scholarly journals The Epstein-Barr Virus-Encoded LMP2A and LMP2B Proteins Promote Epithelial Cell Spreading and Motility

2005 ◽  
Vol 79 (3) ◽  
pp. 1789-1802 ◽  
Author(s):  
Michael D. Allen ◽  
Lawrence S. Young ◽  
Christopher W. Dawson
Keyword(s):  
Protein Kinase ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Tyrosine Kinases ◽  
Epstein Barr Virus ◽  
Cell Spreading ◽  
Mitogen Activated Protein Kinase ◽  
Barr Virus ◽  
Mitogen Activated Protein ◽  
Epstein Barr

ABSTRACT The frequent expression of latent membrane proteins LMP2A and LMP2B in Epstein Barr virus (EBV)-associated tumors suggests that these proteins play a role in EBV-induced epithelial cell growth transformation. Expression of LMP2A and LMP2B had no effect on the morphology of squamous epithelial cells in monolayer culture, but their expression was associated with an increased capacity to spread and migrate on extracellular matrix. Although the mechanisms by which LMP2A and LMP2B promote cell spreading and motility are unclear, the use of selective pharmacological inhibitors has established a role for tyrosine kinases in this phenotype but ruled out contributions of phosphatidylinositol 3-kinase, extracellular signal-regulated kinase/mitogen-activated protein kinase, and protein kinase C. The ability of LMP2B to induce a phenotype that is virtually indistinguishable from that of LMP2A suggests that regions of the LMP2 protein in addition to the cytosolic amino terminus are capable of inducing phenotypic effects in epithelial cells. Thus, rather than serving to modulate the activity of LMP2A, LMP2B may directly engage signaling pathways to influence epithelial cell behavior such as cell adhesion and motility.

scholarly journals Epstein-Barr Virus (EBV) Tegument Protein BGLF2 Promotes EBV Reactivation through Activation of the p38 Mitogen-Activated Protein Kinase

2015 ◽  
Vol 90 (2) ◽  
pp. 1129-1138 ◽  
Author(s):  
XueQiao Liu ◽  
Jeffrey I. Cohen
Keyword(s):  
Protein Kinase ◽  
Signaling Pathways ◽  
Epstein Barr Virus ◽  
Virus Production ◽  
Mitogen Activated Protein Kinase ◽  
Tegument Protein ◽  
Barr Virus ◽  
Ebv Reactivation ◽  
Mitogen Activated Protein ◽  
Epstein Barr

ABSTRACTEpstein-Barr virus (EBV) is a ubiquitous gammaherpesvirus associated with both B cell and epithelial cell malignancies. EBV infection of B cells triggers activation of several signaling pathways that are critical for cell survival, virus latency, and growth transformation. To identify EBV proteins important for regulating cell signaling, we used a proteomic approach to screen viral proteins for AP-1 and NF-κB promoter activity in AP-1– and NF-κB–luciferase reporter assays. We found that EBV BGLF2 activated AP-1 but not NF-κB reporter activity. Expression of EBV BGLF2 in cells activated p38 and c-Jun N-terminal kinase (JNK), both of which are important for mitogen-activated protein kinase (MAPK) signaling. Deletion of the carboxyl-terminal 66 amino acids of BGLF2 reduced the ability of BGLF2 to activate JNK and p38. Expression of BGLF2 enhanced BZLF1 expression in latently EBV-infected lymphoblastoid cell lines, and knockdown of BGLF2 reduced EBV reactivation induced by IgG cross-linking. Expression of BGLF2 induced BZLF1 expression and virus production in EBV-infected gastric carcinoma cells. BGLF2 enhanced BZLF1 expression and EBV production by activating p38; chemical inhibition of p38 and MAPK/ERK kinases 1 and 2 (MEK1/2) reduced expression of BZLF1 and virus production induced by BGLF2. In summary, the EBV tegument protein BGLF2, which is delivered to the cell at the onset of virus infection, activates the AP-1 pathway and enhances EBV reactivation and virus production.IMPORTANCEEpstein-Barr virus (EBV) is associated with both B cell and epithelial cell malignancies, and the virus activates multiple signaling pathways important for its persistence in latently infected cells. We identified a viral tegument protein, BGLF2, which activates members of the mitogen-activated protein kinase signaling pathway. Expression of BGLF2 increased expression of EBV BZLF1, which activates a switch from latent to lytic virus infection, and increased production of EBV. Inhibition of BGFL2 expression or inhibition of p38/MAPK, which is activated by BGLF2, reduced virus reactivation from latency. These results indicate that a viral tegument protein which is delivered to cells upon infection activates signaling pathways to enhance virus production and facilitate virus reactivation from latency.

