The Library

Epstein-Barr virus-encoded EBNA1 inhibits the canonical NF-κB pathway in carcinoma cells by inhibiting IKK phosphorylation

Tools

Valentine, Robert, Dawson, Christopher W., Hu, Chunfang, Shah, Khilan M., Owen, Thomas J., Date, Kathryn L., Maia, Sonia P., Shao, Jianyong, Arrand, J. R. (John R.), Young, Lawrence S. and O'Neil, John D.. (2010) Epstein-Barr virus-encoded EBNA1 inhibits the canonical NF-κB pathway in carcinoma cells by inhibiting IKK phosphorylation. Molecular Cancer, Vol.9 (No.1). p. 1. ISSN 1476-4598

Preview

Text
WRAP_Young_Epstein_Barr_1476-4598-9-1.pdf - Published Version
Download (1617Kb) | Preview

Official URL: http://dx.doi.org/10.1186/1476-4598-9-1

Abstract

Background
The Epstein-Barr virus (EBV)-encoded EBNA1 protein is expressed in all EBV-associated tumours, including undifferentiated nasopharyngeal carcinoma (NPC), where it is indispensable for viral replication, genome maintenance and viral gene expression. EBNA1's transcription factor-like functions also extend to influencing the expression of cellular genes involved in pathways commonly dysregulated during oncogenesis, including elevation of AP-1 activity in NPC cell lines resulting in enhancement of angiogenesis in vitro. In this study we sought to extend these observations by examining the role of EBNA1 upon another pathway commonly deregulated during carcinogenesis; namely NF-κB.

Results
In this report we demonstrate that EBNA1 inhibits the canonical NF-κB pathway in carcinoma lines by inhibiting the phosphorylation of IKKα/β. In agreement with this observation we find a reduction in the phosphorylation of IκBα and reduced phosphorylation and nuclear translocation of p65, resulting in a reduction in the amount of p65 in nuclear NF-κB complexes. Similar effects were also found in carcinoma lines infected with recombinant EBV and in the EBV-positive NPC-derived cell line C666-1. Inhibition of NF-κB was dependent upon regions of EBNA1 essential for gene transactivation whilst the interaction with the deubiquitinating enzyme, USP7, was entirely dispensable. Furthermore, in agreement with EBNA1 inhibiting p65 NF-κB we demonstrate that p65 was exclusively cytoplasmic in 11 out of 11 NPC tumours studied.

Conclusions
Inhibition of p65 NF-κB in murine and human epidermis results in tissue hyperplasia and the development of squamous cell carcinoma. In line with this, p65 knockout fibroblasts have a transformed phenotype. Inhibition of p65 NF-κB by EBNA1 may therefore contribute to the development of NPC by inducing tissue hyperplasia. Furthermore, inhibition of NF-κB is employed by viruses as an immune evasion strategy which is also closely linked to oncogenesis during persistent viral infection. Our findings therefore further implicate EBNA1 in playing an important role in the pathogenesis of NPC.

Item Type:

Journal Article

Subjects:

Q Science > QR Microbiology > QR355 Virology

Divisions:

Faculty of Medicine > Warwick Medical School

Library of Congress Subject Headings (LCSH):

Epstein-Barr virus -- Genetic aspects, Cancer -- Pathogenesis, Cancer -- Physiology

Journal or Publication Title:

Molecular Cancer

Publisher:

BioMed Central Ltd.

ISSN:

1476-4598

Official Date:

5 January 2010

Dates:

Date	Event
5 January 2010	Published

Volume:

Vol.9

Number:

No.1

Page Range:

p. 1

Identification Number:

10.1186/1476-4598-9-1

Status:

Peer Reviewed

Publication Status:

Published

Access rights to Published version:

Open Access

Funder:

Cancer Research UK (CRUK), Sixth Framework Programme (European Commission) (FP6)

Grant number:

LSHC-CT-2005-018704 (FP6)

References:

