Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

Epigenetic repression of E-cadherin expression by hepatitis B virus x antigen in liver cancer

Oncogene volume 31, pages 563–572 (2012)Cite this article

Abstract

Loss of E-cadherin is associated with acquisition of metastatic capacity. Numerous studies suggest that histone deacetylation and/or hypermethylation of CpG islands in E-cadherin gene (CDH1) are major mechanisms responsible for E-cadherin silencing in different tumors and cancer cell lines. The hepatitis B virus (HBV)-encoded X antigen, HBx, contributes importantly to the development of hepatocellular carcinoma using multiple mechanisms. Experiments were designed to test if in addition to CDH1 hypermethylation HBx promotes epigenetic modulation of E-cadherin transcriptional activity through histone deacetylation and miR-373. The relationships between HBx, E-cadherin, mSin3A, Snail-1 and miR-373 were evaluated in HBx expressing (HepG2X) and control (HepG2CAT) cells by western blotting, immunoprecipitation (IP), chromatin IP as well as by immunohistochemical staining of liver and tumor tissue sections from HBV-infected patients. In HepG2X cells, decreased levels of E-cadherin and elevated levels of mSin3A and Snail-1 were detected. Reciprocal IP with anti-HBx and anti-mSin3A demonstrated mutual binding. Furthermore, HBx–mSin3A colocalization was detected by immunofluorescent staining. HBx downregulated E-cadherin expression by the recruitment of the mSin3A/histone deacetylase complex to the Snail-binding sites in human CDH1. Histone deacetylation inhibition by Trichostatin-A treatment restored E-cadherin expression. Mir-373, a positive regulator of E-cadherin expression, was downregulated by HBx in HepG2X cells and tissue sections from HBV-infected patients. Thus, histone deacetylation of CDH1 and downregulation of miR-373, together with the previously demonstrated hypermethylation of CDH1 by HBx, may be important for the understanding of HBV-related carcinogenesis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others

Abbreviations

CDH1 :

E-cadherin gene

DNMT:

DNA-methyltransferase

HBV:

hepatitis B virus

HBx:

hepatitis B x antigen

HCC:

hepatocellular carcinoma

HDAC1:

histone deacetylase 1

IP:

immunoprecipitation

TSA:

Trichostatin A

References

  • Agami R . (2010). Micro-RNAs, RNA binding proteins and cancer. Eur J Clin Invest 40: 370–374.

    Article  CAS  PubMed  Google Scholar 

  • Batlle E, Sancho E, Francí C, Domínguez D, Monfar M, Baulida J et al. (2000). The transcription factor Snail is a repressor of E-cadherin gene expression in epithelial tumour cells. Nat Cell Biol 2: 84–89.

    Article  CAS  PubMed  Google Scholar 

  • Cameron EE, Bachman KE, Myöhänen S, Herman JG, Baylin SB . (1999). Synergy of demethylation and histone deacetylase inhibition in the re-expression of genes silenced in cancer. Nat Genet 21: 103–107.

    Article  CAS  PubMed  Google Scholar 

  • Chung TW, Lee YC, Kim CH . (2004). Hepatitis B viral HBx induces MMP-9 gene expression through activation of ERK and PI-3K/AKT pathways: involvement of invasive potential. FASEB J 18: 1123–1125.

    Article  CAS  PubMed  Google Scholar 

  • Endo K, Ueda T, Ueyama J, Ohta T, Terada T . (2000). Immunoreactive E-cadherin, alpha-catenin, beta-catenin, and gamma-catenin proteins in hepatocellular carcinoma: relationships with tumor grade, clinicopathological parameters, and patients’ survival. Hum Pathol 31: 558–565.

    Article  CAS  PubMed  Google Scholar 

  • Feitelson MA, Millman I, Duncan GD, Blumberg BS . (1988). Presence of antibodies to the polymerase gene product(s) of hepatitis B and woodchuck hepatitis virus in natural and experimental infections. J Med Virol 24: 121–136.

    Article  CAS  PubMed  Google Scholar 

  • Giroldi L, Bringuier P, de Weijert M, Jansen C, van Bokhoven A, Schalken JA . (1997). Role of E boxes in the repression of E-cadherin expression. Biochem Biophys Res Commun 241: 453–458.

    Article  CAS  PubMed  Google Scholar 

  • Guarino M, Rubino B, Ballabio G . (2007). The role of epithelial-mesenchymal transition in cancer pathology. Pathology 39: 305–318.

