West Nile Virus Replication Requires Fatty Acid Synthesis but Is Independent on Phosphatidylinositol-4-Phosphate Lipids
Figure 2
Replication of WNV is dependent on fatty acid synthesis.
(A) WNV infection requires active fatty acid synthesis. Cells infected with WNV (MOI of 0.5 PFU/cell) were treated with 15 µM cerulenin or 15 µM C75 from 0 or 3 h p.i. throughout the rest of the assay and total virus yield was determined at 24 h p.i. (B) Genome replication of WNV is dependent on fatty acid synthesis. Cells were infected and treated with FASN inhibitors from 3 h p.i. as in (A). RNA was extracted at 24 h p.i. and the number of WNV RNA copies was determined by quantitative RT-PCR. (C) Localization of FASN in mock and WNV-infected Huh-7 cells. Infected cells (MOI of 5 PFU/cell) were fixed and processed for immunofluorescence (12 or 24 h p.i.) using a rabbit anti-FASN antibody in combination with a monoclonal antibody against dsRNA. Primary antibodies were detected using suitable AF-488 or 594 labelled secondary antibodies. Scale bar: 10 µm. (D) Analysis of FASN levels during WNV infection. Huh-7 cells were infected with WNV as in (A) and lysed at different times p.i. Western blot analysis was performed to determine the relative levels of FASN protein. Membrane was retested against a tubulin antibody as a control for protein loading.