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.

scholarly journals Hippo signaling effectors YAP and TAZ induce Epstein-Barr Virus (EBV) lytic reactivation through TEADs in epithelial cells

2021 ◽  
Vol 17 (8) ◽  
pp. e1009783
Author(s):  
Nicholas Van Sciver ◽  
Makoto Ohashi ◽  
Nicholas P. Pauly ◽  
Jillian A. Bristol ◽  
Scott E. Nelson ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Hippo Signaling ◽  
Oral Keratinocytes ◽  
Barr Virus ◽  
Ebv Infection ◽  
Lytic Reactivation ◽  
Epstein Barr

The Epstein-Barr virus (EBV) human herpesvirus is associated with B-cell and epithelial-cell malignancies, and both the latent and lytic forms of viral infection contribute to the development of EBV-associated tumors. Here we show that the Hippo signaling effectors, YAP and TAZ, promote lytic EBV reactivation in epithelial cells. The transcriptional co-activators YAP/TAZ (which are inhibited by Hippo signaling) interact with DNA-binding proteins, particularly TEADs, to induce transcription. We demonstrate that depletion of either YAP or TAZ inhibits the ability of phorbol ester (TPA) treatment, cellular differentiation or the EBV BRLF1 immediate-early (IE) protein to induce lytic EBV reactivation in oral keratinocytes, and show that over-expression of constitutively active forms of YAP and TAZ reactivate lytic EBV infection in conjunction with TEAD family members. Mechanistically, we find that YAP and TAZ interact with, and activate, the EBV BZLF1 immediate-early promoter. Furthermore, we demonstrate that YAP, TAZ, and TEAD family members are expressed at much higher levels in epithelial cell lines in comparison to B-cell lines, and find that EBV infection of oral keratinocytes increases the level of activated (dephosphorylated) YAP and TAZ. Finally, we have discovered that lysophosphatidic acid (LPA), a known YAP/TAZ activator that plays an important role in inflammation, induces EBV lytic reactivation in epithelial cells through a YAP/TAZ dependent mechanism. Together these results establish that YAP/TAZ are powerful inducers of the lytic form of EBV infection and suggest that the ability of EBV to enter latency in B cells at least partially reflects the extremely low levels of YAP/TAZ and TEADs in this cell type.

scholarly journals Epstein-Barr-Virus-Encoded LMP2A Induces Primary Epithelial Cell Migration and Invasion: Possible Role in Nasopharyngeal Carcinoma Metastasis

2005 ◽  
Vol 79 (24) ◽  
pp. 15430-15442 ◽  
Author(s):  
Dirk M. Pegtel ◽  
Aravind Subramanian ◽  
Tzung-Shiahn Sheen ◽  
Ching-Hwa Tsai ◽  
Todd R. Golub ◽  
...  
Keyword(s):  
Cell Migration ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Epstein Barr Virus ◽  
Cellular Migration ◽  
Migration And Invasion ◽  
Barr Virus ◽  
Ebv Infection ◽  
Epithelial Cell Migration ◽  
Epstein Barr

ABSTRACT Nonkeratinizing nasopharyngeal carcinomas (NPC) are >95% associated with the expression of the Epstein-Barr virus (EBV) LMP2A latent protein. However, the role of EBV, in particular, LMP2A, in tumor progression is not well understood. Using Affymetrix chips and a pattern-matching computational technique (neighborhood analysis), we show that the level of LMP2A expression in NPC biopsy samples correlates with that of a cellular protein, integrin-alpha-6 (ITGα6), that is associated with cellular migration in vitro and metastasis in vivo. We have recently developed a primary epithelial model from tonsil tissue to study EBV infection in epithelial cells. Here we report that LMP2A expression in primary tonsil epithelial cells causes them to become migratory and invasive, that ITGα6 RNA levels are up-regulated in epithelial cells expressing LMP2, and that ITGα6 protein levels are increased in the migrating cells. Blocking antibodies against ITGα6 abrogated LMP2-induced invasion through Matrigel by primary epithelial cells. Our results provide a link between LMP2A expression, ITGα6 expression, epithelial cell migration, and NPC metastasis and suggest that EBV infection may contribute to the high incidence of metastasis in NPC progression.

scholarly journals Clustered MicroRNAs of the Epstein-Barr Virus Cooperatively Downregulate an Epithelial Cell-Specific Metastasis Suppressor

2014 ◽  
Vol 89 (5) ◽  
pp. 2684-2697 ◽  
Author(s):  
Teru Kanda ◽  
Mamiko Miyata ◽  
Makoto Kano ◽  
Satoru Kondo ◽  
Tomokazu Yoshizaki ◽  
...  
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Mirna Expression ◽  
Epstein Barr Virus ◽  
Epithelial Differentiation ◽  
Metastasis Suppressor ◽  
Barr Virus ◽  
Epstein Barr ◽  
Differentiation Marker ◽  
Bart Mirnas