    Article  CAS  PubMed  Google Scholar 

  • Haviv I, Vaizel D, Shaul Y . (1996). pX, the HBV-encoded co-activator, interacts with components of the transcription machinery and stimulates transcription in a TAF-independent manner. EMBO J 15: 3413–3420.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • He L, Hannon GJ . (2004). Micro-RNAs: small RNAs with a big role in gene regulation. Nat Rev Genet 5: 522–531.

    Article  CAS  PubMed  Google Scholar 

  • Hirohashi S, Kanai Y . (2003). Cell adhesion system and human cancer morphogenesis. Cancer Sci 94: 575–581.

    Article  CAS  PubMed  Google Scholar 

  • Iwatsuki M, Mimori K, Yokobori T, Ishi H, Beppu T, Nakamori S et al. (2009). Epithelial-mesenchymal transition in cancer development and its clinical significance. Cancer Sci 101: 293–299.

    Article  PubMed  Google Scholar 

  • Janowski BA, Younger ST, Hardy DB, Ram R, Huffman KE, Corey DR . (2007). Activating gene expression in mammalian cells with promoter-targeted duplex RNAs. Nat Chem Biol 3: 166–173.

    Article  CAS  PubMed  Google Scholar 

  • Jiao W, Miyazaki K, Kitajima Y . (2002). Inverse correlation between E-cadherin and Snail expression in hepatocellular carcinoma cell lines in vitro and in vivo. Br J Cancer 86: 98–101.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kanai Y, Ushijima S, Hui AM, Ochiai A, Tsuda H, Sakamoto M et al. (1997). The E-cadherin gene is silenced by CpG methylation in human hepatocellular carcinomas. Int J Cancer 71: 355–359.

    Article  CAS  PubMed  Google Scholar 

  • Keasler VV, Hodgson AJ, Madden CR, Slagle BL . (2009). Hepatitis B virus HBx protein localized to the nucleus restores HBx-deficient virus replication in HepG2 cells and in vivo in hydrodynamically-injected mice. Virology 390: 122–129.

    Article  CAS  PubMed  Google Scholar 

  • Kekule AS, Lauer U, Weiss L, Luber B, Hofschneider PH . (1993). Hepatitis B virus trans-activator HBx uses a tumour promoter signaling pathway. Nature 361: 742–745.

    Article  CAS  PubMed  Google Scholar 

  • Lee DK, Park SH, Yi Y, Choi SG, Lee C, Parks WT et al. (2001). The hepatitis B virus encoded oncoprotein pX amplifies TGF-beta family signaling through direct interaction with Smad4: potential mechanism of HBV-induced liver fibrosis. Genes Dev 15: 455–466.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lee JO, Kwun HJ, Jung JK, Choi KH, Min DS, Jang KL . (2005). Hepatitis B virus X protein represses E-cadherin expression via activation of DNA methyltransferase 1. Oncogene 24: 6617–6625.

    Article  CAS  PubMed  Google Scholar 

  • Li LC, Okino ST, Zhao H, Pookot D, Place RF, Urakami S et al. (2006). Small dsRNAs induce transcriptional activation in human cells. Proc Natl Acad Sci USA 103: 17337–17342.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lian Z, Liu J, Li L, Li X, Clayton M, Wu MC et al. (2006). Enhanced cell survival of Hep3B cells by the hepatitis B x antigen effector, URG11, is associated with upregulation of beta-catenin. Hepatology 43: 415–424.

    Article  CAS  PubMed  Google Scholar 

  • Lian Z, Pan J, Liu J, Zhang S, Zhu M, Arbuthnot P et al. (1999). The translation initiation factor, hu-Sui1 may be a target of hepatitis B X antigen in hepatocarcinogenesis. Oncogene 18: 1677–1587.

    Article  CAS  PubMed  Google Scholar 

  • Liu J, Lian Z, Han S, Waye MM, Wang H, Wu MC et al. (2006). Downregulation of E-cadherin by hepatitis B virus X antigen in hepatocellullar carcinoma. Oncogene 25: 1008–1017.

    Article  CAS  PubMed  Google Scholar 

  • Lupberger J, Hildt E . (2007). Hepatitis B virus-induced oncogenesis. World J Gastroenterol 13: 74–81.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • MacPherson MR, Molina P, Souchelnytskyi S, Wernstedt C, Martin-Pérez J, Portillo F et al. (2010). Phosphorylation of serine 11 and serine 92 as new positive regulators of human Snail1 function: potential involvement of casein kinase-2 and the cAMP-activated protein kinase A. Mol Biol Cell 21: 244–253.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Moustakas A, Heldin CH . (2007). Signaling networks guiding epithelial-mesenchymal transitions during embryogenesis and cancer progression. Cancer Sci 98: 1512–1520.

