Hepatitis B virus X protein differentially activates RAS-RAF-MAPK and JNK pathways in X-transforming versus non-transforming AML12 hepatocytes.
The hepatitis B virus (HBV) X protein (pX) is implicated in hepatocarcinogenesis of chronic HBV patients by an unknown mechanism. Activities of pX likely relevant to hepatocyte transformation include activation of the mitogenic RAS-RAF-MAPK and JNK pathways. To assess the importance of mitogenic pathway activation by pX in transformation, we employed a cellular model system composed of two tetracycline-regulated, pX-expressing cell lines, constructed in AML12-immortalized hepatocytes. This system includes the differentiated 3pX-1 and the de-differentiated 4pX-1 hepatocytes. Our studies have demonstrated that conditional pX expression transforms only 3pX-1 cells. Here, comparative in vitro kinase assays and various in vivo analyses demonstrate that pX affects an inverse activation of RAS-RAF-MAPK and JNK pathways in 3pX-1 versus 4pX-1 cells. Sustained pX-dependent RAS-RAF-MAPK pathway activation is observed in pX-transforming 3pX-1 cells, whereas sustained pX-dependent JNK pathway activation is observed in pX non-transforming 4pX-1 cells. This differential, pX-dependent mitogenic pathway activation affects differential activation of cAMP-response element-binding protein and c-Jun and determines the proliferative response of 3pX-1 and 4pX-1 cells. Furthermore, tetracycline-regulated, pX-NLS-expressing cell lines demonstrate that expression of the nuclear pX-NLS variant minimally activates the RAS-RAF-MAPK pathway and results in markedly reduced transformation. These results link sustained, pX-mediated activation of RAS-RAF-MAPK pathway to hepatocyte transformation.
Tarn
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Department of Basic Medical Sciences and Medicinal Chemistry and Molecular Pharmacology, Purdue University, West Lafayette, Indiana 47907, USA.
Lee
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Hu
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Ashendel
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Andrisani
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D002455:Cell Division; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D019925:Cyclin A / Q000235:genetics; D022781:Hepatocytes / Q000187:drug effects* / Q000473:pathology; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D015534:Trans-Activators / Q000633:toxicity*; D054334:Viral Regulatory and Accessory Proteins; D018631:ras Proteins / Q000378:metabolism*
D017362:Cyclic AMP Response Element-Binding Protein; D019925:Cyclin A; D016755:Proto-Oncogene Proteins c-jun; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D019908:Proto-Oncogene Proteins c-raf; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins