scholarly journals Reciprocal regulation of the Epstein-Barr virus BamHI-F promoter by EBNA-1 and an E2F transcription factor.

1994 ◽  
Vol 14 (11) ◽  
pp. 7144-7152 ◽  
Author(s):  
N S Sung ◽  
J Wilson ◽  
M Davenport ◽  
N D Sista ◽  
J S Pagano
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Epithelial Cells ◽  
Binding Site ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Barr Virus ◽  
E2f Transcription Factor ◽  
Epstein Barr

The Epstein-Barr virus BamHI-F promoter (Fp) is one of three used to transcribe the EBNA latency proteins, in particular, EBNA-1, the only viral gene product needed for episomal replication. Fp is distinguished by possession of the only EBNA-1 binding sites (the Q locus) in the Epstein-Barr virus genome outside oriP. Activity of Fp is negatively autoregulated by interaction of EBNA-1 at two sites in the Q locus, which is situated downstream of the RNA start site. We demonstrate in transient assays that this EBNA-1-mediated repression of Fp can be overcome by an E2F transcription factor which interacts with the DNA at a site centered between the two EBNA-1 binding sites within the Q locus. An E2F-1 fusion protein protects the sequence 5'-GGATGGCGGGTAATA-3' from DNase I digestion, and a DNA probe containing this sequence binds an E2F-specific protein complex from cell extracts, although this region is only loosely homologous with known consensus binding sites for E2F transcription factors. In mobility shift assays, E2F can displace the binding of EBNA-1 from the Q locus but not from oriP, where the E2F binding site is not present. E2F also activates expression of Fp in epithelial cells. These findings identify a potentially new binding site for members of the E2F family of transcription factors and suggest that such a factor is important for expression of EBNA-1 in lymphoid and epithelial cells by displacing EBNA-1 from the Q locus. In addition, the possibility that Fp activity is under cell cycle control is raised. Since the supply of functional E2F varies during the cell cycle and since in these assays overexpression of E2F can overcome repression of Fp by EBNA-1, control of transcription of EBNA-1 mRNA by cell cycle regulatory factors may help to bring about ordered replication of episomes.

scholarly journals Reciprocal regulation of the Epstein-Barr virus BamHI-F promoter by EBNA-1 and an E2F transcription factor

1994 ◽  
Vol 14 (11) ◽  
pp. 7144-7152
Author(s):  
N S Sung ◽  
J Wilson ◽  
M Davenport ◽  
N D Sista ◽  
J S Pagano
Keyword(s):  
Cell Cycle ◽  
Transcription Factor ◽  
Transcription Factors ◽  
Epithelial Cells ◽  
Binding Site ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Barr Virus ◽  
E2f Transcription Factor ◽  
Epstein Barr

The Epstein-Barr virus BamHI-F promoter (Fp) is one of three used to transcribe the EBNA latency proteins, in particular, EBNA-1, the only viral gene product needed for episomal replication. Fp is distinguished by possession of the only EBNA-1 binding sites (the Q locus) in the Epstein-Barr virus genome outside oriP. Activity of Fp is negatively autoregulated by interaction of EBNA-1 at two sites in the Q locus, which is situated downstream of the RNA start site. We demonstrate in transient assays that this EBNA-1-mediated repression of Fp can be overcome by an E2F transcription factor which interacts with the DNA at a site centered between the two EBNA-1 binding sites within the Q locus. An E2F-1 fusion protein protects the sequence 5'-GGATGGCGGGTAATA-3' from DNase I digestion, and a DNA probe containing this sequence binds an E2F-specific protein complex from cell extracts, although this region is only loosely homologous with known consensus binding sites for E2F transcription factors. In mobility shift assays, E2F can displace the binding of EBNA-1 from the Q locus but not from oriP, where the E2F binding site is not present. E2F also activates expression of Fp in epithelial cells. These findings identify a potentially new binding site for members of the E2F family of transcription factors and suggest that such a factor is important for expression of EBNA-1 in lymphoid and epithelial cells by displacing EBNA-1 from the Q locus. In addition, the possibility that Fp activity is under cell cycle control is raised. Since the supply of functional E2F varies during the cell cycle and since in these assays overexpression of E2F can overcome repression of Fp by EBNA-1, control of transcription of EBNA-1 mRNA by cell cycle regulatory factors may help to bring about ordered replication of episomes.

