Identification of a rare Epstein-Barr virus variant that enhances early antigen expression in Raji cells.

Rabson, M. S.; Heston, Lee; Miller, George

doi:10.1073/pnas.80.9.2762

Cited by 105 publications

(92 citation statements)

References 23 publications

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“…We conducted an analysis of the regulation of Rp in an EBVpositive B-cell background, where EBV naturally persists in a latent state. We chose the Burkitt's lymphoma (BL) cell line Cl16 (HH514-16) for this purpose, since the EBV contained in these cells is tightly latent and yet can be induced efficiently into the lytic cycle by chemical stimuli or by transfection of ZEBRA or Rta expression vectors (30,62,80).…”

mentioning

confidence: 99%

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

Ragoczy

Miller

2001

J Virol

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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. Upon induction of the lytic cycle of Epstein-Barr virus (EBV), Rp and Zp, the two promoters directing the expression of the immediate-early genes BRLF1 and BZLF1, are activated simultaneously (15, 76). Despite their similar temporal regulation, it remains unclear whether the two promoters respond to the same or different signaling cascades. Only a few known response elements for transcriptional activators are contained within Rp (Fig. 1); these include binding sites for the cellular transcription factors YY1, Zif268 (Egr1), and Sp1 and the viral transactivator ZEBRA, the product of BZLF1 (14,70,(84)(85)(86). YY1 and Sp1 sites, as well as ZEBRA response elements (ZREs), are also present in Zp (14,48,58,70,73). ZEBRA activates Rp from the latent virus, presumably by directly binding to the ZREs (38, 42). The Zif268 sites have been implicated in phorbol ester-mediated activation of Rp, while the Sp1 sites were shown to affect the constitutive activity of Rp in cultured epithelial cells (85,86). However, their physiological roles and contributions to autostimulation of Rp in B cells have not been assessed.EBV remains predominantly latent in B lymphocytes, whereas epithelial cells are more permissive for lytic infection by the virus (5,43,57,71). Consistent with this biologic observation, Rp reporters exhibit significantly higher constitutive activities in epithelial tissue culture than in B cells (18,70,86). We conducted an analysis of the regulation of Rp in an EBVpositive B-cell background, where EBV naturally persists in a latent state. We chose the Burkitt's lymphoma (BL) cell line Cl16 (HH514-16) for this purpose, since the EBV contained in these cells is tightly latent and yet can be induced efficiently into the lytic cycle by chemical stimuli or by transfection of ZEBRA or Rta expression vectors (30,62,80).RpCAT reporters are not constitutively active in Cl16 cells and thus accurately reflect the behavior of the endogenous promoter. Moreover, in these cells Rta activates Rp both in the endogenous virus and when presented as a reporter construct (63). Since Rp lacks cognate binding sites for Rta, the mechanism o...

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confidence: 99%

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

Ragoczy

Miller

2001

J Virol

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“…A number of different stimuli have been shown to trigger the switch into the lytic cycle. These include halogenated pyrimidines (25), nutrient starvation (27), phorbol esters (61), calcium ionophores (16), transforming growth factor ␤1 (17), n-butyrate (36), specific histone deacetylase (HDAC) inhibitors such as trichostatin A (TSA) (9,29), the demethylating agent azacytidine (5), cross-linking of surface immunoglobulin (53), superinfection with EBV stocks containing rearranged defective DNA (41,43), and superinfection with human herpesvirus 6 (21). A general unanswered question is whether all inducing agents operate through a final common pathway.…”

