scholarly journals Identification of protein kinases responsible for phosphorylation of Epstein–Barr virus nuclear antigen leader protein at serine-35, which regulates its coactivator function

2003 ◽  
Vol 84 (12) ◽  
pp. 3381-3392 ◽  
Author(s):  
Kentaro Kato ◽  
Akihiko Yokoyama ◽  
Yukinobu Tohya ◽  
Hiroomi Akashi ◽  
Yukihiro Nishiyama ◽  
...  
Keyword(s):  
Protein Kinases ◽  
Epstein Barr Virus ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr

scholarly journals Conserved Region CR2 of Epstein-Barr Virus Nuclear Antigen Leader Protein Is a Multifunctional Domain That Mediates Self-Association as well as Nuclear Localization and Nuclear Matrix Association

2002 ◽  
Vol 76 (3) ◽  
pp. 1025-1032 ◽  
Author(s):  
Michiko Tanaka ◽  
Akihiko Yokoyama ◽  
Mie Igarashi ◽  
Go Matsuda ◽  
Kentaro Kato ◽  
...  
Keyword(s):  
Amino Acid ◽  
Nuclear Localization ◽  
Nuclear Matrix ◽  
Epstein Barr Virus ◽  
Cellular Proteins ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr ◽  
Self Association

ABSTRACT Self-association of viral proteins is important for many of their functions, including enzymatic, transcriptional, and transformational activities. Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) contains various numbers of W1W2 repeats and a unique carboxyl-terminal Y1Y2 domain. It was reported that EBNA-LP associates with a variety of cellular proteins and plays a critical role in EBV-induced transformation. We report here that EBNA-LP self-associates in vivo and the domain responsible for the homotypic association is a multifunctional domain mediating nuclear localization, nuclear matrix association, and EBNA-2-dependent coactivator function of the protein. Our conclusions are based on the following observations. (i) EBNA-LP interacts with itself or its derivatives in the yeast two-hybrid system. (ii) A purified chimeric protein consisting of glutathione S-transferase fused to EBNA-LP specifically formed complexes with EBNA-LP transiently expressed in COS-7 cells. (iii) When Flag epitope-tagged EBNA-LP with either one or two W1W2 repeats and EBNA-LP containing four W1W2 repeats were coexpressed in COS-7 cells, the latter was specifically coimmunoprecipitated with the former. (iv) Mutational analyses of EBNA-LP with deletion mutants revealed that the region between codons 19 and 39 (relative to the first amino acid residue of the W2 domain) is essential for self-association of the protein. The mapped region almost completely overlaps with CR2 and CR3, regions conserved among a subset of primate γ-herpesviruses and critical for EBNA-2-dependent coactivator function. Amino acid substitutions in CR2 alone abolished the ability of the protein to self-interact. This laboratory previously reported that CR2 is also responsible for nuclear localization and nuclear matrix association (A. Yokoyama, Y. Kawaguchi, I. Kitabayashi, M. Ohki, and K. Hirai, Virology 279:401–413, 2001). (v) Sucrose gradient sedimentation showed that amino acid substitutions in CR2 reduced the ability of the protein to form protein complexes in B cells. These results suggest that self-association of EBNA-LP may be important for its various functions and interactions of the protein with multiple cellular proteins.

scholarly journals Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300

2018 ◽  
Vol 92 (9) ◽  
Author(s):  
Chong Wang ◽  
Hufeng Zhou ◽  
Yong Xue ◽  
Jun Liang ◽  
Yohei Narita ◽  
...  
Keyword(s):  
Transcription Factors ◽  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Nuclear Antigen ◽  
Transcription Activator ◽  
Lymphoproliferative Diseases ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr