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 Epstein-Barr Virus Nuclear Antigen 3C Activates the Latent Membrane Protein 1 Promoter in the Presence of Epstein-Barr Virus Nuclear Antigen 2 through Sequences Encompassing an Spi-1/Spi-B Binding Site

2000 ◽  
Vol 74 (11) ◽  
pp. 5151-5160 ◽  
Author(s):  
Bo Zhao ◽  
Clare E. Sample
Keyword(s):  
Dna Binding ◽  
Binding Site ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Latent Membrane Protein ◽  
Nuclear Antigen ◽  
Latent Membrane Protein 1 ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA-3C) protein is a transcriptional regulator of viral and cellular genes that is essential for EBV-mediated immortalization of B lymphocytes in vitro. EBNA-3C can inhibit transcription through an association with the cellular DNA-binding protein Jκ, a function shared by EBNA-3A and EBNA-3B. Here, we report a mechanism by which EBNA-3C can activate transcription from the EBV latent membrane protein 1 (LMP-1) promoter in conjunction with EBNA-2. Jκ DNA-binding sites were not required for this activation, and a mutant EBNA-3C protein unable to bind Jκ activated transcription as efficiently as wild-type EBNA-3C, indicating that EBNA-3C can regulate transcription through a mechanism that is independent of Jκ. Furthermore, activation of the LMP-1 promoter is a unique function of EBNA-3C, not shared by EBNA-3A and EBNA-3B. The DNA element through which EBNA-3C activates the LMP-1 promoter includes a Spi-1/Spi-B binding site, previously characterized as an important EBNA-2 response element. Although this element has considerable homology to mouse immunoglobulin light chain promoter sequences to which the mouse homologue of Spi-1 binds with its dimerization partner IRF4, we demonstrate that the IRF4-like binding sites in the LMP-1 promoter do not play a role in EBNA-3C-mediated activation. Both EBNA-2 and EBNA-3C were required for transcription mediated through a 41-bp region of the LMP-1 promoter encompassing the Spi binding site. However, EBNA-3C had no effect on transcription mediated in conjunction with the EBNA-2 activation domain fused to the GAL4 DNA-binding domain, suggesting that it does not function as an adapter between EBNA-2 and the cellular transcriptional machinery. Like EBNA-2, EBNA-3C bound directly to both Spi-1 and Spi-B in vitro. This interaction was mediated by a region of EBNA-3C encompassing a likely basic leucine zipper (bZIP) domain and the ets domain of Spi-1 or Spi-B, reminiscent of interactions between bZIP and ets domains of other transcription factors that result in their targeting to DNA. There are many examples of regulation of the hematopoietic-specific Spi transcription factors through protein-protein interactions, and a similar regulation by EBNA-3C, in conjunction with EBNA-2, is likely to be an important and unique contribution of EBNA-3C to EBV-mediated immortalization.

scholarly journals Epstein-Barr Virus Infection Promotes Epithelial Cell Growth by Attenuating Differentiation-Dependent Exit from the Cell Cycle

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Mark R. Eichelberg ◽  
Rene Welch ◽  
J. Tod Guidry ◽  
Ahmed Ali ◽  
Makoto Ohashi ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Epstein Barr Virus ◽  
Latent Infection ◽  
Barr Virus ◽  
Ebv Infection ◽  
Epstein Barr ◽  
Oral Epithelial

