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.

  • Article
  • Published:

Hepatitis B virus X protein promotes DNA damage propagation through disruption of liver polyploidization and enhances hepatocellular carcinoma initiation

Abstract

Hepatitis B virus X protein (HBx) contributes to Hepatitis B virus (HBV)-related liver cancer. However, its impact on hepatocyte proliferation and genomic stability remains elusive. We studied the role of HBx expression on the progression of cell cycle and liver polyploidization during proliferation and liver carcinogenesis. Full-length HBx transgenic mice (FL-HBx) were developed to investigate liver ploidy as well as hepatocyte proliferation, along normal liver maturation and during cancer initiation (chemical carcinogen treatment). Investigation of postnatal liver development in FL-HBx showed an aberrant G1/S and G2/M transitions, triggered (1) a delay of the formation of hepatocytes binucleation, (2) the early synthesis of polyploidy nuclei (≥4n) and (3) DNA damage appearance. Moreover, HBV infection during hepatocytes proliferation in a humanized liver mouse model led, to modifications in polyploidy of hepatocytes. In initiation of hepatocellular carcinoma, FL-HBx protein decreased ChK1 phosphorylation, Mre11 and Rad51 expression, upregulated IL-6 expression and impaired apoptosis. This was related to DNA damage accumulation in FL-HBx mice. At day 75 after initiation of hepatocellular carcinoma, FL-HBx mice revealed significant cell cycle changes related to the increased amount of 4n nuclei and of markers of cancer progenitor cells. Finally, PLK1 upregulation and p38/ERK activation in FL-HBx mice were implicated in aberrant polyploidization favoring DNA damage propagation and hepatocyte transformation. In conclusion, our data indicate that FL-HBx protein increases DNA damage through the hijack of hepatocyte polyploidization. That leads to enhancement of hepatocellular carcinoma initiation in an inflammatory context.

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

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

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

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Levrero M, Zucman-Rossi J. Mechanisms of HBV-induced hepatocellular carcinoma. J Hepatol. 2016;64:S84–101.

    Article  CAS  Google Scholar 

  2. Kew MC. Hepatitis B virus x protein in the pathogenesis of hepatitis B virus-induced hepatocellular carcinoma. J Gastroenterol Hepatol. 2011;26:144–52.

    Article  CAS  Google Scholar 

  3. Bandopadhyay M, Sarkar N, Datta S, Das D, Pal A, Panigrahi R, et al. Hepatitis B virus X protein mediated suppression of miRNA-122 expression enhances hepatoblastoma cell proliferation through cyclin G1-p53 axis. Infect Agent Cancer. 2016;11:40.

    Article  Google Scholar 

  4. Qiao L, Leach K, McKinstry R, Gilfor D, Yacoub A, Park JS, et al. Hepatitis B virus X protein increases expression ofp21(Cip-1/WAF1/MDA6) and p27(Kip-1) in primary mouse hepatocytes, leading to reduced cell cycle progression. Hepatology. 2001;34:906–17.

    Article  CAS  Google Scholar 

  5. Wu BK, Li CC, Chen HJ, Chang JL, Jeng KS, Chou CK, et al. Blocking of G1/S transition and cell death in the regenerating liver of Hepatitis B virus X protein transgenic mice. Biochem Biophys Res Commun. 2006;340:916–28.

    Article  CAS  Google Scholar 

  6. Quetier I, Brezillon N, Duriez M, Massinet H, Giang E, Ahodantin J, et al. Hepatitis B virus HBx protein impairs liver regeneration through enhanced expression of IL-6 in transgenic mice. J Hepatol. 2013;59:285–91.

    Article  CAS  Google Scholar 

  7. Na TY, Ka NL, Rhee H, Kyeong D, Kim MH, Seong JK, et al. Interaction of hepatitis B virus X protein with PARP1 results in inhibition of DNA repair in hepatocellular carcinoma. Oncogene. 2016;35:5435–45.

