

Laurence Colin ◽

Gaëlle Wijmeersch ◽

Caroline Vanhulle ◽

...

Keyword(s):

B Cell ◽

Cell Line ◽

Bovine Leukemia Virus ◽

Cytosine Methylation ◽

Leukemia Virus ◽

Dna Cytosine Methylation ◽

Virus Promoter

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Proliferative Responses of a Bovine Leukemia Virus-Infected Lymphoblastoid B-Cell Line by Its Culture Supernatant and Cytokines.

Journal of Veterinary Medical Science ◽

10.1292/jvms.54.255 ◽

1992 ◽

Vol 54 (2) ◽

pp. 255-259 ◽

Cited By ~ 2

Author(s):

YANG Mhan-Pyo ◽

Ryo GOITSUKA ◽

Hajime TSUJIMOTO ◽

Atsuhiko HASEGAWA

Keyword(s):

B Cell ◽

Cell Line ◽

Bovine Leukemia Virus ◽

Culture Supernatant ◽

Leukemia Virus

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Overlapping CRE and E Box Motifs in the Enhancer Sequences of the Bovine Leukemia Virus 5′ Long Terminal Repeat Are Critical for Basal and Acetylation-Dependent Transcriptional Activity of the Viral Promoter: Implications for Viral Latency

Journal of Virology ◽

10.1128/jvi.78.24.13848-13864.2004 ◽

2004 ◽

Vol 78 (24) ◽

pp. 13848-13864 ◽

Cited By ~ 19

Author(s):

Claire Calomme ◽

Ann Dekoninck ◽

Séverine Nizet ◽

Emmanuelle Adam ◽

Thi Liên-Anh Nguyên ◽

...

Keyword(s):

Long Terminal Repeat ◽

Bovine Leukemia Virus ◽

Transcriptional Repression ◽

Trichostatin A ◽

Leukemia Virus ◽

Viral Latency ◽

Terminal Repeat ◽

Binding Studies ◽

Viral Promoter ◽

E Box

ABSTRACT Bovine leukemia virus (BLV) infection is characterized by viral latency in a large proportion of cells containing an integrated provirus. In this study, we postulated that mechanisms directing the recruitment of deacetylases to the BLV 5′ long terminal repeat (LTR) could explain the transcriptional repression of viral expression in vivo. Accordingly, we showed that BLV promoter activity was induced by several deacetylase inhibitors (such as trichostatin A [TSA]) in the context of episomal LTR constructs and in the context of an integrated BLV provirus. Moreover, treatment of BLV-infected cells with TSA increased H4 acetylation at the viral promoter, showing a close correlation between the level of histone acetylation and transcriptional activation of the BLV LTR. Among the known cis-regulatory DNA elements located in the 5′ LTR, three E box motifs overlapping cyclic AMP responsive elements (CREs) in U3 were shown to be involved in transcriptional repression of BLV basal gene expression. Importantly, the combined mutations of these three E box motifs markedly reduced the inducibility of the BLV promoter by TSA. E boxes are susceptible to recognition by transcriptional repressors such as Max-Mad-mSin3 complexes that repress transcription by recruiting deacetylases. However, our in vitro binding studies failed to reveal the presence of Mad-Max proteins in the BLV LTR E box-specific complexes. Remarkably, TSA increased the occupancy of the CREs by CREB/ATF. Therefore, we postulated that the E box-specific complexes exerted their negative cooperative effect on BLV transcription by steric hindrance with the activators CREB/ATF and/or their transcriptional coactivators possessing acetyltransferase activities. Our results thus suggest that the overlapping CRE and E box elements in the BLV LTR were selected during evolution as a novel strategy for BLV to allow better silencing of viral transcription and to escape from the host immune response.

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