CytoTrap Cytoplasmic Yeast Two-hybrid Screening For The Proteins Interacting With The Hepatitis B Spliced Protein

Spliced variants of hepatitis B virus are a group of subgenomic DNA generated from 3.5 kb pregenomic RNA by splicing and reverse transcription. More than 80% of these spliced variants are 2.2 kb in length. 2447 nt-489 nt HBV spliced variant, encoding Hepatitis B spliced protein (HBSP), was shown to correlate with chronization of HBV infection, liver fibrosis and lamivudine-resistance. However, the biological significance of this type of defective genome is far from being fully elucidated.The CytoTrap two-hybrid system is based upon generating fusion proteins whose interaction in the yeast cytoplasm activates the Ras-signaling pathway, inducing the cdc25 mutated cdc25H yeast strain to grow at 37°C. These properties of the CytoTrap system enable researchers to study protein interactions that cannot be assayed by using conventional two-hybrid. These include proteins that are transcriptional activators or repressors, proteins that require post-translational modification in the cytoplasm, and proteins that are toxic to yeast in conventional two-hybrid systems.In the present study, HBSP-binding cellular proteins were searched by using a CytoTrap two-hybrid system and identified the transforming growth factor beta 1 induced transcript 1 (TGFβ1I1) as a specific binding partner. it was postulated that HBSP may contribute to invasion capacity, apoptosis of hepatocytes and androgen-inducing tumorgenesis by interfering with TGFβ1I1.