Expression of Recombinant GFP-Hsp90P E47A Gene by Lentiviral Vector Transfection and Its Effect on Cell Proliferation in HepG2 Cells

Objective: To construct the three-plasmid packaging cell line of the recombinant lentiviral vector encoding Hsp90αE47A/psin-GFP gene and investigate the molecular chaperon function of hsp90α. Methods: The three-plasmid recombinant lentiviral vector, which was made up of the vector plasmid (Hsp90αE47A/psin-GFP), the packaging plasmid (ΔNRF) and the envelop plasmid encoding the vesicular stomatitis virus-glycoprotein (VSV-G), was isolated and purified. Human embryonic kidney 293T cells were co-transfected with the three plasmids by calcium phosphate method. 48 hours after the transfection, the viral supernatant was col- lected to infect HepG2cells. The expression of reporter gene GFP was detected by fluorescence microscope, and the GFP-Hsp90αE47A protein expressed in Hepg2 cells was detected by Western blot. Cell proliferation was tested by CCK-8 method. Results: There was strong expression of GFP s in 293T and Hepg2 cells after transfection. The viral titer was 3.0 ×103 ifu /μl. The expression of GFP-Hsp90α in Hepg2 cells was confirmed by Western blot. Cell proliferation of GFP-Hsp90αE47A protein expressing Hepg2 cells was obviously inhibited at 4d. (1.051±0.03 vs 1.349±0.05, P0.05,t test). Conclusion: The three-plasmid packaging cell line system of recombinant lentiviral vector was successfully established, cell proliferation was obviously inbited by GFP-Hsp90αE47A gene transfec- tion, which will provide a basis for exploring the molecular chaperon function of Hsp90α.