1. Kieff ERAB: Epstein-Barr virus and its replication. FieldsVirology Philadelphia:
Lippincott Williams & WilkinsKnipe Dm HP 2001, 2511-2574.
2. Raab-Traub N: Epstein-Barr virus in the pathogenesis of NPC. Semin
Cancer Biol 2002, 12:431-441.
3. Young LS, Murray PG: Epstein-Barr virus and oncogenesis: from latent
genes to tumours. Oncogene 2003, 22:5108-5121.
4. Holowaty MN, Frappier L: HAUSP/USP7 as an Epstein-Barr virus target.
Biochem Soc Trans 2004, 32:731-732.
5. Wilson JB, Bell JL, Levine AJ: Expression of Epstein-Barr virus nuclear
antigen-1 induces B cell neoplasia in transgenic mice. EMBO J 1996,
15:3117-3126.
6. Kang MS, Soni V, Bronson R, Kieff E: Epstein-Barr virus nuclear antigen 1
does not cause lymphoma in C57BL/6J mice. J Virol 2008, 82:4180-4183.
7. Kang MS, Hung SC, Kieff E: Epstein-Barr virus nuclear antigen 1 activates
transcription from episomal but not integrated DNA and does not alter
lymphocyte growth. Proc Natl Acad Sci USA 2001, 98:15233-15238.
8. Komano J, Sugiura M, Takada K: Epstein-Barr virus contributes to the
malignant phenotype and to apoptosis resistance in Burkitt’s lymphoma
cell line Akata. J Virol 1998, 72:9150-9156.
9. Ruf IK, Rhyne PW, Yang C, Cleveland JL, Sample JT: Epstein-Barr virus small
RNAs potentiate tumorigenicity of Burkitt lymphoma cells
independently of an effect on apoptosis. J Virol 2000, 74:10223-10228.
10. Baumforth KR, Birgersdotter A, Reynolds GM, Wei W, Kapatai G, Flavell JR,
Kalk E, Piper K, Lee S, Machado L, et al: Expression of the Epstein-Barr
virus-encoded Epstein-Barr virus nuclear antigen 1 in Hodgkin’s
lymphoma cells mediates Up-regulation of CCL20 and the migration of
regulatory T cells. Am J Pathol 2008, 173:195-204.
11. Kube D, Vockerodt M, Weber O, Hell K, Wolf J, Haier B, Grasser FA, Muller-
Lantzsch N, Kieff E, Diehl V, Tesch H: Expression of epstein-barr virus
nuclear antigen 1 is associated with enhanced expression of CD25 in
the Hodgkin cell line L428. J Virol 1999, 73:1630-1636.
12. Srinivas SK, Sixbey JW: Epstein-Barr virus induction of recombinaseactivating
genes RAG1 and RAG2. J Virol 1995, 69:8155-8158.
13. O’Neil JD, Owen TJ, Wood VH, Date KL, Valentine R, Chukwuma MB,
Arrand JR, Dawson CW, Young LS: Epstein-Barr virus-encoded EBNA1
modulates the AP-1 transcription factor pathway in nasopharyngeal
carcinoma cells and enhances angiogenesis in vitro. J Gen Virol 2008,
89:2833-2842.
14. Wood VH, O’Neil JD, Wei W, Stewart SE, Dawson CW, Young LS: Epstein-
Barr virus-encoded EBNA1 regulates cellular gene transcription and
modulates the STAT1 and TGFbeta signaling pathways. Oncogene 2007,
26:4135-4147.
15. Dresang LR, Vereide DT, Sugden B: Identifying sites bound by Epstein-Barr
virus nuclear antigen 1 (EBNA1) in the human genome: defining a
position-weighted matrix to predict sites bound by EBNA1 in viral
genomes. J Virol 2009, 83:2930-2940.
16. Stewart S, Dawson CW, Takada K, Curnow J, Moody CA, Sixbey JW,
Young LS: Epstein-Barr virus-encoded LMP2A regulates viral and cellular
gene expression by modulation of the NF-kappaB transcription factor
pathway. Proc Natl Acad Sci USA 2004, 101:15730-15735.
17. Karin M: NF-kappaB and cancer: mechanisms and targets. Mol Carcinog
2006, 45:355-361.
18. Renne R, Barry C, Dittmer D, Compitello N, Brown PO, Ganem D:
Modulation of cellular and viral gene expression by the latencyassociated
nuclear antigen of Kaposi’s sarcoma-associated herpesvirus. J
Virol 2001, 75:458-468.
19. Rodrigues L, Filipe J, Seldon MP, Fonseca L, Anrather J, Soares MP, Simas JP:
Termination of NF-kappaB activity through a gammaherpesvirus protein
that assembles an EC5S ubiquitin-ligase. EMBO J 2009, 28:1283-1295.
20. Sample J, Henson EB, Sample C: The Epstein-Barr virus nuclear protein 1
promoter active in type I latency is autoregulated. J Virol 1992, 66:4654-
4661.
21. Arenzana-Seisdedos F, Fernandez B, Dominguez I, Jacque JM, Thomas D,
Diaz-Meco MT, Moscat J, Virelizier JL: Phosphatidylcholine hydrolysis
activates NF-kappa B and increases human immunodeficiency virus
replication in human monocytes and T lymphocytes. J Virol 1993,
67:6596-6604.
