title: Regulation of p21 and p27 expression by the hepatitis B virus X protein and the alternate initiation site X proteins, AUG2 and AUG3.
abstract: BACKGROUND
  The hepatitis B virus X gene has three in-frame start codons encoding the pX, AUG2 and AUG3 proteins. The AUG2 and AUG3 genes are 5'-truncated in respect to the full-length pX gene; however, all three genes terminate at the same stop codon. The activity of pX as an oncogene is well characterized; however, less is known about the AUG2 and AUG3 proteins.
  
  
  METHODS
  The effects of pX, AUG2 and AUG3 on p21Cip,1/WAF,1/MDA6 and p27Kip-1 cyclin kinase inhibitor (CKI) protein expression, and the impact they have on proliferation, were investigated in CHO K-1 cells. CHO K-1 cells were chosen because they can be transfected at 100% efficiency.
  
  
  RESULTS
  p21- and p27-luciferase reporter expression is modulated by increasing doses of the hepatitis B X proteins. At low concentrations of pX or AUG2, p21- and p27-luciferase activity was increased, and at high concentrations, p21- and p27-luciferase activity was decreased. Expression of the AUG3 gene showed a different profile: it was increasingly stimulatory with dose for both promoters. Western blot analyses demonstrated that p21 and p27 protein levels were modulated as predicted based on data generated in the promoter-luciferase experiments. Tritiated thymidine labeling of DNA showed biphasic kinetics of incorporation in the presence of varying pX and AUG2 concentrations, whereas labeling decreased with AUG3 concentration. The growth inhibitory effect of pX expression was reduced by antisense ablation of either p21 or p27.
  
  
  CONCLUSIONS
  The relative expression level of pX, AUG2, and AUG3 impacts on CKI expression and cell proliferation. Our findings may explain why divergent effects of pX expression on growth have been observed by different groups, which may be related to relative pX expression levels.
authors.affiliation: Departments of Radiation Oncology, Richmond Commonwealth University, Richmond, USA.
authors.forename: J Kevin
authors.initials: JK
authors.lastname: Leach
authors.forename: Liang
authors.initials: L
authors.lastname: Qiao
authors.forename: Youwen
authors.initials: Y
authors.lastname: Fang
authors.forename: Song L Y
authors.initials: SL
authors.lastname: Han
authors.forename: Dpnna
authors.initials: D
authors.lastname: Gilfor
authors.forename: Paul B
authors.initials: PB
authors.lastname: Fisher
authors.forename: Steven
authors.initials: S
authors.lastname: Grant
authors.forename: Philip B
authors.initials: PB
authors.lastname: Hylemon
authors.forename: Darrell
authors.initials: D
authors.lastname: Peterson
authors.forename: Paul
authors.initials: P
authors.lastname: Dent
mesh_terms: D000818:Animals; D016466:CHO Cells; D018797:Cell Cycle Proteins / Q000032:analysis* / Q000235:genetics*; D018387:Codon, Initiator / Q000235:genetics; D006224:Cricetinae; D050759:Cyclin-Dependent Kinase Inhibitor p21; D050760:Cyclin-Dependent Kinase Inhibitor p27; D016213:Cyclins / Q000032:analysis* / Q000235:genetics*; D004791:Enzyme Inhibitors / Q000032:analysis*; D015967:Gene Expression Regulation, Viral / Q000235:genetics*; D006509:Hepatitis B / Q000235:genetics*; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D066298:In Vitro Techniques; D011401:Promoter Regions, Genetic / Q000235:genetics; D015534:Trans-Activators / Q000235:genetics*; D024363:Transcription Initiation Site; D025521:Tumor Suppressor Proteins / Q000032:analysis* / Q000235:genetics*; D054334:Viral Regulatory and Accessory Proteins
chemical_list: C493956:CDKN1A protein, human; D018797:Cell Cycle Proteins; D018387:Codon, Initiator; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D004791:Enzyme Inhibitors; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D050760:Cyclin-Dependent Kinase Inhibitor p27