scholarly journals Autostimulation of the Epstein-Barr Virus BRLF1 Promoter Is Mediated through Consensus Sp1 and Sp3 Binding Sites

2001 ◽  
Vol 75 (11) ◽  
pp. 5240-5251 ◽  
Author(s):  
Tobias Ragoczy ◽  
George Miller
Keyword(s):  
B Cells ◽  
Transcriptional Activation ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Transcriptional Start Site ◽  
Lytic Cycle ◽  
Site Directed Mutagenesis ◽  
Cellular Transcription Factor ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT As an essential step in the lytic cascade, the Rta homologues of gammaherpesviruses all activate their own expression. Consistent with this biologic function, the Epstein-Barr virus (EBV) Rta protein powerfully stimulates the promoter of its own gene, Rp, in EBV-positive B cells in transient-transfection reporter-based assays. We analyzed the activity of RpCAT in response to Rta by deletional and site-directed mutagenesis. Two cognate Sp1 binding sites located at −279 and −45 relative to the transcriptional start site proved crucial for Rta-mediated activation. Previously described binding sites for the cellular transcription factor Zif268 and the viral transactivator ZEBRA were found to be dispensable for activation of RpCAT by Rta. Gel shift analysis, using extracts of B cells in latency or induced into the lytic cycle, identified Sp1 and Sp3 as the predominant cellular proteins bound to Rp near −45. During the lytic cycle, ZEBRA bound Rp near the Sp1/Sp3 site. The binding of Sp1 and Sp3 to Rp correlated with the reporter activities in the mutagenesis study, establishing a direct link between transcriptional activation of Rp by Rta and DNA binding by Sp1 and/or Sp3. The relative abundance or functional state of the cellular Sp1 and Sp3 transcription factors may be altered in response to stimuli that induce the BRLF1 promoter and thereby contribute to the activation of the viral lytic cycle.

scholarly journals Interaction sites of the Epstein–Barr virus Zta transcription factor with the host genome in epithelial cells

2021 ◽  
Vol 3 (11) ◽  
Author(s):  
Anja Godfrey ◽  
Kay Osborn ◽  
Alison J. Sinclair
Keyword(s):  
Transcription Factor ◽  
B Cells ◽  
Epithelial Cells ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Lytic Cycle ◽  
Barr Virus ◽  
Epstein Barr ◽  
The Impact

Epstein–Barr virus (EBV) is present in a state of latency in infected memory B-cells and EBV-associated lymphoid and epithelial cancers. Cell stimulation or differentiation of infected B-cells and epithelial cells induces reactivation to the lytic replication cycle. In each cell type, the EBV transcription and replication factor Zta (BZLF1, EB1) plays a role in mediating the lytic cycle of EBV. Zta is a transcription factor that interacts directly with Zta response elements (ZREs) within viral and cellular genomes. Here we undertake chromatin-precipitation coupled to DNA-sequencing (ChIP-Seq) of Zta-associated DNA from cancer-derived epithelial cells. The analysis identified over 14 000 Zta-binding sites in the cellular genome. We assessed the impact of lytic cycle reactivation on changes in gene expression for a panel of Zta-associated cellular genes. Finally, we compared the Zta-binding sites identified in this study with those previously identified in B-cells and reveal substantial conservation in genes associated with Zta-binding sites.

scholarly journals The Epstein-Barr virus (EBV) nuclear antigen 1 BamHI F promoter is activated on entry of EBV-transformed B cells into the lytic cycle.

1992 ◽  
Vol 66 (12) ◽  
pp. 7461-7468 ◽  
Author(s):  
A L Lear ◽  
M Rowe ◽  
M G Kurilla ◽  
S Lee ◽  
S Henderson ◽  
...  
Keyword(s):  
B Cells ◽  
Epstein Barr Virus ◽  
Lytic Cycle ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Epstein Barr

scholarly journals Histone Loaders CAF1 and HIRA Restrict Epstein-Barr Virus B-Cell Lytic Reactivation

mBio ◽  
2020 ◽  
Vol 11 (5) ◽  
Author(s):  
Yuchen Zhang ◽  
Chang Jiang ◽  
Stephen J. Trudeau ◽  
Yohei Narita ◽  
Bo Zhao ◽  
...  
Keyword(s):  
B Cells ◽  
Burkitt Lymphoma ◽  
Epstein Barr Virus ◽  
Chromatin Assembly ◽  
Lytic Cycle ◽  
Histone Chaperones ◽  
Viral Genomes ◽  
Barr Virus ◽  
Lytic Reactivation ◽  
Epstein Barr