    Article  CAS  Google Scholar 

  8. Bontron S, Lin-Marq N, Strubin M. Hepatitis B virus X protein associated with UV-DDB1 induces cell death in the nucleus and is functionally antagonized by UV-DDB2. J Biol Chem. 2002;277:38847–54.

    Article  CAS  Google Scholar 

  9. Qadri I, Fatima K, Abde LHH. Hepatitis B virus X protein impedes the DNA repair via its association with transcription factor, TFIIH. BMC Microbiol. 2011;11:48.

    Article  CAS  Google Scholar 

  10. Yue D, Zhang Y, Cheng L, Ma J, Xi Y, Yang L, et al. Hepatitis B virus X protein (HBx)-induced abnormalities of nucleic acid metabolism revealed by (1)H-NMR-based metabonomics. Sci Rep. 2016;6:24430.

    Article  Google Scholar 

  11. Kim S, Lee HS, Ji JH, Cho MY, Yoo YS, Park YY, et al. Hepatitis B virus X protein activates the ATM-Chk2 pathway and delays cell cycle progression. J Gen Virol. 2015;96:2242–51.

    Article  CAS  Google Scholar 

  12. Cheng B, Zheng Y, Guo X, Wang Y, Liu C. Hepatitis B viral X protein alters the biological features and expressions of DNA repair enzymes in LO2 cells. Liver Int. 2010;30:319–26.

    Article  CAS  Google Scholar 

  13. Forgues M, Difilippantonio MJ, Linke SP, Ried T, Nagashima K, Feden J, et al. Involvement of Crm1 in hepatitis B virus X protein-induced aberrant centriole replication and abnormal mitotic spindles. Mol Cell Biol. 2003;23:5282–92.

    Article  CAS  Google Scholar 

  14. Yun C, Cho H, Kim SJ, Lee JH, Park SY, Chan GK. Mitotic aberration coupled with centrosome amplification is induced by hepatitis B virus X oncoprotein via the Ras-mitogen-activated protein/extracellular signal-regulated kinase-mitogen-activated protein pathway. Mol Cancer Res. 2004;2:159–69.

    CAS  PubMed  Google Scholar 

  15. Rakotomalala L, Studach L, Wang WH, Gregori G, Hullinger RL, Andrisani O. Hepatitis B virus X protein increases the Cdt1-to-geminin ratio inducing DNA re-replication and polyploidy. J Biol Chem. 2008;283:28729–40.

    Article  CAS  Google Scholar 

  16. Studach LL, Rakotomalala L, Wang WH, Hullinger RL, Cairo S, Buendia MA, et al. Polo-like kinase 1 inhibition suppresses hepatitis B virus X protein-induced transformation in an in vitro model of liver cancer progression. Hepatology. 2009;50:414–23.

    Article  CAS  Google Scholar 

  17. Gentric G, Desdouets C. Polyploidization in liver tissue. Am J Pathol. 2014;184:322–31.

    Article  CAS  Google Scholar 

  18. Duncan AW, Taylor MH, Hickey RD, Hanlon Newell AE, Lenzi ML, Olson SB, et al. The ploidy conveyor of mature hepatocytes as a source of genetic variation. Nature. 2010;467:707–10.

    Article  CAS  Google Scholar 

  19. Toyoda H, Bregerie O, Vallet A, Nalpas B, Pivert G, Brechot C, et al. Changes to hepatocyte ploidy and binuclearity profiles during human chronic viral hepatitis. Gut. 2005;54:297–302.

    Article  CAS  Google Scholar 

  20. Guidotti JE, Bregerie O, Robert A, Debey P, Brechot C, Desdouets C. Liver cell polyploidization: a pivotal role for binuclear hepatocytes. J Biol Chem. 2003;278:19095–101.

    Article  CAS  Google Scholar 

  21. Kudryavtsev BN, Kudryavtseva MV, Sakuta GA, Stein GI. Human hepatocyte polyploidization kinetics in the course of life cycle. Virchows Arch B Cell Pathol Incl Mol Pathol. 1993;64:387–93.