ABSTRACTThe Epstein-Barr virus (EBV) encodes its own microRNAs (miRNAs); however, their biological roles remain elusive. The commonly used EBV B95-8 strain lacks a 12-kb genomic region, known as BamHI A rightward transcripts (BART) locus, where a number of BART miRNAs are encoded. Here, bacterial artificial chromosome (BAC) technology was used to generate an EBV B95-8 strain in which the 12-kb region was fully restored at its native locus [BART(+) virus]. Epithelial cells were stably infected with either the parental B95-8 virus or the BART(+) virus, and BART miRNA expression was successfully reconstituted in the BART(+) virus-infected cells. Microarray analyses of cellular gene expression identified N-myc downstream regulated gene 1 (NDRG1) as a putative target of BART miRNAs. The NDRG1 protein was barely expressed in B cells, highly expressed in epithelial cells, including primary epithelial cells, and strongly downregulated in the BART(+) virus-infected epithelial cells of various origins. Althoughin vitroreporter assays identified BART22 as being responsible for the NDRG1 downregulation, EBV genetic analyses revealed that BART22 was not solely responsible; rather, the entire BART miRNA cluster 2 was responsible for the downregulation. Immunohistochemical analyses revealed that the expression level of the NDRG1 protein was downregulated significantly in EBV-positive nasopharyngeal carcinoma specimens. Considering thatNDRG1encodes an epithelial differentiation marker and a suppressor of metastasis, these data implicate a causative relationship between BART miRNA expression and epithelial carcinogenesisin vivo.IMPORTANCEEBV-related epithelial cancers, such as nasopharyngeal carcinomas and EBV-positive gastric cancers, encompass more than 80% of EBV-related malignancies. Although it is known that they express high levels of virally encoded BART miRNAs, how these miRNAs contribute to EBV-mediated epithelial carcinogenesis remains unknown. Although a number of screenings have been performed to identify targets of viral miRNAs, many targets likely have not been identified, especially in case of epithelial cell infection. This is the first study to use EBV genetics to perform unbiased screens of cellular genes that are differentially expressed in viral miRNA-positive and -negative epithelial cells. The result indicates that multiple EBV-encoded miRNAs cooperatively downregulate NDRG1, an epithelial differentiation marker and suppressor of metastasis. The experimental system described in this study should be useful for further clarifying the mechanism of EBV-mediated epithelial carcinogenesis.

scholarly journals CD21-Dependent Infection of an Epithelial Cell Line, 293, by Epstein-Barr Virus

1999 ◽  
Vol 73 (3) ◽  
pp. 2115-2125 ◽  
Author(s):  
Joyce D. Fingeroth ◽  
Margaret E. Diamond ◽  
David R. Sage ◽  
Jody Hayman ◽  
John L. Yates
Keyword(s):  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Line ◽  
Epstein Barr Virus ◽  
Lymphoid Cells ◽  
Epithelial Cell Line ◽  
Barr Virus ◽  
293 Cells ◽  
Low Levels ◽  
Epstein Barr

ABSTRACT Epstein-Barr virus (EBV) is invariably present in undifferentiated nasopharyngeal carcinomas, is found sporadically in other carcinomas, and replicates in the differentiated layer of the tongue epithelium in lesions of oral hairy leukoplakia. However, it is not clear how frequently or by what mechanism EBV infects epithelial cells normally. Here, we report that a human epithelial cell line, 293, can be stably infected by EBV that has been genetically marked with a selectable gene. We show that 293 cells express a relatively low level of CD21, that binding of fluorescein-labeled EBV to 293 cells can be detected, and that both the binding of virus to cells and infection can be blocked with antibodies specific for CD21. Two proteins known to form complexes with CD21 on the surface of lymphoid cells, CD35 and CD19, could not be detected at the surface of 293 cells. All infected clones of 293 cells exhibited tight latency with a pattern of gene expression similar to that of type II latency, but productive EBV replication and release of infectious virus could be induced inefficiently by forced expression of the lytic transactivators, R and Z. Low levels of mRNA specific for the transforming membrane protein of EBV, LMP-1, as well as for LMP-2, were detected; however, LMP-1 protein was either undetectable or near the limit of detection at less than 5% of the level typical of EBV-transformed B cells. A slight increase in expression of the receptor for epidermal growth factor, which can be induced in epithelial cells by LMP-1, was detected at the cell surface with two EBV-infected 293 cell clones. These results show that low levels of surface CD21 can support infection of an epithelial cell line by EBV. The results also raise the possibility that in a normal infection of epithelial cells by EBV, the LMP-1 protein is not expressed at levels that are high enough to be oncogenic and that there might be differences in the cells of EBV-associated epithelial cancers that have arisen to allow for elevated expression of LMP-1.

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