ABSTRACT Epstein-Barr virus (EBV) infects 95% of adults worldwide and causes infectious mononucleosis. EBV is associated with endemic Burkitt lymphoma, Hodgkin lymphoma, posttransplant lymphomas, nasopharyngeal and gastric carcinomas. In these cancers and in most infected B-cells, EBV maintains a state of latency, where nearly 80 lytic cycle antigens are epigenetically suppressed. To gain insights into host epigenetic factors necessary for EBV latency, we recently performed a human genome-wide CRISPR screen that identified the chromatin assembly factor CAF1 as a putative Burkitt latency maintenance factor. CAF1 loads histones H3 and H4 onto newly synthesized host DNA, though its roles in EBV genome chromatin assembly are uncharacterized. Here, we found that CAF1 depletion triggered lytic reactivation and virion secretion from Burkitt cells, despite also strongly inducing interferon-stimulated genes. CAF1 perturbation diminished occupancy of histones 3.1 and 3.3 and of repressive histone 3 lysine 9 and 27 trimethyl (H3K9me3 and H3K27me3) marks at multiple viral genome lytic cycle regulatory elements. Suggestive of an early role in establishment of latency, EBV strongly upregulated CAF1 expression in newly infected primary human B-cells prior to the first mitosis, and histone 3.1 and 3.3 were loaded on the EBV genome by this time point. Knockout of CAF1 subunit CHAF1B impaired establishment of latency in newly EBV-infected Burkitt cells. A nonredundant latency maintenance role was also identified for the DNA synthesis-independent histone 3.3 loader histone regulatory homologue A (HIRA). Since EBV latency also requires histone chaperones alpha thalassemia/mental retardation syndrome X-linked chromatin remodeler (ATRX) and death domain-associated protein (DAXX), EBV coopts multiple host histone pathways to maintain latency, and these are potential targets for lytic induction therapeutic approaches. IMPORTANCE Epstein-Barr virus (EBV) was discovered as the first human tumor virus in endemic Burkitt lymphoma, the most common childhood cancer in sub-Saharan Africa. In Burkitt lymphoma and in 200,000 EBV-associated cancers per year, epigenetic mechanisms maintain viral latency, during which lytic cycle factors are silenced. This property complicated EBV’s discovery and facilitates tumor immunoevasion. DNA methylation and chromatin-based mechanisms contribute to lytic gene silencing. Here, we identified histone chaperones CAF1 and HIRA, which have key roles in host DNA replication-dependent and replication-independent pathways, respectively, as important for EBV latency. EBV strongly upregulates CAF1 in newly infected B-cells, where viral genomes acquire histone 3.1 and 3.3 variants prior to the first mitosis. Since histone chaperones ATRX and DAXX also function in maintenance of EBV latency, our results suggest that EBV coopts multiple histone pathways to reprogram viral genomes and highlight targets for lytic induction therapeutic strategies.

scholarly journals Antibodies to gp350/220 Enhance the Ability of Epstein-Barr Virus To Infect Epithelial Cells

2006 ◽  
Vol 80 (19) ◽  
pp. 9628-9633 ◽  
Author(s):  
Susan M. Turk ◽  
Ru Jiang ◽  
Liudmila S. Chesnokova ◽  
Lindsey M. Hutt-Fletcher
Keyword(s):  
B Cells ◽  
Nasopharyngeal Carcinoma ◽  
Epithelial Cells ◽  
Epstein Barr Virus ◽  
Lytic Cycle ◽  
Virus Envelope ◽  
Barr Virus ◽  
High Risk Populations ◽  
Epstein Barr