ABSTRACT Epstein-Barr virus (EBV) is a human herpesvirus that is associated with lymphomas as well as nasopharyngeal and gastric carcinomas. Although carcinomas account for almost 90% of EBV-associated cancers, progress in examining EBV’s role in their pathogenesis has been limited by difficulty in establishing latent infection in nontransformed epithelial cells. Recently, EBV infection of human telomerase reverse transcriptase (hTERT)-immortalized normal oral keratinocytes (NOKs) has emerged as a model that recapitulates aspects of EBV infection in vivo, such as differentiation-associated viral replication. Using uninfected NOKs and NOKs infected with the Akata strain of EBV (NOKs-Akata), we examined changes in gene expression due to EBV infection and differentiation. Latent EBV infection produced very few significant gene expression changes in undifferentiated NOKs but significantly reduced the extent of differentiation-induced gene expression changes. Gene set enrichment analysis revealed that differentiation-induced downregulation of the cell cycle and metabolism pathways was markedly attenuated in NOKs-Akata relative to that in uninfected NOKs. We also observed that pathways induced by differentiation were less upregulated in NOKs-Akata. We observed decreased differentiation markers and increased suprabasal MCM7 expression in NOKs-Akata versus NOKs when both were grown in raft cultures, consistent with our transcriptome sequencing (RNA-seq) results. These effects were also observed in NOKs infected with a replication-defective EBV mutant (AkataΔRZ), implicating mechanisms other than lytic-gene-induced host shutoff. Our results help to define the mechanisms by which EBV infection alters keratinocyte differentiation and provide a basis for understanding the role of EBV in epithelial cancers. IMPORTANCE Latent infection by Epstein-Barr virus (EBV) is an early event in the development of EBV-associated carcinomas. In oral epithelial tissues, EBV establishes a lytic infection of differentiated epithelial cells to facilitate the spread of the virus to new hosts. Because of limitations in existing model systems, the effects of latent EBV infection on undifferentiated and differentiating epithelial cells are poorly understood. Here, we characterize latent infection of an hTERT-immortalized oral epithelial cell line (NOKs). We find that although EBV expresses a latency pattern similar to that seen in EBV-associated carcinomas, infection of undifferentiated NOKs results in differential expression of a small number of host genes. In differentiating NOKs, however, EBV has a more substantial effect, reducing the extent of differentiation and delaying the exit from the cell cycle. This effect may synergize with preexisting cellular abnormalities to prevent exit from the cell cycle, representing a critical step in the development of cancer.

scholarly journals Characterization of the CBF2 Binding Site within the Epstein-Barr Virus Latency C Promoter and Its Role in Modulating EBNA2-Mediated Transactivation

1998 ◽  
Vol 72 (1) ◽  
pp. 693-700 ◽  
Author(s):  
Ezequiel M. Fuentes-Pananá ◽  
Paul D. Ling
Keyword(s):  
Binding Site ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Viral Latency ◽  
Barr Virus ◽  
Core Sequence ◽  
Virus Latency ◽  
Flanking Sequences ◽  
Epstein Barr ◽  
Conserved Core

ABSTRACT The Epstein-Barr virus (EBV) EBNA2 protein is a transcriptional activator that regulates viral and cellular gene expression and is also essential for EBV-driven immortalization of B lymphocytes. The EBNA2-responsive enhancer in the viral latency C promoter (Cp) binds two cellular factors, CBF1 and CBF2. The precise role of the CBF2 protein for Cp enhancer function is presently unclear. CBF2 does not appear to interact with EBNA2 and binds just downstream of CBF1 between positions −339 and −368 in the Cp EBNA2 enhancer. Within this region an 8-bp sequence, CAGTGCGT, can be found, and a similar sequence is also located downstream of CBF1 binding sites in other EBNA2-responsive promoters. Previous studies have indicated that mutations and methylation in this sequence affect EBNA2 responsiveness. To investigate the requirements for CBF2 binding, we synthesized a series of oligonucleotides carrying double transversion mutations spanning both the conserved core sequence and outside flanking sequences. Surprisingly, mutations outside of the conserved core sequence in 4 bases immediately flanking the 5′ end, GGTT, had the most deleterious effect on CBF2 binding. Mutations in the conserved core had a gradient effect, with those near the 5′ end having the most deleterious effects on CBF2 binding. In addition, the affinities of CBF2 for binding to the LMP-1, LMP-2, and CD23 promoters were also measured. These promoters contain the conserved core but lack the 5′ flanking GGTT motif and bound CBF2 weakly or not at all. Using Cp reporter plasmids containing CBF2 mutant binding sites, we were also able to show that at lower doses of EBNA2, Cp transactivation required a functional CBF2 binding site but that higher doses of EBNA2 transactivated CBF2 mutant promoters to 40% of wild-type levels. These assays indicate that CBF2 is important for EBNA2-mediated transactivation of the viral latency Cp. In addition, CBF2 activity was found to be associated with two polypeptides of 27 and 33 kDa.

