scholarly journals Cell Cycle Association of the Retinoblastoma Protein Rb and the Histone Demethylase LSD1 with the Epstein-Barr Virus Latency Promoter Cp

2008 ◽  
Vol 82 (7) ◽  
pp. 3428-3437 ◽  
Author(s):  
Charles M. Chau ◽  
Zhong Deng ◽  
Hyojueng Kang ◽  
Paul M. Lieberman
Keyword(s):  
Cell Cycle ◽  
Epstein Barr Virus ◽  
Affinity Purification ◽  
Histone H3 ◽  
Histone Demethylase ◽  
Stable Complex ◽  
Dependent Manner ◽  
Barr Virus ◽  
A Cell ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus C promoter (Cp) regulates the major multicistronic transcript encoding the EBNA-LP, 1, 2, and 3 genes required for B-cell proliferation during latency. The growth-transforming potential of these viral genes suggests that they must be tightly regulated with the host cell cycle and differentiation process. To better understand Cp regulation, we used DNA affinity purification to identify cellular and viral proteins that bind to Cp in latently infected cells. Several previously unknown factors were identified, including the cell cycle regulatory proteins E2F1 and Rb. E2F1 bound to a specific site in Cp located in the core Cp region 3′ of the known EBNA2-responsive RBP-Jk (CSL, CBF1) binding site. The histone H3 K4 demethylase LSD1 (BCC110) was also identified by DNA affinity and was shown to form a stable complex with Rb. Coimmunoprecipitation assays demonstrated that E2F1, Rb, and LSD1 bind to Cp in a cell cycle-dependent manner. Rb and LSD1 binding to Cp increased after the S phase, corresponding to a decrease in histone H3 K4 methylation and Cp transcription. Coimmunoprecipitation and immunofluorescence assays reveal that LSD1 interacts with Rb. Surprisingly, LSD1 did not coimmunoprecipitate with E2F1, suggesting that it associates with Rb independently of E2F1. Depletion of LSD1 by small interfering RNAs inhibited Cp basal transcription levels, and overexpression of LSD1 altered the cell cycle profile in p53-positive (p53+), but not p53-negative (p53−), HCT cells. These findings indicate that Cp is a cell cycle-regulated promoter that is under the control of Rb and the histone demethylase LSD1 in multiple latency types.

scholarly journals Either ZEB1 or ZEB2/SIP1 Can Play a Central Role in Regulating the Epstein-Barr Virus Latent-Lytic Switch in a Cell-Type-Specific Manner

2010 ◽  
Vol 84 (12) ◽  
pp. 6139-6152 ◽  
Author(s):  
Amy L. Ellis ◽  
Zhenxun Wang ◽  
Xianming Yu ◽  
Janet E. Mertz
Keyword(s):  
Cell Lines ◽  
Epstein Barr Virus ◽  
Cellular Protein ◽  
Lytic Replication ◽  
Viral Reactivation ◽  
Dependent Manner ◽  
Cell Type ◽  
Barr Virus ◽  
A Cell ◽  
Epstein Barr

ABSTRACT We previously reported that the cellular protein ZEB1 can repress expression of the Epstein-Barr virus (EBV) BZLF1 gene in transient transfection assays by directly binding its promoter, Zp. We also reported that EBV containing a 2-bp substitution mutation in the ZEB-binding ZV element of Zp spontaneously reactivated out of latency into lytic replication at a higher frequency than did wild-type EBV. Here, using small interfering RNA (siRNA) and short hairpin RNA (shRNA) technologies, we definitively show that ZEB1 is, indeed, a key player in maintaining EBV latency in some epithelial and B-lymphocytic cell lines. However, in other EBV-positive epithelial and B-cell lines, another zinc finger E-box-binding protein, ZEB2/SIP1, is the key player. Both ZEB1 and ZEB2 can bind Zp via the ZV element. In EBV-positive cells containing only ZEB1, knockdown of ZEB1 led to viral reactivation out of latency, with synthesis of EBV immediate-early and early lytic gene products. However, in EBV-positive cells containing both ZEBs, ZEB2, not ZEB1, was the primary ZEB family member bound to Zp. Knockdown of ZEB2, but not ZEB1, led to EBV lytic reactivation. Thus, we conclude that either ZEB1 or ZEB2 can play a central role in the maintenance of EBV latency, doing so in a cell-type-dependent manner.

