scholarly journals The mTOR inhibitor manassantin B reveals a crucial role of mTORC2 signaling in Epstein-Barr virus reactivation

2020 ◽  
Vol 295 (21) ◽  
pp. 7431-7441 ◽  
Author(s):  
Qian Wang ◽  
Nannan Zhu ◽  
Jiayuan Hu ◽  
Yan Wang ◽  
Jun Xu ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epstein Barr Virus ◽  
Mammalian Target Of Rapamycin ◽  
Antiviral Drug ◽  
Lytic Replication ◽  
Early Gene ◽  
Virus Reactivation ◽  
Barr Virus ◽  
Saururus Chinensis ◽  
Epstein Barr

Lytic replication of Epstein-Barr virus (EBV) is not only essential for its cell–to–cell spread and host–to–host transmission, but it also contributes to EBV-induced oncogenesis. Thus, blocking EBV lytic replication could be a strategy for managing EBV-associated diseases. Previously, we identified a series of natural lignans isolated from the roots of Saururus chinensis (Asian lizard's tail) that efficiently block EBV lytic replication and virion production with low cytotoxicity. In this study, we attempted to elucidate the molecular mechanism by which these lignans inhibit EBV lytic replication. We found that a representative compound, CSC27 (manassantin B), inhibits EBV lytic replication by suppressing the expression of EBV immediate-early gene BZLF1 via disruption of AP-1 signal transduction. Further analysis revealed that manassantin B specifically blocks the mammalian target of rapamycin complex 2 (mTORC2)-mediated phosphorylation of AKT Ser/Thr protein kinase at Ser-473 and protein kinase Cα (PKCα) at Ser-657. Using phosphoinositide 3-kinase–AKT-specific inhibitors for kinase mapping and shRNA-mediated gene silencing, we validated that manassantin B abrogates EBV lytic replication by inhibiting mTORC2 activity and thereby blocking the mTORC2–PKC/AKT-signaling pathway. These results suggest that mTORC2 may have utility as an antiviral drug target against EBV infections and also reveal that manassantin B has potential therapeutic value for managing cancers that depend on mTORC2 signaling for survival.

scholarly journals The BFRF1 Gene of Epstein-Barr Virus Encodes a Novel Protein

2000 ◽  
Vol 74 (7) ◽  
pp. 3235-3244 ◽  
Author(s):  
Antonella Farina ◽  
Roberta Santarelli ◽  
Roberta Gonnella ◽  
Roberto Bei ◽  
Raffaella Muraro ◽  
...  
Keyword(s):  
Cell Lines ◽  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Lytic Cycle ◽  
Particle Identification ◽  
Early Gene ◽  
Cell Fractionation ◽  
Barr Virus ◽  
F Region ◽  
Epstein Barr

ABSTRACT Computer analysis of the Epstein-Barr virus (EBV) genome indicates there are ∼100 open reading frames (ORFs). Thus far about 30 EBV genes divided into the categories latent and lytic have been identified. The BamHI F region of EBV is abundantly transcribed during lytic replication. This region is highly conserved among herpesviruses, thus suggesting that some common function could be retained in the ORFs encompassed within this viral fragment. To identify putative novel proteins and possible new markers for viral replication, we focused our attention on the first rightward ORF in theBamHI F region (BFRF1). Histidine and glutathione S-transferase-tagged BFRF1 fusion proteins were synthesized to produce a mouse monoclonal antibody (MAb). Analysis of human sera revealed a high seroprevalence of antibodies to BFRF1 in patients affected by nasopharyngeal carcinoma or Burkitt's lymphoma, whereas no humoral response to BFRF1 could be detected among healthy donors. An anti-BFRF1 MAb recognizes a doublet migrating at 37 to 38 kDa in cells extracts from EBV-infected cell lines following lytic cycle activation and in an EBV-negative cell line (DG75) transfected with a plasmid expressing the BFRF1 gene. Northern blot analysis allowed the detection of a major transcript of 3.7 kb highly expressed in EBV-positive lytic cycle-induced cell lines. Treatment with inhibitors of viral DNA polymerase, such as phosphonoacetic acid and acyclovir, reduced but did not abolish the transcription ofBFRF1, thus indicating that BFRF1 can be classified as an early gene. Cell fractionation experiments, as well as immunolocalization by immunofluorescence microscopy, immunohistochemistry, and immunoelectron microscopy, showed that BFRF1 is localized on the plasma membrane and nuclear compartments of the cells and is a structural component of the viral particle. Identification of BFRF1 provides a new marker with which to monitor EBV infection and might help us better understand the biology of the virus.