    Article  CAS  Google Scholar 

  22. Margall-Ducos G, Celton-Morizur S, Couton D, Bregerie O, Desdouets C. Liver tetraploidization is controlled by a new process of incomplete cytokinesis. J Cell Sci. 2007;120:3633–9.

    Article  CAS  Google Scholar 

  23. Celton-Morizur S, Merlen G, Couton D, Margall-Ducos G, Desdouets C. The insulin/Akt pathway controls a specific cell division program that leads to generation of binucleated tetraploid liver cells in rodents. J Clin Invest. 2009;119:1880–7.

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Duncan AW, Hanlon Newell AE, Smith L, Wilson EM, Olson SB, Thayer MJ, et al. Frequent aneuploidy among normal human hepatocytes. Gastroenterology. 2012;142:25–8.

    Article  Google Scholar 

  25. Pandit SK, Westendorp B, de Bruin A. Physiological significance of polyploidization in mammalian cells. Trends Cell Biol. 2013;23:556–66.

    Article  CAS  Google Scholar 

  26. Gentric G, Maillet V, Paradis V, Couton D, L’Hermitte A, Panasyuk G, et al. Oxidative stress promotes pathologic polyploidization in nonalcoholic fatty liver disease. J Clin Invest. 2015;125:981–92.

    Article  Google Scholar 

  27. Kawai H, Suda T, Aoyagi Y, Isokawa O, Mita Y, Waguri N, et al. Quantitative evaluation of genomic instability as a possible predictor for development of hepatocellular carcinoma: comparison of loss of heterozygosity and replication error. Hepatology. 2000;31:1246–50.

    Article  CAS  Google Scholar 

  28. Quetier I, Brezillon N, Revaud J, Ahodantin J, DaSilva L, Soussan P, et al. C-terminal-truncated hepatitis B virus X protein enhances the development of diethylnitrosamine-induced hepatocellular carcinogenesis. J Gen Virol. 2015;96:614–25.

    Article  CAS  Google Scholar 

  29. Schmit TL, Ahmad N. Regulation of mitosis via mitotic kinases: new opportunities for cancer management. Mol Cancer Ther. 2007;6:1920–31.

    Article  CAS  Google Scholar 

  30. Awad MM, Enslen H, Boylan JM, Davis RJ, Gruppuso PA. Growth regulation via p38 mitogen-activated protein kinase in developing liver. J Biol Chem. 2000;275:38716–21.

    Article  CAS  Google Scholar 

  31. Kurata S. Selective activation of p38 MAPK cascade and mitotic arrest caused by low level oxidative stress. J Biol Chem. 2000;275:38716–21.

    Article  Google Scholar 

  32. Tarn C, Zou L, Hullinger RL, Andrisani OM. Hepatitis B virus X protein activates the p38 mitogen-activated protein kinase pathway in dedifferentiated hepatocytes. J Virol. 2002;76:9763–72.

    Article  CAS  Google Scholar 

  33. He G, Dhar D, Nakagawa H, Font-Burgada J, Ogata H, Jiang Y, et al. Identification of liver cancer progenitors whose malignant progression depends on autocrine IL-6 signaling. Cell. 2013;155:384–96.

    Article  CAS  Google Scholar 

  34. Pandey V, Kumar V. HBx protein of hepatitis B virus promotes reinitiation of DNA replication by regulating expression and intracellular stability of replication licensing factor CDC6. J Biol Chem. 2012;287:20545–54.

    Article  CAS  Google Scholar 

  35. Gentric G, Desdouets C. Liver polyploidy: Dr Jekyll or Mr Hide? Oncotarget. 2015;6:8430–1.

    Article  Google Scholar 

  36. Tormos AM, Rius-Perez S, Jorques M, Rada P, Ramirez L, Valverde AM, et al. p38alpha regulates actin cytoskeleton and cytokinesis in hepatocytes during development and aging. PLoS ONE. 2017;12:e0171738.

