scholarly journals Efficient Replication of Epstein-Barr Virus-Derived Plasmids Requires Tethering by EBNA1 to Host Chromosomes

2013 ◽  
Vol 87 (23) ◽  
pp. 13020-13028 ◽  
Author(s):  
Theresa L. Hodin ◽  
Tanbir Najrana ◽  
John L. Yates
Keyword(s):  
Dna Binding ◽  
Epstein Barr Virus ◽  
Binding Domain ◽  
Hek293 Cells ◽  
Plasmid Replication ◽  
Nuclear Fraction ◽  
Dna Binding Domain ◽  
Host Chromosome ◽  
Barr Virus ◽  
Epstein Barr

The EBNA1 protein of Epstein-Barr virus enables plasmids carryingoriPboth to duplicate and to segregate efficiently in proliferating cells. EBNA1 recruits the origin recognition complex (ORC) to establish a replication origin at one element oforiP, DS (dyadsymmetry); at another element, FR (family ofrepeats), EBNA1 binds to an array of sites from which it tethers plasmids to host chromosomes for mitotic stability. We report experiments leading to the conclusion that tethering by EBNA1 to host chromosomes is also needed within interphase nuclei in order for plasmids to be replicated efficiently fromoriP. The DNA-binding domain of EBNA1, which lacks chromosome-binding ability, was found to support weak, DS-specific replication in HEK293 cells after transient transfection, being 17% as active as wild-type EBNA1. The low efficiency of replication was not due to the failure of the DNA-binding domain to retain plasmids within nuclei, because plasmids were recovered in similar amounts and entirely from the nuclear fraction of these transiently transfected cells. A derivative of EBNA1 with its chromosome-tethering domains replaced by a 22-amino-acid nucleosome-binding domain was fully active in supportingoriPfunctions. The implication is that EBNA1's DNA-binding domain is able to recruit ORC to DS, but either this step or subsequent replication is only efficient if the plasmid is tethered to a host chromosome. Finally, with some cell lines, DS can hardly support even transient plasmid replication without FR. A loss of plasmids lacking FR from nuclei cannot account for this requirement, suggesting that the stronger tethering to chromosomes by FR is needed for plasmid replication within the nuclei of such cells.

scholarly journals Crystal Structure of the DNA-Binding Domain of the Epstein–Barr Virus Origin-Binding Protein, EBNA1, Bound to DNA

Cell ◽  
1996 ◽  
Vol 84 (5) ◽  
pp. 791-800 ◽  
Author(s):  
Alexey Bochkarev ◽  
Jean A Barwell ◽  
Richard A Pfuetzner ◽  
Elena Bochkareva ◽  
Lori Frappier ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Dna Binding ◽  
Epstein Barr Virus ◽  
Binding Protein ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Barr Virus ◽  
Virus Origin ◽  
Origin Binding Protein ◽  
Epstein Barr

scholarly journals Crystal structure of the DNA-binding domain of the Epstein-Barr virus origin-binding protein EBNA1

Cell ◽  
1995 ◽  
Vol 83 (1) ◽  
pp. 39-46 ◽  
Author(s):  
Alexey Bochkarev ◽  
Jean A. Barwell ◽  
Richard A. Pfuetzner ◽  
William Furey ◽  
Aled M. Edwards ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Dna Binding ◽  
Epstein Barr Virus ◽  
Binding Protein ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Barr Virus ◽  
Virus Origin ◽  
Origin Binding Protein ◽  
Epstein Barr

scholarly journals High-resolution footprints of the DNA-binding domain of Epstein-Barr virus nuclear antigen 1.

1989 ◽  
Vol 9 (6) ◽  
pp. 2738-2742 ◽  
Author(s):  
A S Kimball ◽  
G Milman ◽  
T D Tullius
Keyword(s):  
Dna Binding ◽  
Epstein Barr Virus ◽  
Binding Domain ◽  
Nuclear Antigen ◽  
Dna Binding Domain ◽  
Major Groove ◽  
Barr Virus ◽  
Dna Binding Site ◽  
Epstein Barr ◽  
Hydroxyl Radical Footprinting

The DNA-binding domain of Epstein-Barr virus nuclear antigen 1 was found by hydroxyl radical footprinting to protect backbone positions on one side of its DNA-binding site. The guanines contacted in the major groove by the DNA-binding domain of Epstein-Barr virus nuclear antigen 1 were identified by methylation protection. No difference was found in the interaction of the DNA-binding domain of Epstein-Barr virus nuclear antigen 1 with tandemly repeated and overlapping binding sites.