ABSTRACT Epstein-Barr virus (EBV) is a persistent, orally transmitted herpesvirus that replicates in B cells and epithelial cells and is associated with lymphoid and epithelial malignancies. The virus binds to CD21 on B cells via glycoprotein gp350/220 and infects efficiently. Infection of cultured epithelial cells has not typically been efficient but can occur in the absence of gp350/220 and CD21 and in vivo is thought to be important to the development of nasopharyngeal carcinoma. We report here that antibodies to gp350/220, which inhibit EBV infection of B cells, enhance infection of epithelial cells. The effect is not mediated by Fc receptor binding but is further enhanced by antibody cross-linking, which may patch gp350/220 in the virus envelope. Saliva from EBV-seropositive individuals has similar effects that can be reversed by depletion of antibody. The results are consistent with a model in which gp350/220 interferes with the access of other important players to the epithelial cell surface. The results may have implications for the development of nasopharyngeal carcinoma in high-risk populations in which elevated titers of antibody to EBV lytic cycle proteins are prognostic.

scholarly journals Knockout of Epstein-Barr Virus BPLF1 Retards B-Cell Transformation and Lymphoma Formation in Humanized Mice

mBio ◽  
2015 ◽  
Vol 6 (5) ◽  
Author(s):  
Christopher B. Whitehurst ◽  
Guangming Li ◽  
Stephanie A. Montgomery ◽  
Nathan D. Montgomery ◽  
Lishan Su ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Cell Transformation ◽  
Lytic Cycle ◽  
Humanized Mice ◽  
Barr Virus ◽  
Epstein Barr ◽  
Knockout Virus ◽  
Large B Cell

ABSTRACTBPLF1 of Epstein-Barr virus (EBV) is classified as a late lytic cycle protein but is also found in the viral tegument, suggesting its potential involvement at both initial and late stages of viral infection. BPLF1 possesses both deubiquitinating and deneddylating activity located in its N-terminal domain and is involved in processes that affect viral infectivity, viral DNA replication, DNA repair, and immune evasion. A recently constructed EBV BPLF1-knockout (KO) virus was used in conjunction with a humanized mouse model that can be infected with EBV, enabling the first characterization of BPLF1 functionin vivo. Results demonstrate that the BPLF1-knockout virus is approximately 90% less infectious than wild-type (WT) virus. Transformation of human B cells, a hallmark of EBV infection, was delayed and reduced with BPLF1-knockout virus. Humanized mice infected with EBV BPLF1-knockout virus showed less weight loss and survived longer than mice infected with equivalent infectious units of WT virus. Additionally, splenic tumors formed in 100% of mice infected with WT EBV but in only 25% of mice infected with BPLF1-KO virus. Morphological features of spleens containing tumors were similar to those in EBV-induced posttransplant lymphoproliferative disease (PTLD) and were almost identical to cases seen in human diffuse large B-cell lymphoma. The presence of EBV genomes was detected in all mice that developed tumors. The results implicate BPLF1 in human B-cell transformation and tumor formation in humanized mice.IMPORTANCEEpstein-Barr virus infects approximately 90% of the world's population and is the causative agent of infectious mononucleosis. EBV also causes aggressive lymphomas in individuals with acquired and innate immune disorders and is strongly associated with diffuse large B-cell lymphomas, classical Hodgkin lymphoma, Burkitt lymphoma, and nasopharyngeal carcinoma (NPC). Typically, EBV initially infects epithelial cells in the oropharynx, followed by a lifelong persistent latent infection in B-cells, which may develop into lymphomas in immunocompromised individuals. This work is the first of its kind in evaluating the effects of EBV's BPLF1 in terms of pathogenesis and lymphomagenesis in humanized mice and implicates BPLF1 in B-cell transformation and tumor development. Currently, there is no efficacious treatment for EBV, and therapeutic targeting of BPLF1 may lead to a new path to treatment for immunocompromised individuals or transplant recipients infected with EBV.

scholarly journals Epstein-Barr  Virus Binding to CD21 Activates the Initial Viral Promoter via NF-κB Induction

1997 ◽  
Vol 186 (5) ◽  
pp. 731-737 ◽  
Author(s):  
Naoyuki Sugano ◽  
Weiping Chen ◽  
M. Luisa Roberts ◽  
Neil R. Cooper
Keyword(s):  
B Cells ◽  
Transcriptional Activation ◽  
Intracellular Signaling ◽  
Epstein Barr Virus ◽  
Human Herpesvirus ◽  
Virus Binding ◽  
Intracellular Signaling Pathway ◽  
Barr Virus ◽  
Sequence Of Events ◽  
Epstein Barr