ABSTRACTEpstein-Barr virus nuclear antigen (EBNA) leader protein (EBNALP) is one of the first viral genes expressed upon B-cell infection. EBNALP is essential for EBV-mediated B-cell immortalization. EBNALP is thought to function primarily by coactivating EBNA2-mediated transcription. Chromatin immune precipitation followed by deep sequencing (ChIP-seq) studies highlight that EBNALP frequently cooccupies DNA sites with host cell transcription factors (TFs), in particular, EP300, implicating a broader role in transcription regulation. In this study, we investigated the mechanisms of EBNALP transcription coactivation through EP300. EBNALP greatly enhanced EP300 transcription activation when EP300 was tethered to a promoter. EBNALP coimmunoprecipitated endogenous EP300 from lymphoblastoid cell lines (LCLs). EBNALP W repeat serine residues 34, 36, and 63 were required for EP300 association and coactivation. Deletion of the EP300 histone acetyltransferase (HAT) domain greatly reduced EBNALP coactivation and abolished the EBNALP association. An EP300 bromodomain inhibitor also abolished EBNALP coactivation and blocked the EP300 association with EBNALP. EBNALP sites cooccupied by EP300 had significantly higher ChIP-seq signals for sequence-specific TFs, including SPI1, RelA, EBF1, IRF4, BATF, and PAX5. EBNALP- and EP300-cooccurring sites also had much higher H3K4me1 and H3K27ac signals, indicative of activated enhancers. EBNALP-only sites had much higher signals for DNA looping factors, including CTCF and RAD21. EBNALP coactivated reporters under the control of NF-κB or SPI1. EP300 inhibition abolished EBNALP coactivation of these reporters. Clustered regularly interspaced short palindromic repeat interference targeting of EBNALP enhancer sites significantly reduced target gene expression, including that of EP300 itself. These data suggest a previously unrecognized mechanism by which EBNALP coactivates transcription through subverting of EP300 and thus affects the expression of LCL genes regulated by a broad range of host TFs.IMPORTANCEEpstein-Barr virus was the first human DNA tumor virus discovered over 50 years ago. EBV is causally linked to ∼200,000 human malignancies annually. These cancers include endemic Burkitt lymphoma, Hodgkin lymphoma, lymphoma/lymphoproliferative disease in transplant recipients or HIV-infected people, nasopharyngeal carcinoma, and ∼10% of gastric carcinoma cases. EBV-immortalized human B cells faithfully model key aspects of EBV lymphoproliferative diseases and are useful models of EBV oncogenesis. EBNALP is essential for EBV to transform B cells and transcriptionally coactivates EBNA2 by removing repressors from EBNA2-bound DNA sites. Here, we found that EBNALP can also modulate the activity of the key transcription activator EP300, an acetyltransferase that activates a broad range of transcription factors. Our data suggest that EBNALP regulates a much broader range of host genes than was previously appreciated. A small-molecule inhibitor of EP300 abolished EBNALP coactivation of multiple target genes. These findings suggest novel therapeutic approaches to control EBV-associated lymphoproliferative diseases.

scholarly journals Genetic analysis of the Epstein–Barr virus-coded leader protein EBNA-LP as a co-activator of EBNA2 function

2001 ◽  
Vol 82 (12) ◽  
pp. 3067-3079 ◽  
Author(s):  
Eamon M. McCann ◽  
Gemma L. Kelly ◽  
Alan B. Rickinson ◽  
Andrew I. Bell
Keyword(s):  
Genetic Analysis ◽  
Epstein Barr Virus ◽  
Cell Transformation ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Lmp1 Expression ◽  
Leader Protein ◽  
Epstein Barr ◽  
The Individual ◽  
Unique Domain

Co-operation between the Epstein–Barr virus (EBV)-coded leader protein EBNA-LP and the nuclear antigen EBNA2 appears to be critical for efficient virus-induced B cell transformation. Here we report the genetic analysis of EBNA-LP function using two transient co-transfection assays of co-operativity, activation of latent membrane protein 1 (LMP1) expression from a resident EBV genome in Akata-BL cells and activation of an EBNA2-responsive reporter construct. Small deletions were introduced into each of five conserved regions (CRs) of EBNA-LP sequence present in type 1 and type 2 EBV strains and in several primate lymphocryptovirus EBNA-LP homologues. Deletions within all three CRs in the EBNA-LP W1W2 repeat domain completely abrogated function, through inhibition of nuclear localization in the cases of CR1 and CR2 but not of CR3; deletions within CR4 and CR5 in the Y1Y2 unique domain had relatively little effect, yet loss of the whole Y2 sequence blocked activity. Alanine substitution of serine residues within potential phosphorylation sites identified two mutants of particular interest. Substitution of three such residues (S34,36,63) within W1W2 not only abrogated EBNA-LP activity but was associated with a complete loss of EBNA2 detectability in co-transfected cells, implying possible destabilization of the co-expressed EBNA2 protein. More importantly the individual substitution of S36 completely blocked EBNA-LP/EBNA2 co-operativity while retaining EBNA2 expression. We infer critical roles for the CR3 domain and for the S36 residue in EBNA-LP’s co-operative function.

scholarly journals Epstein-Barr Virus Nuclear Antigen Leader Protein Induces Expression of Thymus- and Activation-Regulated Chemokine in B Cells