scholarly journals The Latency-Associated Nuclear Antigen Encoded by Kaposi's Sarcoma-Associated Herpesvirus Activates Two Major Essential Epstein-Barr Virus Latent Promoters

2001 ◽  
Vol 75 (19) ◽  
pp. 9446-9457 ◽  
Author(s):  
Angela K. Groves ◽  
Murray A. Cotter ◽  
Chitra Subramanian ◽  
Erle S. Robertson
Keyword(s):  
Transcription Factors ◽  
Cell Line ◽  
Binding Sites ◽  
Kaposi's Sarcoma ◽  
Epstein Barr Virus ◽  
Kaposi’S Sarcoma ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Epstein Barr ◽  
Kaposi’S Sarcoma Associated Herpesvirus

ABSTRACT The latency-associated nuclear antigen (LANA) encoded by the Kaposi's sarcoma-associated herpesvirus (KSHV) is expressed in the majority of KSHV-infected cells and in cells coinfected with Epstein-Barr virus (EBV). In coinfected body cavity-based lymphomas (BCBLs), EBV latent membrane protein 1 (LMP1), which is essential for B-lymphocyte transformation, is expressed. EBNA2 upregulates the expression of LMP1 and other cellular genes through specific interactions with cellular transcription factors tethering EBNA2 to its responsive promoters. In coinfected BCBL cells, EBNA2 is not detected but LANA, which is constitutively expressed, contains motifs suggestive of potential transcriptional activity. Additionally, recent studies have shown that LANA is capable of activating cellular promoters. Therefore, we investigated whether LANA can affect transcription from two major EBV latent promoters. In this study, we demonstrated that LANA can efficiently transactivate both the LMP1 and C promoters in the human B-cell line BJAB as well as in the human embryonic kidney 293 cell line. Moreover, we demonstrated that specific domains of LANA containing the putative leucine zipper and the glutamic acid-rich region are highly effective in upregulating these viral promoters, while the amino-terminal region (435 amino acids) exhibited little or no transactivation activity in our assays. We also specifically tested truncations of the LMP1 promoter element and showed that the −204 to +40 region had increased levels of activation compared with a larger region, −512 to +40, which contains two recombination signal-binding protein Jκ binding sites. The smaller, −204 to +40 promoter region contains specific binding sites for the Ets family transcription factor PU.1, transcription activating factor/cyclic AMP response element, and Sp1, all of which are known to function as activators of transcription. Our data therefore suggest a potential role for LANA in regulation of the major EBV latent promoters in KSHV- and EBV-coinfected cells. Furthermore, LANA may be able to activate transcription of viral and cellular promoters in the absence of EBNA2, potentially through association with transcription factors bound to their cognate sequences within the −204 to +40 region. This regulation of viral gene expression is critical for persistence of these DNA tumor viruses and most likely involved in mediating the oncogenic process in these coinfected cells.

scholarly journals Induction of Epstein-Barr Virus Oncoprotein LMP1 by Transcription Factors AP-2 and Early B Cell Factor

2016 ◽  
Vol 90 (8) ◽  
pp. 3873-3889 ◽  
Author(s):  
Takayuki Murata ◽  
Chieko Noda ◽  
Yohei Narita ◽  
Takahiro Watanabe ◽  
Masahiro Yoshida ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cell ◽  
Binding Sites ◽  
Epstein Barr Virus ◽  
Cell Transformation ◽  
Latent Membrane Protein 1 ◽  
Barr Virus ◽  
Pou Domain ◽  
Epstein Barr ◽  
Early B Cell Factor