22. Marechal V, Dehee A, Chikhi-Brachet R, Piolot T, Coppey-Moisan M,
Nicolas JC: Mapping EBNA-1 domains involved in binding to metaphase
chromosomes. J Virol 1999, 73:4385-4392.
23. Wu H, Kapoor P, Frappier L: Separation of the DNA replication,
segregation, and transcriptional activation functions of Epstein-Barr
nuclear antigen 1. J Virol 2002, 76:2480-2490.
24. Hussain SA, Ganesan R, Reynolds G, Gross L, Stevens A, Pastorek J,
Murray PG, Perunovic B, Anwar MS, Billingham L, et al: Hypoxia-regulated
carbonic anhydrase IX expression is associated with poor survival in
patients with invasive breast cancer. Br J Cancer 2007, 96:104-109.
25. Vermeulen L, De Wilde G, Van Damme P, Berghe Vanden W, Haegeman G:
Transcriptional activation of the NF-kappaB p65 subunit by mitogenand
stress-activated protein kinase-1 (MSK1). EMBO J 2003, 22:1313-1324.
26. Hayden MS, Ghosh S: Shared principles in NF-kappaB signaling. Cell 2008,
132:344-362.
27. Yamaoka S, Courtois G, Bessia C, Whiteside ST, Weil R, Agou F, Kirk HE,
Kay RJ, Israel A: Complementation cloning of NEMO, a component of the
IkappaB kinase complex essential for NF-kappaB activation. Cell 1998,
93:1231-1240.
28. Nishikori M: Classical and Alternative NF-�B Activation Pathways and
Their Roles in Lymphoid Malignancies. J Clin Exp Hematopathol 2005,
45:15-24.
29. Busson P, Ganem G, Flores P, Mugneret F, Clausse B, Caillou B, Braham K,
Wakasugi H, Lipinski M, Tursz T: Establishment and characterization of
three transplantable EBV-containing nasopharyngeal carcinomas. Int J
Cancer 1988, 42:599-606.
30. Huang YT, Sheen TS, Chen CL, Lu J, Chang Y, Chen JY, Tsai CH: Profile of
cytokine expression in nasopharyngeal carcinomas: a distinct expression
of interleukin 1 in tumor and CD4+ T cells. Cancer Res 1999, 59:1599-
1605.
31. Zwergal A, Quirling M, Saugel B, Huth KC, Sydlik C, Poli V, Neumeier D,
Ziegler-Heitbrock HW, Brand K: C/EBP beta blocks p65 phosphorylation
and thereby NF-kappa B-mediated transcription in TNF-tolerant cells. J
Immunol 2006, 177:665-672.
32. Ma N, Kawanishi M, Hiraku Y, Murata M, Huang GW, Huang Y, Luo DZ,
Mo WG, Fukui Y, Kawanishi S: Reactive nitrogen species-dependent DNA
damage in EBV-associated nasopharyngeal carcinoma: the relation to
STAT3 activation and EGFR expression. Int J Cancer 2008, 122:2517-2525.
33. Thornburg NJ, Pathmanathan R, Raab-Traub N: Activation of nuclear
factor-kappaB p50 homodimer/Bcl-3 complexes in nasopharyngeal
carcinoma. Cancer Res 2003, 63:8293-8301.
34. Bowie AG, Zhan J, Marshall WL: Viral appropriation of apoptotic and NFkappaB
signaling pathways. J Cell Biochem 2004, 91:1099-1108.
35. Cai QL, Knight JS, Verma SC, Zald P, Robertson ES: EC5S ubiquitin complex
is recruited by KSHV latent antigen LANA for degradation of the VHL
and p53 tumor suppressors. PLoS Pathog 2006, 2:e116.
36. Amir-Zilberstein L, Dikstein R: Interplay between E-box and NF-kappaB in
regulation of A20 gene by DRB sensitivity-inducing factor (DSIF). J Biol
Chem 2008, 283:1317-1323.
37. Seitz CS, Deng H, Hinata K, Lin Q, Khavari PA: Nuclear factor kappaB
subunits induce epithelial cell growth arrest. Cancer Res 2000, 60:4085-
4092.
38. Gapuzan ME, Yufit PV, Gilmore TD: Immortalized embryonic mouse
fibroblasts lacking the RelA subunit of transcription factor NF-kappaB
have a malignantly transformed phenotype. Oncogene 2002, 21:2484-
2492.
39. Seitz CS, Lin Q, Deng H, Khavari PA: Alterations in NF-kappaB function in
transgenic epithelial tissue demonstrate a growth inhibitory role for NFkappaB.
Proc Natl Acad Sci USA 1998, 95:2307-2312.
40. Dajee M, Lazarov M, Zhang JY, Cai T, Green CL, Russell AJ, Marinkovich MP,
Tao S, Lin Q, Kubo Y, Khavari PA: NF-kappaB blockade and oncogenic Ras
trigger invasive human epidermal neoplasia. Nature 2003, 421:639-643.
41. Chen RA, Ryzhakov G, Cooray S, Randow F, Smith GL: Inhibition of IkappaB
kinase by vaccinia virus virulence factor B14. PLoS Pathog 2008, 4:e22.
42. Hiscott J, Kwon H, Genin P: Hostile takeovers: viral appropriation of the
NF-kappaB pathway. J Clin Invest 2001, 107:143-151.
43. Santoro MG, Rossi A, Amici C: NF-kappaB and virus infection: who
controls whom. EMBO J 2003, 22:2552-2560.