scholarly journals Expression of EBNA-1 mRNA Is Regulated by Cell Cycle during Epstein-Barr Virus Type I Latency

1999 ◽  
Vol 73 (4) ◽  
pp. 3154-3161 ◽  
Author(s):  
Matthew G. Davenport ◽  
Joseph S. Pagano
Keyword(s):  
Cell Cycle ◽  
Epstein Barr Virus ◽  
S Phase ◽  
Luciferase Reporter ◽  
Infected Cell Line ◽  
Type I ◽  
Mrna Levels ◽  
Dependent Manner ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT Expression of EBNA-1 protein is required for the establishment and maintenance of the Epstein-Barr virus (EBV) genome during latent infection. During type I latency, the BamHI Q promoter (Qp) gives rise to EBNA-1 expression. The dominant regulatory mechanism for Qp appears to be mediated through the Q locus, located immediately downstream of the transcription start site. Binding of EBNA-1 to the Q locus represses Qp constitutive activity, and repression has been reported to be overcome by an E2F family member that binds to the Q locus and displaces EBNA-1 (N. S. Sung, J. Wilson, M. Davenport, N. D. Sista, and J. S. Pagano, Mol. Cell. Biol. 14:7144–7152, 1994). These data suggest that the final outcome of Qp activity is reciprocally controlled by EBNA-1 and E2F. Since E2F activity is cell cycle regulated, Qp activity and EBNA-1 expression are predicted to be regulated in a cell cycle-dependent manner. Proliferation of the type I latently infected cell line, Akata, was synchronized with the use of the G2/M blocking agent nocodazole. From 65 to 75% of cells could be made to peak in S phase without evidence of viral reactivation. Following release from G2/M block, EBNA-1 mRNA levels declined as the synchronized cells entered the G1 phase of the cell cycle. As cells proceeded into S phase, EBNA-1 mRNA levels increased parallel to the peak in cell numbers in S phase. However, EBNA-1 protein levels showed no detectable change during the cell cycle, most likely due to the protein’s long half-life as estimated by inhibition of protein synthesis by cycloheximide. Finally, in Qp luciferase reporter assays, the activity of Qp was shown to be regulated by cell cycle and to be dependent on the E2F sites within the Q locus. These findings demonstrate that transcriptional activity of Qp is cell cycle regulated and indicated that E2F serves as the stimulus for this regulation.

scholarly journals Telomere Repeat Binding Factors TRF1, TRF2, and hRAP1 Modulate Replication of Epstein-Barr Virus OriP

2003 ◽  
Vol 77 (22) ◽  
pp. 11992-12001 ◽  
Author(s):  
Zhong Deng ◽  
Constandache Atanasiu ◽  
John S. Burg ◽  
Dominique Broccoli ◽  
Paul M. Lieberman
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Epstein Barr Virus ◽  
Affinity Purification ◽  
Dominant Negative ◽  
Full Length ◽  
Barr Virus ◽  
Amino Terminal ◽  
Epstein Barr ◽  
Dyad Symmetry