scholarly journals Effects of Maribavir and Selected Indolocarbazoles on Epstein-Barr Virus Protein Kinase BGLF4 and on Viral Lytic Replication

2004 ◽  
Vol 48 (5) ◽  
pp. 1900-1903 ◽  
Author(s):  
Edward Gershburg ◽  
Ke Hong ◽  
Joseph S. Pagano
Keyword(s):  
Protein Kinase ◽  
Dna Synthesis ◽  
Human Cytomegalovirus ◽  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Virus Protein ◽  
Barr Virus ◽  
Ebv Dna ◽  
Epstein Barr

ABSTRACT The human cytomegalovirus (HCMV) homolog of the Epstein-Barr virus (EBV) protein kinase (PK), UL97, is inhibited by maribavir (1263W94) and selected indolocarbazoles. Here we show that only one of these indolocarbazoles (K252a), but not maribavir, inhibits autophosphorylation of the EBV PK, BGLF4. However, maribavir and another indolocarbazole, NGIC-I, do inhibit EBV DNA synthesis, suggesting that although these last compounds inhibit both HCMV and EBV, they seem to operate through differ-ent pathways.

scholarly journals Phosphorylation of p27Kip1by Epstein-Barr Virus Protein Kinase Induces Its Degradation through SCFSkp2Ubiquitin Ligase Actions during Viral Lytic Replication

2009 ◽  
Vol 284 (28) ◽  
pp. 18923-18931 ◽  
Author(s):  
Satoko Iwahori ◽  
Takayuki Murata ◽  
Ayumi Kudoh ◽  
Yoshitaka Sato ◽  
Sanae Nakayama ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Virus Protein ◽  
Barr Virus ◽  
Epstein Barr

scholarly journals B Cell Receptor-Responsive miR-141 and viral miR-BART9 promote Epstein-Barr Virus Reactivation Through FOXO3 Inhibition

2019 ◽  
Author(s):  
Yan Chen ◽  
Devin Fachko ◽  
Nikita S. Ivanov ◽  
Rebecca L. Skalsky
Keyword(s):  
B Cell ◽  
Epstein Barr Virus ◽  
Cell Receptor ◽  
Lytic Replication ◽  
B Cell Receptor ◽  
Cross Linking ◽  
Virus Reactivation ◽  
Barr Virus ◽  
Lytic Reactivation ◽  
Epstein Barr

AbstractAntigen 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 replication in B cells. The roles of host miRNAs in virus reactivation are not completely understood. To investigate this process, we profiled the small RNAs in latently infected and reactivated Burkitt’s 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 transcriptional regulators of the EBV immediate early promoters and B cell transcription factors, leading us to hypothesize that these miRNAs modulate kinetics of the latent to lytic switch in B cells. Through functional assays, we identified roles for miR-141 and EBV miR-BART9 and one specific target, FOXO3, in lytic reactivation. 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 virus replication.ImportanceEBV 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. Mechanisms governing latency and lytic reactivation are only partly understood, and how the EBV latent to lytic switch is post-transcriptionally regulated remains an outstanding question. This study sheds light on how miR-141 expression is regulated in Burkitt’s lymphoma cells, and identifies a role for FOXO3, a common target of both miR-141 and viral miR-BART9, in modulating EBV reactivation.

scholarly journals Ubiquitin Modification of the Epstein-Barr Virus Immediate Early Transactivator Zta