High-resolution footprints of the DNA-binding domain of Epstein-Barr virus nuclear antigen 1

1989 ◽  
Vol 9 (6) ◽  
pp. 2738-2742
Author(s):  
A S Kimball ◽  
G Milman ◽  
T D Tullius
Keyword(s):  
Dna Binding ◽  
Epstein Barr Virus ◽  
Binding Domain ◽  
Nuclear Antigen ◽  
Dna Binding Domain ◽  
Major Groove ◽  
Barr Virus ◽  
Dna Binding Site ◽  
Epstein Barr ◽  
Hydroxyl Radical Footprinting

The DNA-binding domain of Epstein-Barr virus nuclear antigen 1 was found by hydroxyl radical footprinting to protect backbone positions on one side of its DNA-binding site. The guanines contacted in the major groove by the DNA-binding domain of Epstein-Barr virus nuclear antigen 1 were identified by methylation protection. No difference was found in the interaction of the DNA-binding domain of Epstein-Barr virus nuclear antigen 1 with tandemly repeated and overlapping binding sites.

scholarly journals BZLF1 Activation of the Methylated Form of the BRLF1 Immediate-Early Promoter Is Regulated by BZLF1 Residue 186

2005 ◽  
Vol 79 (12) ◽  
pp. 7338-7348 ◽  
Author(s):  
Prasanna M. Bhende ◽  
William T. Seaman ◽  
Henri-Jacques Delecluse ◽  
Shannon C. Kenney
Keyword(s):  
Dna Binding ◽  
Gene Transcription ◽  
Viral Genome ◽  
Gene Promoter ◽  
Binding Domain ◽  
Immediate Early ◽  
Dna Binding Domain ◽  
Barr Virus ◽  
Z Dna ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV) genome is highly methylated in latently infected cells. We recently reported that the EBV immediate-early (IE) protein BZLF1 (Z) preferentially binds to and activates transcription from the methylated form of the BRLF1 IE gene promoter (Rp). We now report that serine residue 186 in the Z DNA-binding domain plays an important role in the ability of Z to bind to and activate methylated Rp. A Z mutant containing an alanine residue at position 186 [Z(S186A)] was significantly defective in binding to methylated, as well as unmethylated, ZREs (Z-responsive elements) in Rp and was unable to activate lytic EBV gene transcription from the methylated or demethylated form of the viral genome. A Z mutant containing threonine at residue 186 [Z(S186T)] bound only to the methylated form of the ZRE-2 site in Rp and induced lytic EBV gene transcription from the methylated, but not demethylated, form of the viral genome. The defect in both of these mutants was primarily due to an inability to activate the Rp in the context of the viral genome. Finally, a Z mutant containing an aspartic acid at position 186 [Z(S186D)] did not bind to either the consensus AP-1 site or to the methylated or unmethylated Rp ZRE-2 site and did not induce lytic gene transcription. These results indicate that replacement of serine with threonine at residue 186 in the Z DNA-binding domain differentially affects its ability to reactivate the unmethylated, versus methylated, viral genome.

scholarly journals TRAF1 Is a Critical Regulator of JNK Signaling by the TRAF-Binding Domain of the Epstein-Barr Virus-Encoded Latent Infection Membrane Protein 1 but Not CD40

2003 ◽  
Vol 77 (2) ◽  
pp. 1316-1328 ◽  
Author(s):  
Aristides G. Eliopoulos ◽  
Elyse R. Waites ◽  
Sarah M. S. Blake ◽  
Clare Davies ◽  
Paul Murray ◽  
...  
Keyword(s):  
Membrane Protein ◽  
Epstein Barr Virus ◽  
Latent Infection ◽  
Binding Domain ◽  
Cell Phenotype ◽  
Tnf Receptor ◽  
Jnk Signaling ◽  
Barr Virus ◽  
Epstein Barr ◽  
Jnk Activation

