Induction of p16INK4a Is the Major Barrier to Proliferation when Epstein-Barr Virus (EBV) Transforms Primary B Cells into Lymphoblastoid Cell Lines

Skalska, Lenka; White, Rob; Parker, Gillian A.; Sinclair, Alison J.; Paschos, Kostas; Allday, Martin J.

doi:10.1371/journal.ppat.1003187

Cited by 78 publications

(166 citation statements)

References 65 publications

(130 reference statements)

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“…INK4a is a major barrier to proliferation in B cells transformed by EBV, a virus that has been implicated in the pathogenesis of cHL (47). The pathogenic role of EBV infection in cHL (48) is well known, and EBV has been found in HRS cells in about 40% of cases (10) and EBV may have a role in inhibiting p16…”

Section: Discussionmentioning

confidence: 99%

Cellular Senescence Markers p16INK4a and p21CIP1/WAF Are Predictors of Hodgkin Lymphoma Outcome

Caliò

Zamò

Ponzoni

et al. 2015

Clinical Cancer Research

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Purpose: There is evidence that Hodgkin Reed-Sternberg (HRS) cells in classical Hodgkin lymphoma (cHL) could display some molecular and morphologic markers of cellular senescence (CS). We hypothesized that CS mechanisms may have potential prognostic relevance in cHL and investigated whether the expression of the well-established CS biomarkers p21 CIP1/WAF1 and p16 INK4a by HRS cells might be predictive of the probability of event-free survival (EFS).Experimental Design: The study analyzed a retrospective cohort of 147 patients and the results were validated on a cohort of 91 patients independently diagnosed and treated in a different institution. p16INK4a and p21 CIP1/WAF1 were categorized as dichotomous variables (< or ! 30% of HRS cells at diagnosis) and evaluated in univariate and multivariate analysis.Results: Both molecules were independent prognostic factors. A positive staining of one of the two molecules in more than 30% HRS cells predicted a better EFS (P < 0.01). p16INK4a /p21together as a unique categorical variable (both <30%, either <30%, both ! 30%) sorted out three prognostic groups with better, intermediate, or worse outcome either overall or within I-II, bulky and advanced stages. The presence or the lack of the robust expression of p21 CIP1/WAF1 and/or p16 INK4a defined the prognosis in our series.Conclusions: These findings point to (i) the relevance of CS-related mechanisms in cHL, and to (ii) the prognostic value of a simple, reproducible, and low-cost immunohistochemical evaluation of p16INK4a and p21 CIP1/WAF1 expression.

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Section: Discussionmentioning

confidence: 99%

Cellular Senescence Markers p16INK4a and p21CIP1/WAF Are Predictors of Hodgkin Lymphoma Outcome

Caliò

Zamò

Ponzoni

et al. 2015

Clinical Cancer Research

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“…The highly proliferative nature of the LMP1-KO lymphomas (indicated by their high level of Ki67 expression) may also require expression of the EBV EBNA2 protein, which mimics the effect of constitutively activated Notch signaling (3). The EBV-encoded protein EBNA3C, which inhibits expression of the cell cycle inhibitor p16 (49), may also promote proliferation of the LMP1-KO EBVinfected cells. Notably, EBNA3C was also recently shown to stabilize the IRF4 protein in EBV-infected B cells (32), and we found that the LMP1-KO EBV-induced lymphomas expressed a high level of IRF4.…”

Section: Methodsmentioning

confidence: 99%

LMP1-deficient Epstein-Barr virus mutant requires T cells for lymphomagenesis

Dong¹,

Xu²,

Plowshay³

et al. 2014

J. Clin. Invest.

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formation comparison, the P value was calculated using a 2-tailed Fisher exact test. For viral load comparison, the P value was calculated using the Wilcoxon rank-sum test. A P value less than 0.05 was considered significant.Study approval. All animal experiments were approved by the University of Wisconsin-Madison IACUC and conducted in accordance with the NIH Guide for the Care and Use of Laboratory Animals. For human AIDS-related lymphomas, cases were collected from the Department of Pathology and Laboratory Medicine of Weill Cornell Medical College. All human samples, obtained with IRB and biospecimen use approval, were residual cases after diagnosis.

