Hepatitis B virus X protein differentially regulates cell cycle progression in X-transforming versus nontransforming hepatocyte (AML12) cell lines

J Biol Chem. 2002 Mar 8;277(10):8730-40. doi: 10.1074/jbc.M108025200. Epub 2001 Dec 26.

Abstract

Hepatitis B virus (HBV) X protein (pX) is implicated in hepatocarcinogenesis of chronically infected HBV patients. To understand mechanism(s) of pX-mediated cellular transformation, we employed two tetracycline-regulated, pX-expressing cell lines, constructed in AML12 immortalized hepatocytes: one a differentiated (3pX-1) and the other a de-differentiated (4pX-1) hepatocyte cell line. Only 3pX-1 cells undergo pX-mediated transformation, via sustained Ras-Raf-mitogen-activated protein kinase pathway activation. pX-nontransforming 4pX-1 cells display sustained, pX-dependent JNK pathway activation. To understand how pX mediates different growth characteristics in 3pX-1 and 4pX-1 cells, we report, herein, comparative cell cycle analyses. pX-transforming 3pX-1 cells display pX-dependent G(1), S, and G(2)/M progression evidenced by cyclin D(1), A, and B(1) induction, and Cdc2 kinase activation. pX-nontransforming 4pX-1 cells display pX-dependent G(1) and S phase entry, followed by S phase pause and absence of Cdc2 kinase activation. Interestingly, 4pX-1 cells exhibit selective pX-induced expression of cyclin-dependent kinase inhibitor p21(Cip1), tumor suppressor p19(ARF), and proapoptotic genes bax and IGFBP-3. Despite the pX-mediated induction of growth arrest and apoptotic genes and the absence of pX-dependent Cdc2 activation, 4pX-1 cells do not undergo pX-dependent G(2)/M arrest or apoptosis. Nocodazole-treated, G(2)/M-arrested 4pX-1 cells exhibit pX-dependent formation of multinucleated cells, similar to human T-cell lymphotropic virus type I Tax-expressing cells. We propose that in 4pX-1 cells, pX deregulates the G(2)/M checkpoint, thus rescuing cells from pX-mediated apoptosis.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Antineoplastic Agents / pharmacology
  • Blotting, Western
  • CDC2 Protein Kinase / metabolism
  • Cell Cycle
  • Cell Line
  • Cell Line, Transformed
  • Cyclin A / metabolism
  • Cyclin B / metabolism
  • Cyclin B1
  • Cyclin D1 / metabolism
  • Cyclin-Dependent Kinase Inhibitor p16
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins / metabolism
  • Flow Cytometry
  • Gene Expression Regulation
  • Hepatocytes / metabolism*
  • Humans
  • Insulin-Like Growth Factor Binding Protein 3 / metabolism
  • Mitogen-Activated Protein Kinase 8
  • Mitogen-Activated Protein Kinases / metabolism
  • Nocodazole / pharmacology
  • Promoter Regions, Genetic
  • Protein Binding
  • Protein Synthesis Inhibitors / pharmacology
  • Proto-Oncogene Proteins / metabolism
  • Proto-Oncogene Proteins c-bcl-2*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Tetracycline / pharmacology
  • Time Factors
  • Trans-Activators / metabolism*
  • Transcription, Genetic
  • Transfection
  • Tumor Cells, Cultured
  • Tumor Suppressor Protein p14ARF / metabolism
  • Viral Regulatory and Accessory Proteins
  • bcl-2-Associated X Protein

Substances

  • Antineoplastic Agents
  • BAX protein, human
  • CCNB1 protein, human
  • CDKN1A protein, human
  • Ccnb1 protein, mouse
  • Cdkn2a protein, mouse
  • Cyclin A
  • Cyclin B
  • Cyclin B1
  • Cyclin-Dependent Kinase Inhibitor p16
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins
  • Insulin-Like Growth Factor Binding Protein 3
  • Protein Synthesis Inhibitors
  • Proto-Oncogene Proteins
  • Proto-Oncogene Proteins c-bcl-2
  • Trans-Activators
  • Tumor Suppressor Protein p14ARF
  • Viral Regulatory and Accessory Proteins
  • bcl-2-Associated X Protein
  • hepatitis B virus X protein
  • Cyclin D1
  • CDC2 Protein Kinase
  • Mitogen-Activated Protein Kinase 8
  • Mitogen-Activated Protein Kinases
  • Tetracycline
  • Nocodazole