Pre-s1 Protein In Hepatitis B Virus Trans-regulatory Function Of Gene 2 Study

Hepatitis B virus (HBV) is the prototypic member of the hepadnavirus family, which is the main cause of acute and chronic liver diseases, including chronic active hepatitis, as well as cirrhosis and hepatocellular carcinoma. The molecular mechanism responsible for hepatocellular damage caused by HBV infection are strongly associated with the interaction between virus proteins and hapetocellular proteins. The investigation on these possible protein-protein interaction with each peptides of HBV can partly explain the mechanism of HBV infection, and may provide some new clues for the prevention and cure of Hepatitis B.JiDong constructed a subtractive cDNA library of genes transactivated by pre-S1 protein of hepatitis B virus (HBV) using suppression subtractive hybridization (SSH) technique and cloned genes associated with transactivation, found a new gene named human gene 2 transactivated by pre-S1 protein of hepatitis B virus (PS1TP2). We cloned the new gene PS1TP2, and explored its function and structure by bioinformatics analysis.To explore the possible function of PS1TP2, Green fluorescent protein (GFP) expression vector pEGFP-C1-PS1TP2 was established, which makes PS1TP2 fusing with green fluorescent protein in cells. The result suggests that PS1TP2 were subcellularly located in cell plasma. Advanced research was to screen proteins in white blood cell interacting with PS1TP2 protein using yeast two-hybrid technique. The obtained protein genes involve in cell immunity, signal transduction, energy metabolism and transcription regulation in vivo. The recombined expression plasmid pET32a(+)-PSlTP2 was constructed and transduced into E.coli strain Rosetta and induced to express by IPTG. In result, the recombinant plasmid was over-expressed in E.coli as inclusion body with molecular weight 41kD which could specifically react with His antibody by Western-blotting.To explore the possible function of PS1TP2, the recombined expression plasmid pcDNA3.1(-)-PS1TP2 was constructed, and HepG2 cells were transfected, and pcDNA3.1(-) vector was used as control. The mRNA was isolated from HepG2 cells transfected with pcDNA3.1(-)-PSlTP2 and pcDNA3.1(-) vector, respectively, and suppression subtractive hybridization (SSH) method was employed to analyze the differentially expressed DNA sequence between the two groups. The obtained product was subcloned into T/A plasmid vectors to set up the subtractive library. Amplification of the library was carried out with E. coli strain JM109. The cDNA was sequenced and analyzed in GenBank with BLAST search after PCR. The obtained sequences may be target genes transactivated by PS1TP2, among which some genes coding proteins involved in cell cycle regulation, signal transduction, correlated with tumor and metabolism. These may explain the possible mechanism of PS1TP2 in vivo.It is the first step to investigate the biological functions of PS1TP2, and pave a way for the following investigation of the more exact biological functions and the modulation of the gene expression.