    Article  CAS  PubMed  Google Scholar 

  • Nomura T, Lin Y, Dorjsuren D, Ohno S, Yamashita T, Murakami S et al. (1999). Human hepatitis B virus X protein is detectable in nuclei of transfected cells, and is active for transactivation. Biochim Biophys Acta 1453: 330–340.

    Article  CAS  PubMed  Google Scholar 

  • Osada T, Sakamoto M, Ino Y, Iwamatsu A, Matsuno Y, Muto T et al. (1996). E-cadherin is involved in the intrahepatic metastasis of hepatocellular carcinoma. Hepatology 24: 1460–1467.

    Article  CAS  PubMed  Google Scholar 

  • Ou JN, Torrisani J, Unterberger A, Provençal N, Shikimi K, Karimi M et al. (2007). Histone deacetylase inhibitor Trichostatin A induces global and gene-specific DNA demethylation in human cancer cell lines. Biochem Pharmacol 73: 1297–1307.

    Article  CAS  PubMed  Google Scholar 

  • Pan J, Clayton M, Feitelson MA . (2004). Hepatitis B virus X antigen promotes TGF-beta1 activity by up-regulation of TGF-beta1 and down-regulation of alpha2-macroglobulin. J Gen Virol 85 (Part 2): 275–282.

    Article  CAS  PubMed  Google Scholar 

  • Park IY, Sohn BH, Yu E, Suh DJ, Chung YH, Lee JH et al. (2007). Aberrant epigenetic modifications in hepatocarcinogenesis induced by hepatitis B virus X protein. Gastroenterology 132: 1476–1494.

    Article  CAS  PubMed  Google Scholar 

  • Park NH, Song IH, Chung YH . (2006). Chronic hepatitis B in hepatocarcinogenesis. Postgrad Med J 82: 507–515.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Peinado H, Ballestar E, Esteller M, Cano A . (2004). Snail mediates E-cadherin repression by the recruitment of the Sin3A/HDAC1/HDAC2 complex. Mol Cell Biol 24: 306–319.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Peinado H, Quintanilla M, Cano A . (2003). TGFβ-1 induces snail transcription factor in epithelial cell lines: mechanisms for epithelial mesenchymal transitions. J Biol Chem 278: 113–123.

    Article  Google Scholar 

  • Perl AK, Wilgenbus P, Dahl U, Semb H, Christofori G . (1998). A causal role for E-cadherin in the transition from adenoma to carcinoma. Nature 392: 190–193.

    Article  CAS  PubMed  Google Scholar 

  • Place R, Li L, Pookot D, Noonan EJ, Dahiya R . (2008). MicroRNA-373 induces expression of genes with complementary promoter sequences. Proc Natl Acad Sci USA 105: 1608–1613.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Rikimaru T, Taketomi A, Yamashita Y, Shirabe K, Hamatsu T, Shimada M et al. (2007). Clinical significance of histone deacetylase 1 expression in patients with hepatocellular carcinoma. Oncology 72: 69–74.

    Article  CAS  PubMed  Google Scholar 

  • Ropero S, Esteller M . (2007). The role of histone deacetylases (HDACs) in human cancer. Mol Oncol 1: 19–25.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shon JK, Shon BH, Park IY, Lee SU, Fa L, Chang KY et al. (2009). Hepatitis B virus-X protein recruits histone deacetylase 1 to repress insulin-like growth factor binding protein 3 transcription. Virus Res 139: 14–21.

    Article  CAS  PubMed  Google Scholar 

  • Sirma H, Weil R, Rosmorduc O, Urban S, Israël A, Kremsdorf D et al. (1998). Cytosol is the prime compartment of hepatitis B virus X protein where it co-localizes with the proteasome. Oncogene 16: 2051–2063.

    Article  CAS  PubMed  Google Scholar 

  • Sugimachi K, Tanaka S, Kameyama T, Taguchi K, Aishima S, Shimada M et al. (2003). Transcriptional repressor Snail and progression of human hepatocellular carcinoma. Clin Cancer Res 9: 2657–2664.

    CAS  PubMed  Google Scholar 

  • Takeichi M . (1991). Cadherin cell adhesion receptors as a morphogenetic regulator. Science 251: 1451–1455.

    Article  CAS  PubMed  Google Scholar 

  • Takeno S, Noguchi T, Fumoto S, Kimura Y, Shibata T, Kawahara K . (2004). E-cadherin expression in patients with esophageal squamous cell carcinoma: Promoter hypermethylation, Snail overexpression, and clinico-pathologic implications. Am J Clin Pathol 122: 78–84.