ABSTRACT Epstein-Barr virus OriP confers cell cycle-dependent DNA replication and stable maintenance on plasmids in EBNA1-positive cells. The dyad symmetry region of OriP contains four EBNA1 binding sites that are punctuated by 9-bp repeats referred to as nonamers. Previous work has shown that the nonamers bind to cellular factors associated with human telomeres and contribute to episomal maintenance of OriP. In this work, we show that substitution mutation of all three nonamer sites reduces both DNA replication and plasmid maintenance of OriP-containing plasmids by 2.5- to 5-fold. The nonamers were required for high-affinity binding of TRF1, TRF2, and hRap1 to the dyad symmetry element but were not essential for the binding of EBNA1 as determined by DNA affinity purification from nuclear extracts. Chromatin immunoprecipitation assays indicated that TRF1, TRF2, and hRap1 bound OriP in vivo. Cell cycle studies indicate that TRF2 binding to OriP peaks in G1/S while TRF1 binding peaks in G2/M. OriP replication was inhibited by transfection of full-length TRF1 but not by deletion mutants lacking the myb DNA binding domain. In contrast, OriP replication was not affected by transfection of full-length TRF2 or hRap1 but was potently inhibited by dominant-negative TRF2 or hRap1 amino-terminal truncation mutants. Knockdown experiments with short interfering RNAs (siRNAs) directed against TRF2 and hRap1 severely reduced OriP replication, while TRF1 siRNA had a modest stimulatory effect on OriP replication. These results indicate that TRF2 and hRap1 promote, while TRF1 antagonizes, OriP-dependent DNA replication and suggest that these telomeric factors contribute to the establishment of replication competence at OriP.

scholarly journals Epstein-Barr Virus miR-BART1-3p Regulates the miR-17-92 Cluster by Targeting E2F3

2021 ◽  
Vol 22 (20) ◽  
pp. 10936
Author(s):  
Myung Chan Park ◽  
Hyoji Kim ◽  
Hoyun Choi ◽  
Mee Soo Chang ◽  
Suk Kyeong Lee
Keyword(s):  
Cell Cycle ◽  
Cell Growth ◽  
Gastric Carcinoma ◽  
Epstein Barr Virus ◽  
Carcinoma Cells ◽  
G1 Arrest ◽  
Protein Levels ◽  
Barr Virus ◽  
A Cell ◽  
Epstein Barr

Epstein-Barr virus (EBV) is associated with several tumors and generates BamHI A rightward transcript (BART) microRNAs (miRNAs) from BART transcript introns. These BART miRNAs are expressed at higher levels in EBV-associated epithelial malignancies than in EBV-infected B lymphomas. To test the effects of EBV miRNA on the cell cycle and cell growth, we transfected miR-BART1-3p, a highly expressed EBV-associated miRNA, into gastric carcinoma cells. We found that miR-BART1-3p induced G0/G1 arrest and suppressed cell growth in gastric carcinoma cells. As our microarray analyses showed that E2F3, a cell cycle regulator, was inhibited by EBV infection, we hypothesized that miR-BART1-3p regulates E2F3. Luciferase assays revealed that miR-BART1-3p directly targeted the 3′-UTR of E2F3 mRNA. Both E2F3 mRNA and encoded protein levels were reduced following miR-BART1-3p transfection. In contrast, E2F3 expression in AGS-EBV cells transfected with a miR-BART1-3p inhibitor was enhanced. As E2F3 has been shown to regulate the expression of highly conserved miR-17-92 clusters in vertebrates, we examined whether this expression is affected by miR-BART1-3p, which can downregulate E2F3. The expression of E2F3, miR-17-92a-1 cluster host gene (MIR17HG), and miR-17-92 cluster miRNAs was significantly reduced in EBV-associated gastric carcinoma (EBVaGC) patients compared with EBV-negative gastric carcinoma (EBVnGC) patients. Further, miR-BART1-3p as well as the siRNA specific to E2F3 inhibited the expression of the miR-17-92 cluster, while inhibition of miR-BART1-3p enhanced the expression of the miR-17-92 cluster in cultured GC cells. Our results suggest a possible role of miR-BART1-3p in cell cycle regulation and in regulation of the miR-17-92 cluster through E2F3 suppression.

scholarly journals Inhibition of Epstein-Barr Virus OriP Function by Tankyrase, a Telomere-Associated Poly-ADP Ribose Polymerase That Binds and Modifies EBNA1

2005 ◽  
Vol 79 (8) ◽  
pp. 4640-4650 ◽  
Author(s):  
Zhong Deng ◽  
Constandache Atanasiu ◽  
Kehao Zhao ◽  
Ronen Marmorstein ◽  
Juan I. Sbodio ◽  
...  
Keyword(s):  
Dna Replication ◽  
Epstein Barr Virus ◽  
Affinity Purification ◽  
Dependent Manner ◽  
Telomere Repeat ◽  
Barr Virus ◽  
Amino Terminal ◽  
Epstein Barr

