MAP-Kinase Regulated Cytosolic Phospholipase A2 Activity Is Essential for Production of Infectious Hepatitis C Virus Particles
Figure 7
HCV particles produced in the presence of Py-2 display aberrant protein composition.
(A) Virus particles produced in the presence or absence of Py-2 were separated by using ultracentrifugation and an iodixanol step gradient. Ten fractions were collected from the bottom, and core protein abundance (left panel) and infectivity titers (right panel) were determined. One example of two independent experiments is given. (B) FLAG-Jc1 RNA [13] was transfected into Huh-7.5-HA-ApoE cells (Figure S11) and cells were treated with Py-2 as outlined in Figure 1A. The total amount of ApoE secreted from transfected cells was determined using an ELISA (top panel). Virus containing culture fluid of Py-2 and untreated cells were normalized for equal quantities of ApoE and precipitated with ApoE-specific antibodies. The amount of co-precipitated core protein was determined by ELISA and is expressed relative to the untreated control. Data are shown as means +/− SD of three independent experiments (C) FLAG-Jc1 transfected Huh-7.5-HA-ApoE cells were treated as in (B). The amount of secreted HCV core protein was determined by ELISA (left panel) and normalized to equal amounts of core prior to precipitation with FLAG-specific antibodies. The amount of co-precipitated ApoE and core was determined by Western blot and ELISA, respectively and is expressed relative to the untreated control. Data are shown as means +/− SD of three independent experiments.