Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300

Wang, Chong; Zhou, Hufeng; Xue, Yong; Liang, Jun; Narita, Yohei; Gerdt, Catherine; Zheng, Amy Y.; Runsheng, Jiang; Trudeau, Stephen J.; Peng, Chih‐Wen; Gewurz, Benjamin E.

doi:10.1128/jvi.02155-17

Cited by 19 publications

(27 citation statements)

References 53 publications

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“…Superenhancers have been proposed to form non-membrane-bound condensates by LLPS (33). Previous studies verified that EBV EBNA2 and EBNALP are two key components of superenhancers that regulate the transcription of key target genes (20,22,23,25). Here, we demonstrate that transcription factors EBNA2 and EBNALP function by forming LLPS nuclear condensates to drive superenhancer-mediated transcription (Fig.…”

Section: Discussionsupporting

confidence: 66%

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Phase Separation of Epstein-Barr Virus EBNA2 and Its Coactivator EBNALP Controls Gene Expression

Qiu

Wang

Qin

et al. 2020

J Virol

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Biological macromolecule condensates formed by liquid-liquid phase separation (LLPS) have been discovered in recent years to be prevalent in biology. These condensates are involved in diverse processes, including the regulation of gene expression. LLPS of proteins have been found in animal, plant, and bacterial species but have scarcely been identified in viral proteins. Here, we discovered that Epstein-Barr virus (EBV) EBNA2 and EBNALP form nuclear puncta that exhibit properties of liquid-like condensates (or droplets), which are enriched in superenhancers of MYC and Runx3. EBNA2 and EBNALP are transcription factors, and the expression of their target genes is suppressed by chemicals that perturb LLPS. Intrinsically disordered regions (IDRs) of EBNA2 and EBNALP can form phase-separated droplets, and specific proline residues of EBNA2 and EBNALP contribute to droplet formation. These findings offer a foundation for understanding the mechanism by which LLPS, previously determined to be related to the organization of P bodies, membraneless organelles, nucleolus homeostasis, and cell signaling, plays a key role in EBV-host interactions and is involved in regulating host gene expression. This work suggests a novel anti-EBV strategy where developing appropriate drugs of interfering LLPS can be used to destroy the function of the EBV’s transcription factors. IMPORTANCE Protein condensates can be assembled via liquid-liquid phase separation (LLPS), a process involving the concentration of molecules in a confined liquid-like compartment. LLPS allows for the compartmentalization and sequestration of materials and can be harnessed as a sensitive strategy for responding to small changes in the environment. This study identified the Epstein-Barr virus (EBV) proteins EBNA2 and EBNALP, which mediate virus and cellular gene transcription, as transcription factors that can form liquid-like condensates at superenhancer sites of MYC and Runx3. This study discovered the first identified LLPS of EBV proteins and emphasized the importance of LLPS in controlling host gene expression.

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Section: Discussionsupporting

confidence: 66%

“…Sequencing by chromatin immunoprecipitation (ChIP-seq) also reported that EBNALP converges at EBV superenhancer sites that have high and broad H3K4me1 and H3K27ac signals, which is a characteristic of superenhancers (24). Approximately 29% of EBNALP superenhancer sites, including MYC and Runx3, colocalize with EBNA2 (23,25).…”

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confidence: 99%

Phase Separation of Epstein-Barr Virus EBNA2 and Its Coactivator EBNALP Controls Gene Expression

Qiu

Wang

Qin

et al. 2020

J Virol

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“…The EBNA-LP protein may also contribute to the ability of P3HR1 to induce Wp-restricted lymphomas. EBNA-LP has been reported to function as a co-activator protein that collaborates with EBNA2 to activate certain viral promoters (including the LMP1 promoter) [70]. Interestingly, a recent study using an EBNA-LP deleted B95.8 EBV mutant reported that EBNA-LP is essential for the ability of EBV to transform human cord blood B cells in vitro although it is not absolutely required for the ability of EBV to transform adult peripheral blood B cells [71]; the exact reason for this phenotype is currently unknown.…”

Section: Plos Pathogensmentioning

confidence: 99%

EBNA2-deleted Epstein-Barr virus (EBV) isolate, P3HR1, causes Hodgkin-like lymphomas and diffuse large B cell lymphomas with type II and Wp-restricted latency types in humanized mice