    Article  Google Scholar 

  37. Parrilla A, Cirillo L, Thomas Y, Gotta M, Pintard L, Santamaria A. Mitotic entry: The interplay between Cdk1, Plk1 and Bora. Cell Cycle. 2016;15:3177–82.

    Article  CAS  Google Scholar 

  38. Studach L, Wang WH, Weber G, Tang J, Hullinger RL, Malbrue R, et al. Polo-like kinase 1 activated by the hepatitis B virus X protein attenuates both the DNA damage checkpoint and DNA repair resulting in partial polyploidy. J Biol Chem. 2010;285:30282–93.

    Article  CAS  Google Scholar 

  39. Wang SN, Lee KT, Tsai CJ, Chen YJ, Yeh YT. Phosphorylated p38 and JNK MAPK proteins in hepatocellular carcinoma. Eur J Clin Invest. 2012;42:1295–301.

    Article  CAS  Google Scholar 

  40. Wang Y, Cui R, Zhang X, Qiao Y, Liu X, Chang Y, et al. SIRT1 increases YAP- and MKK3-dependent p38 phosphorylation in mouse liver and human hepatocellular carcinoma. Oncotarget. 2016;7:11284–98.

    PubMed  PubMed Central  Google Scholar 

  41. He ZL, Zheng H, Lin H, Miao XY, Zhong DW. Overexpression of polo-like kinase1 predicts a poor prognosis in hepatocellular carcinoma patients. World J Gastroenterol. 2009;15:4177–82.

    Article  CAS  Google Scholar 

  42. Chaisaingmongkol J, Budhu A, Dang H, Rabibhadana S, Pupacdi B, Kwon SM, et al. Common molecular subtypes among Asian hepatocellular carcinoma and cholangiocarcinoma. Cancer Cell. 2017;32:57–70 e53.

    Article  CAS  Google Scholar 

  43. Wang C, Yang W, Yan HX, Luo T, Zhang J, Tang L, et al. Hepatitis B virus X (HBx) induces tumorigenicity of hepatic progenitor cells in 3,5-diethoxycarbonyl-1,4-dihydrocollidine-treated HBx transgenic mice. Hepatology. 2012;55:108–20.

    Article  CAS  Google Scholar 

  44. Wong VW, Yu J, Cheng AS, Wong GL, Chan HY, Chu ES, et al. High serum interleukin-6 level predicts future hepatocellular carcinoma development in patients with chronic hepatitis B. Int J Cancer. 2009;124:2766–70.

    Article  CAS  Google Scholar 

  45. Brezillon N, Brunelle MN, Massinet H, Giang E, Lamant C, DaSilva L, et al. Antiviral activity of Bay 41-4109 on hepatitis B virus in humanized Alb-uPA/SCID mice. PLoS ONE. 2011;6:e25096.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank S. Berissi and N. Gadessaud (Inserm US24/CNRS UMS3633) for tissue processing and slide staining, S. Morosan and O. Bregerie (UMS28, phénotyage du petit animal, UPMC-Paris VI) for animal care.

Funding

This work was supported by grants from the Agence Nationale de Recherches sur le Sida et les Hépatites Virales (ANRS) (AO 2014-1 16032), Ligue Contre le Cancer (2016, RS16/75-7 and 2017 RS17/75-23), Institut National de la Santé et de la Recherche Médicale (Inserm) and Université Pierre et Marie Curie (UPMC). J.A. was supported by an ANRS doctoral grant.

Author contributions

DK, JA, and CD contributed to conception of the study and design. JA, MB, CC, and JM performed experiments. All authors contributed to the interpretation of results. DK, JA, and CD drafted the manuscript.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Dina Kremsdorf.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Suppementary information

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Ahodantin, J., Bou-Nader, M., Cordier, C. et al. Hepatitis B virus X protein promotes DNA damage propagation through disruption of liver polyploidization and enhances hepatocellular carcinoma initiation. Oncogene 38, 2645–2657 (2019). https://doi.org/10.1038/s41388-018-0607-3

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41388-018-0607-3

This article is cited by

Search

Quick links