    Article  CAS  PubMed  Google Scholar 

  • van Roy F, Berx G . (2008). The cell-cell adhesion molecule E-cadherin. Cell Mol Life Sci 65: 3756–3788.

    Article  CAS  PubMed  Google Scholar 

  • Wadia JS, Stan RV, Dowdy SF . (2004). Transducible TAT-HA fusogenic peptide enhances escape of TAT-fusion proteins after lipid raft macropinocytosis. Nat Med 1: 310–315.

    Article  Google Scholar 

  • Wang W, London WT, Feitelson MA . (1991a). Hepatitis B x antigen in hepatitis B virus carrier patients with liver cancer. Cancer Res 51: 4971–4977.

    CAS  PubMed  Google Scholar 

  • Wang W, London WT, Lega L, Feitelson MA . (1991b). HBxAg in liver from carrier patients with chronic hepatitis and cirrhosis. Hepatology 14: 29–37.

    Article  CAS  PubMed  Google Scholar 

  • Waris G, Huh KW, Siddiqui A . (2001). Mitochondrially associated hepatitis B virus X protein constitutively activates transcription factors STAT-3 and NF-kB via oxidative stress. Mol Cell Biol 21: 7721–7730.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Weil R, Sirma H, Giannini C, Kremsdorf D, Bessia C, Dargemont C et al. (1999). Direct association and nuclear import of the hepatitis B virus X protein with the NF-kB inhibitor IkBa. Mol Cell Biol 19: 6345–6354.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xu J, Lamouille S, Derynck R . (2009). TGF-beta-induced epithelial to mesenchymal transition. Cell Res 19: 156–172.

    Article  CAS  PubMed  Google Scholar 

  • Yoo YD, Ueda H, Park K, Flanders KC, Lee YI, Jay G et al. (1996). Regulation of TGF-beta 1 expression by the hepatitis B virus (HBV) X transactivator. Role in HBV pathogenesis. J Clin Invest 97: 388–395.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yoo YG, Na TY, Seo HW, Seong JK, Park CK, Shin YK et al. (2008). Hepatitis B virus X protein induces the expression of MTA1 and HDAC1, which enhances hypoxia signaling in hepatocellular carcinoma cells. Oncogene 27: 3405–3413.

    Article  CAS  PubMed  Google Scholar 

  • Zeng PY, Vakoc CR, Chen ZC, Blobel GA, Berger SL . (2006). In vivo dual cross-linking for identification of indirect DNA-associated proteins by chromatin immunoprecipitation. Biotechniques 41: 694–698.

    Article  CAS  PubMed  Google Scholar 

  • Zheng DL, Zhang L, Cheng N, Xu X, Deng Q, Teng XM et al. (2009). Epigenetic modification induced by hepatitis B virus X protein via interaction with de novo DNA methyltransferase DNMT3A. J Hepatol 50: 377–387.

    Article  CAS  PubMed  Google Scholar 

  • Zhou H, Wu S, Joo JY, Zhu S, Han DW, Lin T et al. (2009). Generation of induced pluripotent stem cells using recombinant proteins. Cell Stem Cell 4: 381–384.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This study was supported by NIH Grants 5R01CA104025 and 5R01CA111427 (to M Feitelson). We thank Drs Yongwen Chen and Cheng-ying Yang from the Third Military Medical University for the paraffin blocks of human liver tissue, and Dr Eugeney Bichenkov for assistance with the figures.

Author information

Authors and Affiliations

  1. Department of Biology, College of Science and Technology, Temple University, Philadelphia, PA, USA

    A Arzumanyan, T Friedman, E Kotei, Z Lian & M A Feitelson

  2. Center for Biotechnology, Sbarro Health Research Organization, College of Science and Technology, Temple University, Philadelphia, PA, USA

    A Arzumanyan & M A Feitelson

  3. Department of Pathology, University of Hong Kong, Queen Mary Hospital, Pokfulam, Hong Kong

    I O L Ng

Authors
  1. A Arzumanyan
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. T Friedman
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. E Kotei
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. I O L Ng
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Z Lian
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M A Feitelson
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to A Arzumanyan.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Additional information

Supplementary Information accompanies the paper on the Oncogene website

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Arzumanyan, A., Friedman, T., Kotei, E. et al. Epigenetic repression of E-cadherin expression by hepatitis B virus x antigen in liver cancer. Oncogene 31, 563–572 (2012). https://doi.org/10.1038/onc.2011.255

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/onc.2011.255

Keywords

This article is cited by

Search

Advanced search

Quick links