Establishment of cell lines using a doxycycline-inducible gene expression system to regulate expression of hepatitis B virus X protein.
The hepatitis B virus (HBV) X gene plays an important role in HBV-associated pathogenesis, especially hepatocarcinogenesis. Establishment of a stable and regulable HBx expression system will allow study of the function of this gene. Here, we describe the development of a doxycycline-inducible recombinant plasmid (pBPSTR3-FlagX) with the full-length HBV X gene and all components of the tetracycline-on ("Tet-on") gene expression system. This vector exhibited dose-dependent doxycycline-dependent induction of the Flag-HBx protein in HepG2 and Hep3B cells. We also observed dose-dependent doxycycline transactivation of HBx in HepG2 cells. After transfecting HepG2 cells with the pBPSTR3-FlagX plasmid, we isolated five puromycin-resistant cell clones with stable HBx expression, two of which exhibited stable and tight control of HBx expression by doxycycline. This new system has great potential for functional studies of the HBV X gene.
Tang
Hong
H
Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, People's Republic of China.
Liu
Li
L
Liu
Feng-Jun
FJ
Chen
En-Qiang
EQ
Murakami
Seishi
S
Lin
Yong
Y
He
Fang
F
Zhou
Tao-You
TY
Huang
Fei-Jun
FJ
D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D045744:Cell Line, Tumor; D004318:Doxycycline / Q000494:pharmacology*; D015967:Gene Expression Regulation, Viral / Q000187:drug effects*; D017930:Genes, Reporter; D006515:Hepatitis B virus / Q000235:genetics*; D006801:Humans; D008113:Liver Neoplasms; D010957:Plasmids; D015183:Restriction Mapping; D015534:Trans-Activators / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection; D054334:Viral Regulatory and Accessory Proteins
D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; C057628:hepatitis B virus X protein; D004318:Doxycycline