title: Cooperation of tumor-derived HBx mutants and p53-249(ser) mutant in regulating cell proliferation, anchorage-independent growth and aneuploidy in a telomerase-immortalized normal human hepatocyte-derived cell line.
abstract: Hepatocellular carcinoma (HCC) is a common cancer, and hepatitis B virus (HBV) is a major etiological agent. Convincing epidemiological and experimental evidence also links HCC to aflatoxin, a naturally occurring mycotoxin that produces a signature p53-249(ser) mutation. Recently, we have reported that tumor-derived HBx variants encoded by HBV exhibited attenuated transactivation and proapoptotic functions but retained their ability to block p53-mediated apoptosis. These results indicate that mutations in HBx may contribute to the development of HCC. In this study, we determined whether tumor-derived HBx mutants along, or in cooperation with p53-249(ser), could alter cell proliferation and chromosome stability of normal human hepatocytes. To test this hypothesis, we established a telomerase immortalized normal human hepatocycte line HHT4 that exhibited a near diploid karyotype and expressed many hepatocyte-specific genes. We found that overexpression one of the tumor-derived HBx mutants, CT, significantly increased colony forming efficiency (CFE) while its corresponding wild-type allele CNT significantly decreased CFE in HHT4 cells. p53-249(ser) rescued CNT-mediated inhibition of colony formation. Although HHT4 cells lacked an anchorage independent growth capability as they did not form any colonies in soft agar, the CT-expressing HHT4 cells could form colonies, which could be significantly enhanced by p53-249(ser). Induction of aneuploidy could be observed in HHT4 cells expressing CT, but additionally recurring chromosome abnormalities could only be detected in cells coexpressing CT and p53-249(ser). Our results are consistent with the hypothesis that certain mutations in HBx and p53 at codon 249 may cooperate in contributing to liver carcinogenesis.
authors.affiliation: Laboratory of Human Carcinogenesis, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
authors.forename: Weidong
authors.initials: W
authors.lastname: Jiang
authors.forename: Xin Wei
authors.initials: XW
authors.lastname: Wang
authors.forename: Tamar
authors.initials: T
authors.lastname: Unger
authors.forename: Marshonna
authors.initials: M
authors.lastname: Forgues
authors.forename: Jin Woo
authors.initials: JW
authors.lastname: Kim
authors.forename: S Perwez
authors.initials: SP
authors.lastname: Hussain
authors.forename: Elise
authors.initials: E
authors.lastname: Bowman
authors.forename: Elisa A
authors.initials: EA
authors.lastname: Spillare
authors.forename: Michael M
authors.initials: MM
authors.lastname: Lipsky
authors.forename: Jeanne M
authors.initials: JM
authors.lastname: Meck
authors.forename: Luciane R
authors.initials: LR
authors.lastname: Cavalli
authors.forename: Bassem R
authors.initials: BR
authors.lastname: Haddad
authors.forename: Curtis C
authors.initials: CC
authors.lastname: Harris
mesh_terms: D000782:Aneuploidy; D017209:Apoptosis; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D002448:Cell Adhesion; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002871:Chromosome Banding; D003114:Colony-Forming Units Assay; D020869:Gene Expression Profiling; D006515:Hepatitis B virus; D022781:Hepatocytes / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D009154:Mutation / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis; D012333:RNA, Messenger / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D032681:Spectral Karyotyping; D019098:Telomerase / Q000378:metabolism*; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D016159:Tumor Suppressor Protein p53 / Q000235:genetics* / Q000378:metabolism; D054334:Viral Regulatory and Accessory Proteins
chemical_list: D015415:Biomarkers; D012333:RNA, Messenger; C495901:TP53 protein, human; D015534:Trans-Activators; D016159:Tumor Suppressor Protein p53; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D019098:Telomerase