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confidence: 99%

Protein Kinase C-Independent Activation of the Epstein-Barr Virus Lytic Cycle

Gradoville¹,

Kwa²,

El-Guindy

et al. 2002

J Virol

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The protein kinase C (PKC) pathway has been considered to be essential for activation of latent EpsteinBarr virus (EBV) into the lytic cycle. The phorbol ester tetradecanoyl phorbol acetate (TPA), a PKC agonist, is one of the best understood activators of EBV lytic replication. Zp, the promoter of the EBV immediate-early gene BZLF1, whose product, ZEBRA, drives the lytic cycle, contains several phorbol ester response elements. We investigated the role of the PKC pathway in lytic cycle activation in prototype cell lines that differed dramatically in their response to inducing agents. We determined whether PKC was involved in lytic cycle induction by histone deacetylase ( The switch between latency and the productive lytic cycle of Epstein-Barr virus (EBV) has been studied extensively in cultured lymphoid cell lines, a system that is advantageous for study of the molecular mechanism of the switch. Viral genes that are expressed during latency rather than during the lytic cycle are well characterized (31). The major viral products that activate the lytic cycle, the transcriptional activators encoded by the genes BZLF1 and BRLF1, have been identified (10,11,26). A number of different stimuli have been shown to trigger the switch into the lytic cycle. These include halogenated pyrimidines (25), nutrient starvation (27), phorbol esters (61), calcium ionophores (16), transforming growth factor ␤1 (17), n-butyrate (36), specific histone deacetylase (HDAC) inhibitors such as trichostatin A (TSA) (9, 29), the demethylating agent azacytidine (5), cross-linking of surface immunoglobulin (53), superinfection with EBV stocks containing rearranged defective DNA (41, 43), and superinfection with human herpesvirus 6 (21). A general unanswered question is whether all inducing agents operate through a final common pathway.The mechanisms mediating control of EBV lytic-cycle gene expression are understood in rough outline, but many important details are missing. During latency the genes BRLF1 and BZLF1, encoding the lytic-cycle activators, are repressed. No mRNA or proteins encoded by these genes are detectable during latency (55). Inducing stimuli that activate the lytic cycle lead to expression of BRLF1 and BZLF1.

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“…B95-8 cells are EBV-transformed marmoset B lymphocytes (15). HR-1 clone 5 and HR-1 clone 16 are single-cell clones of an EBV-positive human B-lymphocyte line derived from a Burkitt lymphoma (16,17). Raji is an EBV-positive Burkitt lymphoma cell line unable to produce viral particles (18).…”

Section: Methodsmentioning

confidence: 99%

Viral proteins associated with the Epstein-Barr virus transactivator, ZEBRA.

Katz

Baumann²,

Sun³

et al. 1992

Proc. Natl. Acad. Sci. U.S.A.

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The BamHI Z Epstein-Barr replication activator (ZEBRA) mediates disruption of latency and induction of Epstein-Barr virus (EBV) early gene expression in latently infected lymphocytes. Polyclonal rabbit sera raised against ZEBRA were used to immunoprecipitate ZEBRA-associated proteins (ZAPs). ZAPs of 19, 21, 23, and 42 kDa were coimmunoprecipitated with ZEBRA from extracts of EBVproducing lymphoid cell lines. ZAPs were not recognized directly by the rabbit sera, but they were antigenic for EBV+ human sera. Immunoprecipitation of ZAPs by ZEBRA-specific antisera required the presence of ZEBRA. ZAPs were not coprecipitated with ZEBRA from mouse cells expressing only ZEBRA, from Raji (a cell line in which EBV is unable to complete lytic replication), or from cells treated with inhibitors of viral DNA synthesis. Thus, ZAPs are late EBV-encoded proteins. ZEBRA and ZAPs colocalized to a salt-insoluble nuclear fraction, and both were found extracellularly in crude preparations of virions. ZAPs might function to affect the cellular localization of ZEBRA, to alter its capacity to transactivate, or to influence its target gene specificity.

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Identification of a rare Epstein-Barr virus variant that enhances early antigen expression in Raji cells.

Abstract: Early antigens (EAs) are made when the P3J-HR-1 strain of Epstein-Barr virus (EBV)

Cited by 105 publications

References 23 publications

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

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

Protein Kinase C-Independent Activation of the Epstein-Barr Virus Lytic Cycle

Viral proteins associated with the Epstein-Barr virus transactivator, ZEBRA.

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