Romero-Masters

Huebner

et al. 2020

PLoS Pathog

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EBV transforms B cells in vitro and causes human B-cell lymphomas including classical Hodgkin lymphoma (CHL), Burkitt lymphoma (BL) and diffuse large B-cell lymphoma (DLBCL). The EBV latency protein, EBNA2, transcriptionally activates the promoters of all latent viral protein-coding genes expressed in type III EBV latency and is essential for EBV's ability to transform B cells in vitro. However, EBNA2 is not expressed in EBV-infected CHLs and BLs in humans. EBV-positive CHLs have type II latency and are largely driven by the EBV LMP1/LMP2A proteins, while EBV-positive BLs, which usually have type I latency are largely driven by c-Myc translocations, and only express the EBNA1 protein and viral noncoding RNAs. Approximately 15% of human BLs contain naturally occurring EBNA2-deleted viruses that support a form of viral latency known as Wp-restricted (expressing the EBNA-LP, EBNA3A/3B/3C, EBNA1 and BHRF1 proteins), but whether Wp-restricted latency and/ or EBNA2-deleted EBV can induce lymphomas in humanized mice, or in the absence of c-Myc translocations, is unknown. Here we show that a naturally occurring EBNA2-deleted EBV strain (P3HR1) isolated from a human BL induces EBV-positive B-cell lymphomas in a subset of infected cord blood-humanized (CBH) mice. Furthermore, we find that P3HR1infected lymphoma cells support two different viral latency types and phenotypes that are mutually exclusive: 1) Large (often multinucleated), CD30-positive, CD45-negative cells reminiscent of the Reed-Sternberg (RS) cells in CHL that express high levels of LMP1 but not EBNA-LP (consistent with type II viral latency); and 2) smaller monomorphic CD30-negative DLBCL-like cells that express EBNA-LP and EBNA3A but not LMP1 (consistent with Wp-restricted latency). These results reveal that EBNA2 is not absolutely required for EBV

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“…EBNA2 activates key oncogenes, including MYC (8,(10)(11)(12)(13)(14). EBNALP strongly coactivates EBNA2 by dislodging transcription repressors and coactivating EP300 (15)(16)(17)(18)(19). EBNA3A and EBNA3C can repress p16 INK4A , p14 ARF , and BIM to prevent senescence and apoptosis (13,14,(20)(21)(22)(23)(24)(25).…”

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confidence: 99%

RNA Sequencing Analyses of Gene Expression during Epstein-Barr Virus Infection of Primary B Lymphocytes

Wang

Zhang

et al. 2019

J Virol

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Epstein-Barr virus (EBV) infection of human primary resting B lymphocytes (RBLs) leads to the establishment of lymphoblastoid cell lines (LCLs) that can grow indefinitely in vitro. EBV transforms RBLs through the expression of viral latency genes, and these genes alter host transcription programs. To globally measure the transcriptome changes during EBV transformation, primary human resting B lymphocytes (RBLs) were infected with B95.8 EBV for 0, 2, 4, 7, 14, 21, and 28 days, and poly(A) plus RNAs were analyzed by transcriptome sequencing (RNA-seq). Analyses of variance (ANOVAs) found 3,669 protein-coding genes that were differentially expressed (false-discovery rate [FDR] < 0.01). Ninety-four percent of LCL genes that are essential for LCL growth and survival were differentially expressed. Pathway analyses identified a significant enrichment of pathways involved in cell proliferation, DNA repair, metabolism, and antiviral responses. RNA-seq also identified long noncoding RNAs (lncRNAs) differentially expressed during EBV infection. Clustered regularly interspaced short palindromic repeat (CRISPR) interference (CRISPRi) and CRISPR activation (CRISPRa) found that CYTOR and NORAD lncRNAs were important for LCL growth. During EBV infection, type III EBV latency genes were expressed rapidly after infection. Immediately after LCL establishment, EBV lytic genes were also expressed in LCLs, and ∼4% of the LCLs express gp350. Chromatin immune precipitation followed by deep sequencing (ChIP-seq) and POLR2A chromatin interaction analysis followed by paired-end tag sequencing (ChIA-PET) data linked EBV enhancers to 90% of EBV-regulated genes. Many genes were linked to enhancers occupied by multiple EBNAs or NF-κB subunits. Incorporating these assays, we generated a comprehensive EBV regulome in LCLs. IMPORTANCE Epstein-Barr virus (EBV) immortalization of resting B lymphocytes (RBLs) is a useful model system to study EBV oncogenesis. By incorporating transcriptome sequencing (RNA-seq), chromatin immune precipitation followed by deep sequencing (ChIP-seq), chromatin interaction analysis followed by paired-end tag sequencing (ChIA-PET), and genome-wide clustered regularly interspaced short palindromic repeat (CRISPR) screen, we identified key pathways that EBV usurps to enable B cell growth and transformation. Multiple layers of regulation could be achieved by cooperations between multiple EBV transcription factors binding to the same enhancers. EBV manipulated the expression of most cell genes essential for lymphoblastoid cell line (LCL) growth and survival. In addition to proteins, long noncoding RNAs (lncRNAs) regulated by EBV also contributed to LCL growth and survival. The data presented in this paper not only allowed us to further define the molecular pathogenesis of EBV but also serve as a useful resource to the EBV research community.

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Epstein-Barr Virus Nuclear Antigen Leader Protein Coactivates EP300

Cited by 19 publications

References 53 publications

Phase Separation of Epstein-Barr Virus EBNA2 and Its Coactivator EBNALP Controls Gene Expression

Phase Separation of Epstein-Barr Virus EBNA2 and Its Coactivator EBNALP Controls Gene Expression

EBNA2-deleted Epstein-Barr virus (EBV) isolate, P3HR1, causes Hodgkin-like lymphomas and diffuse large B cell lymphomas with type II and Wp-restricted latency types in humanized mice

RNA Sequencing Analyses of Gene Expression during Epstein-Barr Virus Infection of Primary B Lymphocytes

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