title: Induction of apoptosis after switch-on of the hepatitis B virus X gene mediated by the Cre/loxP recombination system.
abstract: The HBx protein of hepatitis B virus is a multifunctional protein that is implicated in the pathogenesis of hepatocellular carcinoma by regulating gene transcription, causing cell proliferation and, as shown recently, inducing cell death. However, analysis of the effects of HBx in stable cultured cell clones has been hampered because only cell lines that adapted to the effects of HBx were selected during the establishment of cell clones. Here, we describe a system in which transcription of the X gene of hepatitis B virus is switched on by the use of the site-specific Cre recombinase. Two human liver cell lines, HLF and HepG2, were used, the former with a mutant p53 allele and the latter with wild-type p53. The stable cell clones isolated, which carried the X gene in a transcriptionally silent state, were infected with recombinant adenovirus carrying Cre recombinase. Ninety-six hours after adenovirus infection, cell clones that expressed HBx had undergone TUNEL-positive cell death with characteristics of apoptosis. Apoptosis was induced despite concomitant inactivation of the p53 protein as a result of its cytoplasmic translocation by HBx. In contrast, neither the X gene-carrying cells infected with wild-type adenovirus nor various control cells infected with Cre-expressing adenovirus exhibited apoptosis. These results indicate that the expression of HBx protein leads to liver cell apoptosis independently of the p53 pathway. The significance of HBx-induced apoptosis in natural infection is unclear, but it may contribute to the development of hepatitis and serve to spread progeny virus to neighbouring cells while evading the host immune responses.
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Yoshizumi
authors.initials: Y
authors.lastname: Shintani
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Yotsuyanagi
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Kyoji
authors.initials: K
authors.lastname: Moriya
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Hajime
authors.initials: H
authors.lastname: Fujie
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Takeya
authors.initials: T
authors.lastname: Tsutsumi
authors.affiliation: Laboratory of Molecular Genetics, Institute of Medical Science, University of Tokyo, Tokyo 108-0072, Japan2.
authors.forename: Yumi
authors.initials: Y
authors.lastname: Kanegae
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kimura
authors.affiliation: Laboratory of Molecular Genetics, Institute of Medical Science, University of Tokyo, Tokyo 108-0072, Japan2.
authors.forename: Izumu
authors.initials: I
authors.lastname: Saito
authors.affiliation: First Department of Internal Medicine, University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8655, Japan1.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Koike
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis; D015139:Blotting, Southern; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D016158:Genes, p53; D005822:Genetic Vectors; D006801:Humans; D019426:Integrases / Q000235:genetics / Q000378:metabolism; D007763:Lac Operon; D011995:Recombination, Genetic; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism*; D015533:Transcriptional Activation; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D015534:Trans-Activators; D016159:Tumor Suppressor Protein p53; D014764:Viral Proteins; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; C045073:Cre recombinase; D019426:Integrases