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“…Although EBNA3s bind an RBPJ domain that is distinct from the EBNA2/ICN binding site, they nevertheless limit EBNA2 activation by competing for RBPJ binding (15)(16)(17)(18). In LCLs and Burkitt lymphoma tumor cells, the EBNA3 proteins have been shown to regulate distinct but extensively overlapping sets of cell genes (19)(20)(21)(22)(23)(24)(25)(26). EBNA3 proteins have been implicated in the pathogenesis of Burkitt lymphoma and in attenuating an antiproliferative DNA damage response during EBV transformation of primary B lymphocytes (19,(27)(28)(29).…”

Section: Epstein-barr Virus (Ebv) Latent Gene Products Cause Human Camentioning

confidence: 99%

“…EBNA3 proteins have been implicated in the pathogenesis of Burkitt lymphoma and in attenuating an antiproliferative DNA damage response during EBV transformation of primary B lymphocytes (19,(27)(28)(29). Moreover, EBNA3A and EBNA3C repression of the CDKN2A-encoded tumor suppressors p16 and p14 is essential for LCL growth, requires interaction with RBPJ, and is associated with increased H3K27me3 modification at the CDKN2A promoter (22,23,(30)(31)(32).Despite significant advances in our understanding of the role of EBNA3 proteins in LCL growth, the basis for their different effects via RBPJ remains an area of active investigation. The selectivity in gene regulation suggests that EBNA3 proteins either target different RBPJ-bound sites or exert different effects at the same sites.…”

mentioning

confidence: 99%

Epstein-Barr Virus Nuclear Antigen 3 (EBNA3) Proteins Regulate EBNA2 Binding to Distinct RBPJ Genomic Sites

Wang

Welch

et al. 2016

J Virol

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Latent infection of B lymphocytes by Epstein-Barr virus (EBV) in vitro results in their immortalization into lymphoblastoid cell lines (LCLs); this latency program is controlled by the EBNA2 viral transcriptional activator, which targets promoters via RBPJ, a DNA binding protein in the Notch signaling pathway. Three other EBNA3 proteins (EBNA3A, EBNA3B, and EBNA3C) interact with RBPJ to regulate cell gene expression. The mechanism by which EBNAs regulate different genes via RBPJ remains unclear. Our chromatin immunoprecipitation with deep sequencing (ChIP-seq) analysis of the EBNA3 proteins analyzed in concert with prior EBNA2 and RBPJ data demonstrated that EBNA3A, EBNA3B, and EBNA3C bind to distinct, partially overlapping genomic locations. Although RBPJ interaction is critical for EBNA3A and EBNA3C growth effects, only 30 to 40% of EBNA3-bound sites colocalize with RBPJ. Using LCLs conditional for EBNA3A or EBNA3C activity, we demonstrate that EBNA2 binding at sites near EBNA3A-or EBNA3C-regulated genes is specifically regulated by the respective EBNA3. To investigate EBNA3 binding specificity, we identified sequences and transcription factors enriched at EBNA3A-, EBNA3B-, and EBNA3C-bound sites. This confirmed the prior observation that IRF4 is enriched at EBNA3A-and EBNA3C-bound sites and revealed IRF4 enrichment at EBNA3B-bound sites. Using IRF4-negative BJAB cells, we demonstrate that IRF4 is essential for EBNA3C, but not EBNA3A or EBNA3B, binding to specific sites. These results support a model in which EBNA2 and EBNA3s compete for distinct subsets of RBPJ sites to regulate cell genes and where EBNA3 subset specificity is determined by interactions with other cell transcription factors. IMPORTANCE Epstein-Barr virus (EBV) latent gene products cause human cancers and transform B lymphocytes into immortalized lymphoblastoid cell lines in vitro.EBV nuclear antigens (EBNAs) and membrane proteins constitutively activate pathways important for lymphocyte growth and survival. An important unresolved question is how four different EBNAs (EBNA2, -3A, -3B, and -3C) exert unique effects via a single transcription factor, RBPJ. Here, we report that each EBNA binds to distinct but partially overlapping sets of genomic sites. EBNA3A and EBNA3C specifically regulate EBNA2's access to different RBPJ sites, providing a mechanism by which each EBNA can regulate distinct cell genes. We show that IRF4, an essential regulator of B cell differentiation, is critical for EBNA3C binding specificity; EBNA3A and EBNA3B specificities are likely due to interactions with other cell transcription factors. EBNA3 titration of EBNA2 transcriptional function at distinct sites likely limits cell defenses that would be triggered by unchecked EBNA2 prooncogenic activity. E pstein-Barr virus (EBV) is a herpesvirus that infects over 90% of the population by adulthood. Primary EBV infection usually presents as a nonspecific illness in early childhood but often manifests as infectious mononucleosis in adolescents (1). Thereafter, EBV es...

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Induction of p16INK4a Is the Major Barrier to Proliferation when Epstein-Barr Virus (EBV) Transforms Primary B Cells into Lymphoblastoid Cell Lines

Cited by 78 publications

References 65 publications

Cellular Senescence Markers p16INK4a and p21CIP1/WAF Are Predictors of Hodgkin Lymphoma Outcome

Cellular Senescence Markers p16INK4a and p21CIP1/WAF Are Predictors of Hodgkin Lymphoma Outcome

LMP1-deficient Epstein-Barr virus mutant requires T cells for lymphomagenesis

Epstein-Barr Virus Nuclear Antigen 3 (EBNA3) Proteins Regulate EBNA2 Binding to Distinct RBPJ Genomic Sites

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