2020 ◽  
Vol 94 (22) ◽  
Author(s):  
Mengmeng Zhao ◽  
Asuka Nanbo ◽  
David Becnel ◽  
Zhiqiang Qin ◽  
Gilbert F. Morris ◽  
...  
Keyword(s):  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Early Gene ◽  
Immediate Early ◽  
Viral Gene ◽  
Barr Virus ◽  
Amino Terminal ◽  
Lysine Residues ◽  
Infected Cells ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV) immediate early transactivator Zta plays a key role in regulating the transition from latency to the lytic replication stages of EBV infection. Regulation of Zta is known to be controlled through a number of transcriptional and posttranscriptional events. Here, we show that Zta is targeted for ubiquitin modification and that this can occur in EBV-negative and in EBV-infected cells. Genetic studies show critical roles for both an amino-terminal region of Zta and the basic DNA binding domain of Zta in regulating Zta ubiquitination. Pulse-chase experiments demonstrate that the bulk population of Zta is relatively stable but that at least a subset of ubiquitinated Zta molecules are targeted for degradation in the cell. Mutation of four out of a total of nine lysine residues in Zta largely abrogates its ubiquitination, indicating that these are primary ubiquitination target sites. A Zta mutant carrying mutations at these four lysine residues (lysine 12, lysine 188, lysine 207, and lysine 219) cannot induce latently infected cells to produce and/or release infectious virions. Nevertheless, this mutant can induce early gene expression, suggesting a possible defect at the level of viral replication or later in the lytic cascade. As far as we know, this is the first study that has investigated the targeting of Zta by ubiquitination or its role in Zta function. IMPORTANCE Epstein-Barr virus (EBV) is a ubiquitous human pathogen and associated with various human diseases. EBV undergoes latency and lytic replication stages in its life cycle. The transition into the lytic replication stage, at which virus is produced, is mainly regulated by the viral gene product, Zta. Therefore, the regulation of Zta function becomes a central issue regarding viral biology and pathogenesis. Known modifications of Zta include phosphorylation and sumoylation. Here, we report the role of ubiquitination in regulating Zta function. We found that Zta is subjected to ubiquitination in both EBV-infected and EBV-negative cells. The ubiquitin modification targets 4 lysine residues on Zta, leading to both mono- and polyubiquitination of Zta. Ubiquitination of Zta affects the protein’s stability and likely contributes to the progression of viral lytic replication. The function and fate of Zta may be determined by the specific lysine residue being modified.

Potential Antiviral Lignans from the Roots of Saururus chinensis with Activity against Epstein–Barr Virus Lytic Replication

2013 ◽  
Vol 77 (1) ◽  
pp. 100-110 ◽  
Author(s):  
Hui Cui ◽  
Bo Xu ◽  
Taizong Wu ◽  
Jun Xu ◽  
Yan Yuan ◽  
...  
Keyword(s):  
Epstein Barr Virus ◽  
Lytic Replication ◽  
Barr Virus ◽  
Saururus Chinensis ◽  
Epstein Barr

scholarly journals Inhibition of Epstein-Barr Virus (EBV) Reactivation by Short Interfering RNAs Targeting p38 Mitogen-Activated Protein Kinase or c-myc in EBV-Positive Epithelial Cells

2004 ◽  
Vol 78 (21) ◽  
pp. 11798-11806 ◽  
Author(s):  
Xiangrong Gao ◽  
Haoran Wang ◽  
Takeshi Sairenji
Keyword(s):  
Protein Kinase ◽  
Epstein Barr Virus ◽  
Antiviral Effect ◽  
Mitogen Activated Protein Kinase ◽  
Early Gene ◽  
No Donor ◽  
Barr Virus ◽  
Ebv Reactivation ◽  
Mitogen Activated Protein ◽  
Epstein Barr

ABSTRACT Latent Epstein-Barr virus (EBV) is reactivated by 12-O-tetradecanoylphorbol-13-acetate (TPA) in EBV-infected cells. In this study, we found that TPA up-regulated phosphorylation of p38, a mitogen-activated protein kinase, and activated c-myc mRNA in EBV-positive epithelial GT38 cells. The EBV immediate-early gene BZLF1 mRNA and its product ZEBRA protein were induced following TPA treatment. Protein kinase C inhibitors, 1-(5-isoquinolinesulphonyl)-2, 5-dimethylpiperazine (H7) and staurosporine, inhibited the induction of p38 phosphorylation and the activation of c-Myc by TPA. The p38 inhibitor SB203580 blocked both p38 phosphorylation and ZEBRA expression by TPA. Pretreatment of GT38 cells with the nitric oxide (NO) donor S-nitroso-N-acetylpenicillamine inhibited p38 phosphorylation and c-Myc activation by TPA, suggesting that NO may inhibit EBV reactivation via both p38 and c-Myc. By using short interfering RNA (siRNA) targeting either p38 or c-myc, we found that p38 or c-myc siRNA specifically inhibited expression of the respective gene and also suppressed the induction of ZEBRA and EBV early antigen. The interferon (IFN)-responsive gene expression tests ruled out the possibility that the antiviral effect of siRNA is dependent on IFN. Our present study demonstrates for the first time that either p38 or c-myc siRNA can efficiently inhibit TPA-induced EBV reactivation in GT38 cells, indicating that p38- and/or c-myc-associated signaling pathways may play critical roles in the disruption of EBV latency by TPA.

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