Figure 2. Inhibition of PLA2G4A by Py-2 impairs infectivity of HCV.
(A) U0126 prevents phosphorylation of PLA2G4A in the absence of serum. Cells were cultured in presence or absence of serum and the U0126-assay was carried out as described in Figure 1A. Cell lysates were analyzed using antibodies specific for PLA2G4A or the S505-phosphorylated enzyme. (B) Luc-Jc1-transfected cells were treated with indicated doses of Py-2 as outlined in Figure 1A. The influence on RNA replication (left panel) and production of infectious particles (right panel) was determined as described in the legend to Figure 1. Data are shown as means +/− SD of three independent experiments (the dotted line represents background luciferase activity in mock infected cells). (C) Analysis of NS5A and core protein levels in Jc1-transfected cells in the presence or absence of Py-2. (D) Secreted and intracellular infectious HCV was quantified using a limiting dilution assay. (E) An iPLA2-specific inhibitor does not impede HCV RNA replication or virus production. Huh-7.5 cells were transfected with Luc-Jc1 and treated with given doses of BEL, an iPLA2-specific inhibitor, using the procedure outlined in Figure 1A. Luciferase activity was measured in transfected (left panel) and in the inoculated cells (right panel). Data are shown as means +/− SD of three independent experiments (the dotted line represents background luciferase activity in mock infected cells).