ABSTRACTLatent membrane protein 1 (LMP1) is a major oncogene essential for primary B cell transformation by Epstein-Barr virus (EBV). Previous studies suggested that some transcription factors, such as PU.1, RBP-Jκ, NF-κB, and STAT, are involved in this expression, but the underlying mechanism is unclear. Here, we identified binding sites for PAX5, AP-2, and EBF in the proximal LMP1 promoter (ED-L1p). We first confirmed the significance of PU.1 and POU domain transcription factor binding for activation of the promoter in latency III. We then focused on the transcription factors AP-2 and early B cell factor (EBF). Interestingly, among the three AP-2-binding sites in the LMP1 promoter, two motifs were also bound by EBF. Overexpression, knockdown, and mutagenesis in the context of the viral genome indicated that AP-2 plays an important role in LMP1 expression in latency II in epithelial cells. In latency III B cells, on the other hand, the B cell-specific transcription factor EBF binds to the ED-L1p and activates LMP1 transcription from the promoter.IMPORTANCEEpstein-Barr virus (EBV) latent membrane protein 1 (LMP1) is crucial for B cell transformation and oncogenesis of other EBV-related malignancies, such as nasopharyngeal carcinoma and T/NK lymphoma. Its expression is largely dependent on the cell type or condition, and some transcription factors have been implicated in its regulation. However, these previous reports evaluated the significance of specific factors mostly by reporter assay. In this study, we prepared point-mutated EBV at the binding sites of such transcription factors and confirmed the importance of AP-2, EBF, PU.1, and POU domain factors. Our results will provide insight into the transcriptional regulation of the major oncogene LMP1.

scholarly journals ΔNp63α promotes Epstein-Barr virus latency in undifferentiated epithelial cells

2021 ◽  
Vol 17 (11) ◽  
pp. e1010045
Author(s):  
Nicholas Van Sciver ◽  
Makoto Ohashi ◽  
Dhananjay M. Nawandar ◽  
Nicholas P. Pauly ◽  
Denis Lee ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Nasopharyngeal Carcinoma ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Viral Latency ◽  
Human Herpesvirus ◽  
Barr Virus ◽  
Epstein Barr

Epstein-Barr virus (EBV) is a human herpesvirus that causes infectious mononucleosis and contributes to both B-cell and epithelial-cell malignancies. EBV-infected epithelial cell tumors, including nasopharyngeal carcinoma (NPC), are largely composed of latently infected cells, but the mechanism(s) maintaining viral latency are poorly understood. Expression of the EBV BZLF1 (Z) and BRLF1 (R) encoded immediate-early (IE) proteins induces lytic infection, and these IE proteins activate each other’s promoters. ΔNp63α (a p53 family member) is required for proliferation and survival of basal epithelial cells and is over-expressed in NPC tumors. Here we show that ΔNp63α promotes EBV latency by inhibiting activation of the BZLF1 IE promoter (Zp). Furthermore, we find that another p63 gene splice variant, TAp63α, which is expressed in some Burkitt and diffuse large B cell lymphomas, also represses EBV lytic reactivation. We demonstrate that ΔNp63α inhibits the Zp promoter indirectly by preventing the ability of other transcription factors, including the viral IE R protein and the cellular KLF4 protein, to activate Zp. Mechanistically, we show that ΔNp63α promotes viral latency in undifferentiated epithelial cells both by enhancing expression of a known Zp repressor protein, c-myc, and by decreasing cellular p38 kinase activity. Furthermore, we find that the ability of cis-platinum chemotherapy to degrade ΔNp63α contributes to the lytic-inducing effect of this agent in EBV-infected epithelial cells. Together these findings demonstrate that the loss of ΔNp63α expression, in conjunction with enhanced expression of differentiation-dependent transcription factors such as BLIMP1 and KLF4, induces lytic EBV reactivation during normal epithelial cell differentiation. Conversely, expression of ΔNp63α in undifferentiated nasopharyngeal carcinoma cells and TAp63α in Burkitt lymphoma promotes EBV latency in these malignancies.

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