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Carcinogenesis Advance Access originally published online on May 13, 2008
Carcinogenesis 2008 29(6):1207-1214; doi:10.1093/carcin/bgn111
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Mutations in the C-terminus of the X protein of hepatitis B virus regulate Wnt-5a expression in hepatoma Huh7 cells: cDNA microarray and proteomic analyses

Xiaohong Liu1,4, Li Wang1, Shuhui Zhang1, Jing Lin1, Shunmin Zhang1, Mark A. Feitelson2, Hengjun Gao3 and Minghua Zhu1,*

1 Department of Pathology, Changhai Hospital, Second Military Medical University, 174 Changhai Road, Shanghai 200433, People’s Republic of China
2 Department of Biology, College of Science and Technology, Temple University, 1900 North 12th Street, Philadelphia, PA 19122, USA
3 National Engineering Center for Biochip, Shanghai 201203, People’s Republic of China
4 Department of Pathology, Jinan Military General Hospital, Jinan 250031, People’s Republic of China

* To whom correspondence should be addressed. Tel: +86 21 25074604; Fax: +86 21 25074604; Email: mhzhu2000{at}hotmail.com

Background: The hepatitis B virus x gene (HBx) is a promiscuous transactivator implicated in the development of hepatocellular carcinoma (HCC). The present study was designed to investigate the molecular events regulated by HBx. Methods: Genomic and proteomic expression profiling was performed in Huh7 HCC cells transfected with HBx mutants with a C-terminal deletion. The gene and protein expression of wingless-type murine-mammary-tumour virus (MMTV) integration site family, member 5A (Wnt-5a) was validated by analyses of reverse transcription–polymerase chain reaction (RT–PCR), real-time RT–PCR, western blot and immunohistochemistry. Results: Differentially expressed genes and proteins were found in the transfected Huh7 HCC cells; most of them were involved in transcriptional regulation, although others including oncogenes or tumor suppressor genes, and molecules involved in cell junctions, signal transduction pathways, metabolism or the immune response were also observed. The expression of the Wnt-5a gene was elevated >10-fold in Huh7 cells transfected with the HBx3'-30 amino acid deletion mutant. However, the expression was downregulated by the transfection with the HBx3'-40 amino acid deletion mutant. The changes in Wnt-5a expression were also observed in human HCC tissues, compared with corresponding non-cancerous liver tissues. A negative correlation was found between the expression of Wnt-5a and HBx COOH mutations in HCC tissues. Conclusions: HBx mutants may participate in the development and progression of HCC, at least in part through the Wnt-5a pathway.

Abbreviations: cDNA, complementary DNA; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; HBV, hepatitis B virus; HBx, hepatitis B virus x gene; HCC, hepatocellular carcinoma; mRNA, messenger RNA; MMTV, murine-mammary-tumour virus; PCR, polymerase chain reaction; RT–PCR, reverse transcription–polymerase chain reaction; Wnt-5a, wingless-type MMTV integration site family, member 5A

Received January 7, 2008; revised April 27, 2008; accepted May 3, 2008.


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