Epigenetic silencing of SFRP1 and SFRP5 by hepatitis B virus X protein enhances hepatoma cell tumorigenicity through Wnt signaling pathway

Authors

  • Qing Xie,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Linlin Chen,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Xuefeng Shan,

    1. The First Affiliated Hospital of Chongqing Medical University, Chongqing, China
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  • Xiaoliang Shan,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Jia Tang,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Fan Zhou,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Qingmei Chen,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Huiqin Quan,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Dan Nie,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Wenlu Zhang,

    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
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  • Ai-Long Huang,

    Corresponding author
    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
    • Correspondence to: Ni Tang, Tel.: +86-23-68486780, Fax: +86-23-68486780, E-mail: nitang@cqmu.edu.cn or Ai-Long Huang, Tel.: +86-23-68486135, Fax: +86-23-68486780, E-mail: ahuang@cqu.edu.cn

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  • Ni Tang

    Corresponding author
    1. The Second Affiliated Hospital and the Key Laboratory of Molecular Biology of Infectious Diseases designated by the Chinese Ministry of Education, Chongqing Medical University, Chongqing, China
    • Correspondence to: Ni Tang, Tel.: +86-23-68486780, Fax: +86-23-68486780, E-mail: nitang@cqmu.edu.cn or Ai-Long Huang, Tel.: +86-23-68486135, Fax: +86-23-68486780, E-mail: ahuang@cqu.edu.cn

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  • Conflict of Interest Statement: The authors have no conflicts of interest to declare.

  • Q.X., L.C. and X.S. contributed equally to this work

Abstract

Secreted frizzled-related proteins (SFRPs) are antagonists of the Wnt signaling pathway whose epigenetic downregulation have been shown to be involved in hepatocarcinogenesis. However, dysregulation of SFRPs induced by hepatitis B virus (HBV) X protein (HBx) has never been studied in HBV-related hepatocellular carcinoma (HBV-HCC). In this study, we sought to determine the clinical significance and underlying mechanism of HBx-induced SFRPs dysregulation in hepatoma cells and HBV-HCC patients. Our results showed that SFRP1 and SFRP5 expression were dramatically decreased by HBx in hepatoma cells. The repressed expression in hepatoma cells was partially rescued by a DNA methylation inhibitor and synergistically increased by a combination treatment with a histone deacetyltransferases inhibitor. In addition, we identified that SFRP1 and SFRP5 promoters were hypermethylated in both HBx-expressing hepatoma cells and HBV-HCC tissues. Downregulation of SFRP1 and SFRP5 in HBV-HCC tissues was significantly correlated with overexpression of DNA methyltransferase 1 (DNMT1) and poor tumor differentiation. HBx facilitated the binding of DNMT1 and DNMT3A to SFRP1 and SFRP5 promoters, and resulted in epigenetic silencing of SFRP1 and SFRP5. Moreover, overexpression of SFRP1, SFRP5 or RNA interference mediated silencing of DNMT1 inactivated the Wnt signaling pathway and decreased the expression levels of Wnt target genes c-Myc and CyclinD1, thus impeding HCC growth in vitro and in vivo, and regressing HBx-induced epithelial–mesenchymal transition (EMT). Our findings strongly suggest that epigenetic silencing of SFRP1 and SFRP5 by HBx allows constitutive activation of Wnt signaling pathway and hence contributes to hepatocarcinogenesis.

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