Epstein-Barr virus (EBV), an oncogenic human herpesvirus, binds to and infects normal human B lymphocytes via CD21, the CR2 complement receptor. Studies of the mechanisms that enable EBV to infect nonactivated, noncycling B cells provide compelling evidence for a sequence of events in which EBV binding to CD21 on purified resting human B cells rapidly activates the NF-κB transcription factor, which, in turn, binds to and mediates transcriptional activation of Wp, the initial viral latent gene promoter. Thus, EBV binding to its cellular receptor on resting B cells triggers an NF-κB–dependent intracellular signaling pathway which is required for infection.

scholarly journals Marked Variation in Response of Consensus Binding Elements for the Rta Protein of Epstein-Barr Virus

2005 ◽  
Vol 79 (15) ◽  
pp. 9635-9650 ◽  
Author(s):  
Lee-Wen Chen ◽  
Pey-Jium Chang ◽  
Henri-Jacques Delecluse ◽  
George Miller
Keyword(s):  
Dna Sequences ◽  
Transcriptional Activation ◽  
Mammalian Cells ◽  
Epstein Barr Virus ◽  
Lytic Cycle ◽  
Maximal Response ◽  
Binding Affinities ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT The R transactivator (Rta) protein activates Epstein-Barr virus (EBV) lytic-cycle genes by several distinct mechanisms that include direct binding to viral promoters, synergy with BamHI Z EBV replication activator (ZEBRA), and activation of cellular signaling pathways. In the direct and synergistic mechanisms of action, Rta binds to specific DNA sequences that are present in the promoters of responsive genes. It has been difficult to demonstrate the capacity of Rta expressed in mammalian cells to bind DNA in vitro in order to study the relative affinities of Rta binding elements. We discovered that a short C-terminal region of Rta inhibits the ability of Rta to bind DNA in vitro. C-terminally truncated versions of Rta bind DNA efficiently and thus facilitate a comparison of consensus Rta binding elements (CRBEs) found in promoters of five Rta-responsive genes: BMLF1, BHLF1, BMRF1, BaRF1, and BLRF2. All CRBEs in the promoters of the five genes conform to the proposed recognition sequence GNCCN9GGNG, where N is any nucleotide and N9 represents a sequence of nine nucleotides. Nonetheless, CRBEs varied markedly in their abilities to bind Rta in electrophoretic mobility shift assays. Not all CRBEs bound or responded to Rta. Binding affinities of the CRBEs and the capacity to be activated by Rta in reporter assays were strongly correlated. The CRBEs from the BMLF1 and BHLF1 promoters conferred the greatest response. The response of the BMRF1, BaRF1, and BLRF2 CRBEs was less robust. By creation of chimeras, inversions, and point mutations, differences in binding affinities and transcriptional activation levels could be attributed to N9 sequence variation. The length of N9 was also critical for a maximal response. In Raji and BZLF1-knockout cells, the mRNAs of the five Rta-responsive lytic-cycle genes differed dramatically in kinetics of expression, abundance, and synergistic responses to ZEBRA and Rta. Affinities of Rta response elements for Rta are likely to play an important role in temporal regulation and the level of lytic-cycle EBV gene expression.

scholarly journals Essential role of PKCδ in histone deacetylase inhibitor-induced Epstein–Barr virus reactivation in nasopharyngeal carcinoma cells

2008 ◽  
Vol 89 (4) ◽  
pp. 878-883 ◽  
Author(s):  
Heng-Huan Lee ◽  
Shih-Shin Chang ◽  
Sue-Jane Lin ◽  
Huey-Huey Chua ◽  
Tze-Jiun Tsai ◽  
...  
Keyword(s):  
Transcriptional Activation ◽  
Epstein Barr Virus ◽  
Nuclear Translocation ◽  
Chemotherapeutic Agents ◽  
Lytic Cycle ◽  
Virus Reactivation ◽  
Barr Virus ◽  
Ebv Reactivation ◽  
Deacetylase Inhibitor ◽  
Epstein Barr

Histone deactylase inhibitors (HDACi) are common chemotherapeutic agents that stimulate Epstein–Barr virus (EBV) reactivation; the detailed mechanism remains obscure. In this study, it is demonstrated that PKCδ is required for induction of the EBV lytic cycle by HDACi. Inhibition of PKCδ abrogates HDACi-mediated transcriptional activation of the Zta promoter and downstream lytic gene expression. Nuclear translocation of PKCδ is observed following HDACi stimulation and its overexpression leads to progression of the EBV lytic cycle. Our study suggests that PKCδ is a crucial mediator of EBV reactivation and provides a novel insight to study the regulation of the EBV lytic cycle.

