Heat shock protein 90ß in the Vero cell membrane binds Japanese encephalitis virus

Wang, Yuan; Li, Yan; Ding, Tianbing

Several siRNA fragments were synthesised based on the human HSP90β mRNA transcript sequence from the NCBI PDB database. Following the general principles of siRNA design (30) , several siRNA fragments were synthesised, subcloned into lentiviral vectors (Cat. no. HPK-LvTR-20; GeneCopoeia, Rockville, MD, USA), packaged in the 293Ta lentiviral packaging cell line (Cat. no. Clv-PK-01; GeneCopoeia), and the pseudoviruses thus produced infected intact Vero cells in the presence of 5 µg/ml hexadimethrine bromide. We considered HSP90β expression inhibition over 50% effective as void of off-target effects, and the fragment was selected for further experiments. Cells with reduced HSP90β expression were then subjected to JEV infection (MOI=0.1), and progeny virus yields were measured using plaque formation assay at 12, 24, 36 and 48 hours post-infection (hpi).