ABSTRACT The oncogenic Epstein-Barr virus (EBV)-encoded latent infection membrane protein 1 (LMP1) mimics a constitutive active tumor necrosis factor (TNF) family receptor in its ability to recruit TNF receptor-associated factors (TRAFs) and TNF receptor-associated death domain protein (TRADD) in a ligand-independent manner. As a result, LMP1 constitutively engages signaling pathways, such as the JNK and p38 mitogen-activated protein kinases (MAPK), the transcription factor NF-κB, and the JAK/STAT cascade, and these activities may explain many of its pleiotropic effects on cell phenotype, growth, and transformation. In this study we demonstrate the ability of the TRAF-binding domain of LMP1 to signal on the JNK/AP-1 axis in a cell type- dependent manner that critically involves TRAF1 and TRAF2. Thus, expression of this LMP1 domain in TRAF1-positive lymphoma cells promotes significant JNK activation, which is blocked by dominant-negative TRAF2 but not TRAF5. However, TRAF1 is absent in many established epithelial cell lines and primary nasopharyngeal carcinoma (NPC) biopsy specimens. In these cells, JNK activation by the TRAF-binding domain of LMP1 depends on the reconstitution of TRAF1 expression. The critical role of TRAF1 in the regulation of TRAF2-dependent JNK signaling is particular to the TRAF-binding domain of LMP1, since a homologous region in the cytoplasmic tail of CD40 or the TRADD-interacting domain of LMP1 signal on the JNK axis independently of TRAF1 status. These data further dissect the signaling components used by LMP1 and identify a novel role for TRAF1 as a modulator of oncogenic signals.

DNA-binding-defective mutants of the Epstein-Barr virus lytic switch activator Zta transactivate with altered specificities

1994 ◽  
Vol 14 (5) ◽  
pp. 3041-3052
Author(s):  
E K Flemington ◽  
J P Lytle ◽  
C Cayrol ◽  
A M Borras ◽  
S H Speck
Keyword(s):  
Dna Binding ◽  
Epstein Barr Virus ◽  
Lytic Cycle ◽  
Promoter Elements ◽  
Barr Virus ◽  
Binding Forms ◽  
Viral Genes ◽  
Direct Binding ◽  
Epstein Barr ◽  
Necessary And Sufficient

The Epstein-Barr virus BRLF1 and BZLF1 genes are the first viral genes transcribed upon induction of the viral lytic cycle. The protein products of both genes (referred to here as Rta and Zta, respectively) activate expression of other viral genes, thereby initiating the lytic cascade. Among the viral antigens expressed upon induction of the lytic cycle, however, Zta is unique in its ability to disrupt viral latency; expression of the BZLF1 gene is both necessary and sufficient for triggering the viral lytic cascade. We have previously shown that Zta can activate its own promoter (Zp), through binding to two Zta recognition sequences (ZIIIA and ZIIIB). Here we describe mutant Zta proteins that do not bind DNA (referred to as Zta DNA-binding mutants [Zdbm]) but retain the ability to transactivate Zp. Consistent with the inability of these mutants to bind DNA, transactivation of Zp by Zdbm is not dependent on the Zta recognition sequences. Instead, transactivation by Zdbm is dependent upon promoter elements that bind cellular factors. An examination of other viral and cellular promoters identified promoters that are weakly responsive or unresponsive to Zdbm. An analysis of a panel of artificial promoters containing one copy of various promoter elements demonstrated a specificity for Zdbm activation that is distinct from that of Zta. These results suggest that non-DNA-binding forms of some transactivators retain the ability to transactivate specific target promoters without direct binding to DNA.

scholarly journals Epstein–Barr virus induces a distinct form of DNA-bound STAT1 compared with that found in interferon-stimulated B lymphocytes

2007 ◽  
Vol 88 (7) ◽  
pp. 1876-1886 ◽  
Author(s):  
James McLaren ◽  
Martin Rowe ◽  
Paul Brennan
Keyword(s):  
Dna Binding ◽  
Epstein Barr Virus ◽  
Lymphoblastoid Cell Lines ◽  
Distinct Form ◽  
Post Translational Modifications ◽  
Constitutive Activation ◽  
Barr Virus ◽  
Molecular Identity ◽  
Infected Cells ◽  
Epstein Barr

Since ‘constitutive activation’ of STAT1 was first described in Epstein–Barr virus (EBV)-immortalized lymphoblastoid cell lines (LCLs), there has been controversy regarding the molecular identity of the STAT1 DNA-binding complex found in these cells. The post-translational modifications of STAT1 in LCLs have been analysed and an LMP1-induced STAT1 DNA-binding complex, different from that generated by alpha interferon (IFN) stimulation and not involving tyrosine phosphorylation, is demonstrated. STAT1 is serine-phosphorylated downstream of PI3K and MEK in LCLs and this modification restricts IFN-stimulated STAT1–DNA binding. These data suggest that EBV induces a distinct form of DNA-bound STAT1 in virus-infected cells.

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