ABSTRACT Tankyrase (TNKS) is a telomere-associated poly-ADP ribose polymerase (PARP) that has been implicated along with several telomere repeat binding factors in the regulation of Epstein-Barr virus origin of plasmid replication (OriP). We now show that TNKS1 can bind to the family of repeats (FR) and dyad symmetry regions of OriP by using a chromatin immunoprecipitation assay and DNA affinity purification. TNKS1 and TNKS2 bound to EBNA1 in coimmunoprecipitation experiments with transfected cell lysates and with purified recombinant proteins in vitro. Two RXXPDG-like TNKS-interacting motifs in the EBNA1 amino-terminal domain mediated binding with the ankyrin repeat domain of TNKS. Mutations of both motifs at EBNA1 G81 and G425 abrogated TNKS binding and enhanced EBNA1-dependent replication of OriP. Small hairpin RNA targeted knock-down of TNKS1 enhanced OriP-dependent DNA replication. Overexpression of TNKS1 or TNKS2 inhibited OriP-dependent DNA replication, while a PARP-inactive form of TNKS2 (M1045V) was compromised for this inhibition. We show that EBNA1 is subject to PAR modification in vivo and to TNKS1-mediated PAR modification in vitro. These results indicate that TNKS proteins can interact directly with the EBNA1 protein, associate with the FR region of OriP in vivo, and inhibit OriP replication in a PARP-dependent manner.

scholarly journals Recurrent Erythema Annulare Centrifugum due to Influenza Type A

2021 ◽  
pp. 134-140
Author(s):  
Luca Ena ◽  
Vittorio Mazzarello ◽  
Marco Ferrari ◽  
Pasquale Ena
Keyword(s):  
Epstein Barr Virus ◽  
Local Treatment ◽  
Type A ◽  
Barr Virus ◽  
Ebv Infection ◽  
Influenza Type ◽  
Erythema Annulare Centrifugum ◽  
A Cell ◽  
Epstein Barr ◽  
Calcipotriol Betamethasone

Erythema annulare centrifugum (EAC) is a rare erythema characterized by erythematous and urticarial papules or annular plaques that enlarges centrifugally. The lesions usually involve the thighs and the legs. Several disorders are occasionally associated with EAC, infections, including mycoses, bacteria, or viruses and drugs have also been regarded as possible causes of this eruption. We present a 42-year-old dark-skinned woman affected by recurrent EAC that appeared secondary to influenza type A (H1N1). Histopathology showed a superficial form of EAC. In our case, a previous cytomegalovirus and Epstein-Barr virus (EBV) infection were identified and no underlying other diseases were found. Clarithromycin with calcipotriol betamethasone treatment was temporarily efficacious. In the last 3 years, the lesions started to appear every 2 weeks and tended to regress with local treatment after a variable period. We believe that the latent cytomegalovirus and the reactivity induced by EBV combined with influenza can determine, in our case, a cell mediate cutaneous immune response, which leads to the peculiar inflammatory disease known as EAC.

scholarly journals HCF1 and OCT2 Cooperate with EBNA1 To Enhance OriP-Dependent Transcription and Episome Maintenance of Latent Epstein-Barr Virus

2016 ◽  
Vol 90 (11) ◽  
pp. 5353-5367 ◽  
Author(s):  
Jayaraju Dheekollu ◽  
Andreas Wiedmer ◽  
Daniel Sentana-Lledo ◽  
Joel Cassel ◽  
Troy Messick ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Epstein Barr Virus ◽  
Histone H3 ◽  
Type I ◽  
Barr Virus ◽  
Cellular Factors ◽  
Epstein Barr ◽  
Simplex Virus

