Expression of Hepatitis B Virus X Protein Does Not Alter the Accumulation of Spontaneous Mutations in Transgenic Mice

Abstract

Chronic infection with hepatitis B virus (HBV) is one of the major etiological factors in the development of human hepatocellular carcinoma. Transgenic mice that express the HBV X protein (HBx) have previously been shown to be more sensitive to the effects of hepatocarcinogens, although the mechanism for this cofactor role remains unknown. The ability of HBx to inhibit DNA repair in transiently transfected cell lines suggests one possible pathway. In the present study, primary hepatocytes isolated from transgenic mice that possess the HBV X gene under the control of the human ␣-1-antitrypsin regulatory region (ATX mice) were found to be deficient in their ability to conduct unscheduled DNA synthesis in response to UV-induced DNA damage. In order to measure the impact of HBx expression on DNA repair in vivo, double-transgenic mice that express HBx and possess a bacteriophage lambda transgene were sacrificed at 30, 90, and 240 days of age. Mutation frequency was determined for high-molecular-weight liver DNA of ATX and control mice by functional analysis of the lambda transgene. Expression of HBx did not significantly increase the accumulation of spontaneous mutations. These results are consistent with previous studies of HBx transgenic mice in which no effect of HBx on liver histology was apparent. This new animal model provides a powerful system in which to investigate the in vivo cooperation between HBx expression and environmental carcinogens.The hepatitis B virus (HBV) X protein (HBx) is a 17-kDa regulatory protein necessary for the establishment of hepadnaviral infection in woodchucks and, presumably, in all mammals (9, 65). Detectable in both the cytoplasm and nucleus of infected cells (13,38,48,56), the essential role(s) for HBx in the life cycle of the virus remains to be established. Although HBx does not bind DNA directly, it is capable of transactivating cellular genes (reviewed in references 6, 8, 45, and 51) and can induce protein kinase signaling cascades (3,13,29,37). In addition to interacting with numerous cellular proteins (reviewed in reference 16), HBx demonstrates a deoxy-ATPase activity (12).Chronic infection with HBV is considered a major risk factor for the development of human hepatocellular carcinoma (HCC) (1, 49), but the exact mechanism(s) by which HBV infection leads to the development of liver cancer remains to be elucidated. An increased rate of hepatocyte cell death and regeneration caused by the host immune response to viral antigens undoubtedly contributes to the pathology of chronic HBV infection (reviewed in references 11 and 49). Several lines of evidence indicate that HBx may also contribute to the development of HCC. Although the majority of HBx transgenic mouse lines do not demonstrate an increased incidence of liver cancer under normal conditions (5,14,21,34,39,43), such mice are more susceptible to the tumorigenic effects of hepatocarcinogens (14, 50) or the oncogene c-myc (54). Based on these observations, HBx is believed to be an important cofactor in HBV-associated liv...

HBX causes cyclin D1 overexpression and development of breast cancer in transgenic animals that are heterozygous for p53

“…Taken together, these experiments demonstrate that HBX, as the SV40 t-antigen, does not exert a spontaneous proapoptotic effect on ME-cells in monoor multiparous animals. These findings are comparable to results obtained with ATX mice, where HBX did not induce apoptosis in liver cells (Madden et al, 2000). However, related studies suggest that HBX triggers an apoptotic process in hepatocytes of transgenic animals ).…”

Gap Junction–Mediated Intercellular Communication in A Long–Term Primary Mouse Hepatocyte Culture System

“…Primary cultures of adult C57BL6 mouse hepatocytes were isolated by collagenase perfusion of adult male C57BL6 mice (8-12 weeks of age, Jackson Laboratories, Bar Harbor, ME) as described previously. 20 The animals used for perfusion received humane care in accordance with the guidelines established by the National Research Council as outlined in the "Guide for the Care and Use of Laboratory Animals" prepared by the National Academy of Sciences, published by the National Institutes of Health (National Institutes of Health publication no. 86-23, revised 1985).…”

Transcriptional Regulation of the TFIIH Transcription Repair Components XPB and XPD by the Hepatitis B Virus x Protein in Liver Cells and Transgenic Liver Tissue

“…HBx has been shown to inhibit nucleotide excision repair (NER), a major mechanism by which cells repair DNA bulky adducts induced by carcinogens such as aflatoxins and UV radiation (25)(26)(27). Interestingly, altered NER activity has been observed in liver cells from HBx-transgenic mice (8). The observation that apoptotic signals are impaired in the presence of HBx (14,28,29) also suggests that HBx may alter the balance between DNA repair and apoptosis, two important cellular defense mechanisms.…”

“…This is consistent with two main features of human HCC: its long latency and its large regional variation. More important is the evidence of synergistic interaction between HBV and exposure to environmental factors in the high incidence of HCC in humans and transgenic mice (3)(4)(5)(6)(7)(8). Therefore, understanding molecular mechanisms by which HBV affects cellular susceptibility to hepatocarcinogenesis is essential to identify preventive strategies for liver cancer.…”