2004 ◽  
Vol 78 (8) ◽  
pp. 3984-3993 ◽  
Author(s):  
Mikiko Kanamori ◽  
Shinya Watanabe ◽  
Reiko Honma ◽  
Masayuki Kuroda ◽  
Shosuke Imai ◽  
...  
Keyword(s):  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Epstein Barr Virus ◽  
Critical Role ◽  
Nuclear Antigen ◽  
Viral Gene ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr

ABSTRACT Epstein-Barr virus (EBV) nuclear antigen leader protein (EBNA-LP) plays a critical role in transformation of primary B lymphocytes to continuously proliferating lymphoblastoid cell lines (LCLs). To identify cellular genes in B cells whose expression is regulated by EBNA-LP, we performed microarray expression profiling on an EBV-negative human B-cell line, BJAB cells, that were transduced by a retroviral vector expressing the EBV EBNA-LP (BJAB-LP cells) and on BJAB cells that were transduced with a control vector (BJAB-vec cells). Microarray analysis led to the identification of a cellular gene encoding the CC chemokine TARC as a novel target gene that was induced by EBNA-LP. The levels of TARC mRNA expression and TARC secretion were significantly up-regulated in BJAB-LP compared with BJAB-vec cells. Induction of TARC was also observed when a subline of BJAB cells was converted by a recombinant EBV. Among the EBV-infected B-cell lines with the latency III phenotype that were tested, the LCLs especially secreted significantly high levels of TARC. The level of TARC secretion appeared to correlate with the level of full-length EBNA-LP expression. These results indicate that EBV infection induces TARC expression in B cells and that EBNA-LP is one of the viral gene products responsible for the induction.

scholarly journals The Conserved Domain CR2 of Epstein–Barr Virus Nuclear Antigen Leader Protein Is Responsible Not Only for Nuclear Matrix Association but Also for Nuclear Localization

Virology ◽  
2001 ◽  
Vol 279 (2) ◽  
pp. 401-413 ◽  
Author(s):  
Akihiko Yokoyama ◽  
Yasushi Kawaguchi ◽  
Issay Kitabayashi ◽  
Misao Ohki ◽  
Kanji Hirai
Keyword(s):  
Nuclear Localization ◽  
Nuclear Matrix ◽  
Epstein Barr Virus ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Conserved Domain ◽  
Leader Protein ◽  
Epstein Barr

scholarly journals The Epstein-Barr virus nuclear antigen leader protein associates with hsp72/hsc73.

1995 ◽  
Vol 69 (12) ◽  
pp. 8169-8172 ◽  
Author(s):  
J B Mannick ◽  
X Tong ◽  
A Hemnes ◽  
E Kieff
Keyword(s):  
Epstein Barr Virus ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr

Ultrastructural Characterization of Hearts from Transgenic Mice Expressing the Epstein-Barr Virus Nuclear Antigen-Leader Protein

1999 ◽  
Vol 5 (S2) ◽  
pp. 1172-1173
Author(s):  
J.R. Megill ◽  
T. M. Monticello ◽  
M.H. French ◽  
P.L. Smith ◽  
N.C. Trippodo ◽  
...  
Keyword(s):  
Heart Failure ◽  
Transgenic Mice ◽  
Epstein Barr Virus ◽  
Transgenic Line ◽  
Complex Life Cycle ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Nuclear Antigens ◽  
Leader Protein ◽  
Epstein Barr

Mice expressing the Epstein-Barr virus nuclear antigen-leader protein (EBNA-LP) develop congestive heart failure and atrial thrombosis as early as four months of age. Epstein-Barr virus (EBV) is a common human herpes virus with a complex life cycle involving chronic replication in epithelial tissues. The EBNA-LP, one of six EBV nuclear antigens, has an unknown biochemical function but is associated with EBV-induced growth transformation. This transgenic line was originally produced to investigate oncogenic consequences, but unexpectedly resulted in mice with progressive heart failure. Since the progression and development of the heart failure is highly predictable, this transgenic line may serve as a valuable model for studying the pathophysiologic changes associated with human dilated cardiomyopathy. We describe here the ultrastructural characteristics of the myocardium from EBNA-LP transgenic mice in order to better delineate the pathogenesis of the associated heart disease.

scholarly journals Epstein-Barr virus nuclear antigen leader protein localizes to promoters and enhancers with cell transcription factors and EBNA2

2013 ◽  
Vol 110 (46) ◽  
pp. 18537-18542 ◽  
Author(s):  
D. Portal ◽  
H. Zhou ◽  
B. Zhao ◽  
P. V. Kharchenko ◽  
E. Lowry ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Epstein Barr Virus ◽  
Nuclear Antigen ◽  
Barr Virus ◽  
Leader Protein ◽  
Epstein Barr
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