ABSTRACTEpstein-Barr virus (EBV) establishes latent infections as multicopy episomes with complex patterns of viral gene transcription and chromatin structure. The EBV origin of plasmid replication (OriP) has been implicated as a critical control element for viral transcription, as well as viral DNA replication and episome maintenance. Here, we examine cellular factors that bind OriP and regulate histone modification, transcription regulation, and episome maintenance. We found that OriP is enriched for histone H3 lysine 4 (H3K4) methylation in multiple cell types and latency types. Host cell factor 1 (HCF1), a component of the mixed-lineage leukemia (MLL) histone methyltransferase complex, and transcription factor OCT2 (octamer-binding transcription factor 2) bound cooperatively with EBNA1 (Epstein-Barr virus nuclear antigen 1) at OriP. Depletion of OCT2 or HCF1 deregulated latency transcription and histone modifications at OriP, as well as the OriP-regulated latency type-dependent C promoter (Cp) and Q promoter (Qp). HCF1 depletion led to a loss of histone H3K4me3 (trimethylation of histone H3 at lysine 4) and H3 acetylation at Cp in type III latency and Qp in type I latency, as well as an increase in heterochromatic H3K9me3 at these sites. HCF1 depletion resulted in the loss of EBV episomes from Burkitt's lymphoma cells with type I latency and reactivation from lymphoblastoid cells (LCLs) with type III latency. These findings indicate that HCF1 and OCT2 function at OriP to regulate viral transcription, histone modifications, and episome maintenance. As HCF1 is best known for its function in herpes simplex virus 1 (HSV-1) immediate early gene transcription, our findings suggest that EBV latency transcription shares unexpected features with HSV gene regulation.IMPORTANCEEBV latency is associated with several human cancers. Viral latent cycle gene expression is regulated by the epigenetic control of the OriP enhancer region. Here, we show that cellular factors OCT2 and HCF1 bind OriP in association with EBNA1 to maintain elevated histone H3K4me3 and transcriptional enhancer function. HCF1 is known as a transcriptional coactivator of herpes simplex virus (HSV) immediate early (IE) transcription, suggesting that OriP enhancer shares aspects of HSV IE transcription control.

scholarly journals Direct and Indirect Regulation of Cytokine and Cell Cycle Proteins by EBNA-2 during Epstein-Barr Virus Infection

2001 ◽  
Vol 75 (8) ◽  
pp. 3537-3546 ◽  
Author(s):  
Lindsay C. Spender ◽  
Georgina H. Cornish ◽  
Benjamin Rowland ◽  
Bettina Kempkes ◽  
Paul J. Farrell
Keyword(s):  
Cell Cycle ◽  
Epstein Barr Virus ◽  
Transient Transfection ◽  
Cyclin D2 ◽  
Transient Transfection Assay ◽  
Rnase Protection ◽  
Barr Virus ◽  
Tnf Α ◽  
Epstein Barr ◽  
Transfection Assay

ABSTRACT We have studied the pathways of regulation of cytokine and cell cycle control proteins during infection of human B lymphocytes by Epstein-Barr virus (EBV). Among 30 cytokine RNAs analyzed by the RNase protection assay, tumor necrosis factor alpha (TNF-α), granulocyte colony-stimulating factor, lymphotoxin (LT), and LTβ were found to be regulated within 20 h of EBV infection of primary B cells. Similar results were obtained using the estrogen-regulated EBNA-2 cell line EREB2.5, in which RNAs for LT and TNF-α were induced within 6 h of activation of EBNA-2. Expression of Notch also caused an induction of TNF-α RNA. The induction of TNF-α RNA by EBNA-2 was indirect, and constitutive expression of either LMP-1 or c-myc proteins did not substitute for EBNA-2 in induction of TNF-α RNA. Cyclin D2 is also an indirect target of EBNA-2-mediated transactivation. EBNA-2 was found to activate the cyclin D2 promoter in a transient-transfection assay. A mutant of EBNA-2 that does not bind RBP-Jκ retained some activity in this assay, and activation did not depend on the presence of B-cell-specific factors. Deletion analysis of the cyclin D2 promoter revealed that removal of sequences containing E-box c-myc consensus DNA binding sequences did not reduce EBNA-2-mediated activation of the cyclin D2 promoter in the transient-transfection assay. The results indicate that cytokines are an early target of EBNA-2 and that EBNA-2 can regulate cyclin D2 transcription in EBV-infected cells by mechanisms additional to the c-myc pathway.

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