Figure 8

Analysis of viral protein degradation in infected cells. (A-D) Untreated or (E, F) Bafilomycin A1-preincubated HeLa cells were incubated for 30 minutes at ~10°C with double-tagged viruses composed of Gag-eGFP labeled capsids and (A) PE Ch, (B) PE Ch iCS, (C, E) SE Ch, (D, F) SE Ch iCS. Subsequently, cells were either lysed immediately (0 h pb) or warmed to 37°C to allow virus entry for the time periods indicated (1–24 h pb). At the respective time points, the cells were lysed, and the cell lysates were analyzed by immunoblotting. The capsid proteins were detected using a PFV Gag-specific antibody (α-Gag) and the LP-Ch protein using a mCherry-specific antibody (α-Ch). Abbreviations: h pb, hours post binding; GAPDH, Glyceraldehyde 3-phosphate dehydrogenase