scholarly journals Interleukin-5 is an autocrine growth factor for Epstein-Barr virus- transformed B lymphocytes

Blood ◽  
1992 ◽  
Vol 79 (7) ◽  
pp. 1763-1767 ◽  
Author(s):  
MA Baumann ◽  
CC Paul
Keyword(s):  
B Cells ◽  
B Lymphocytes ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Maximum Effect ◽  
Surface Binding ◽  
Interleukin 5 ◽  
Barr Virus ◽  
Epstein Barr ◽  
Cells Cultured

Because of the recent finding that interleukin-5 (IL-5) is produced by Epstein-Barr virus-transformed B lymphocytes (EBV-B cells), we performed studies to ascertain whether EBV-B cells might use IL-5 by an autocrine mechanism. EBV-B cells known to be IL-5 producers were capable of responding to addition of exogenous IL-5 by dose-related augmented proliferation. The addition of a neutralizing anti-IL-5 antibody reduced these effects and also dose-dependently inhibited proliferation and reduced viability of unsupplemented EBV-B cells, having a maximum effect at about 120 hours. In contrast, no stimulatory effect of IL-5 was noted on Burkitt's lymphoma cell lines, nor were these lines growth-inhibited by anti-IL-5 antibody. With biotinylated IL-5, (b-IL-5) second labeling with streptavidin-FITC, and flow cytometric analysis, binding of IL-5 to EBV-B cells cultured in fresh medium was demonstrated and could be competed for by excess unlabeled IL-5, suggesting the presence of IL-5-specific binding sites. Binding of IL-5 was reduced on cells cultured for longer periods before study but could be restored by extensively washing cells before labeling them with b-IL-5, suggesting that surface binding sites had become occupied by endogenously produced IL-5. These findings support a role for IL-5 in autocrine support of EBV-B cell growth.

scholarly journals B Cell Receptor-Responsive miR-141 Enhances Epstein-Barr Virus Lytic Cycle via FOXO3 Inhibition

mSphere ◽  
2021 ◽  
Vol 6 (2) ◽  
Author(s):  
Yan Chen ◽  
Devin N. Fachko ◽  
Nikita S. Ivanov ◽  
Rebecca L. Skalsky
Keyword(s):  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Cell Receptor ◽  
Lytic Replication ◽  
B Cell Receptor ◽  
Lytic Cycle ◽  
Cross Linking ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT Antigen recognition by the B cell receptor (BCR) is a physiological trigger for reactivation of Epstein-Barr virus (EBV) and can be recapitulated in vitro by cross-linking of surface immunoglobulins. Previously, we identified a subset of EBV microRNAs (miRNAs) that attenuate BCR signal transduction and subsequently dampen lytic reactivation in B cells. The roles of host miRNAs in the EBV lytic cycle are not completely understood. Here, we profiled the small RNAs in reactivated Burkitt lymphoma cells and identified several miRNAs, such as miR-141, that are induced upon BCR cross-linking. Notably, EBV encodes a viral miRNA, miR-BART9, with sequence homology to miR-141. To better understand the functions of these two miRNAs, we examined their molecular targets and experimentally validated multiple candidates commonly regulated by both miRNAs. Targets included B cell transcription factors and known regulators of EBV immediate-early genes, leading us to hypothesize that these miRNAs modulate kinetics of the lytic cascade in B cells. Through functional assays, we identified roles for miR-141 and EBV miR-BART9 and one specific target, FOXO3, in progression of the lytic cycle. Our data support a model whereby EBV exploits BCR-responsive miR-141 and further mimics activity of this miRNA family via a viral miRNA to promote productive lytic replication. IMPORTANCE EBV is a human pathogen associated with several malignancies. A key aspect of lifelong virus persistence is the ability to switch between latent and lytic replication modes. The mechanisms governing latency, reactivation, and progression of the lytic cycle are only partly understood. This study reveals that specific miRNAs can act to support the EBV lytic phase following BCR-mediated reactivation triggers. Furthermore, this study identifies a role for FOXO3, commonly suppressed by both host and viral miRNAs, in modulating progression of the EBV lytic cycle.

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