Figure 5.

Impaired autophagic flux in HCV infection is due to an Arl8b-dependent mechanism. A, Huh-7.5 cells stably expressing nontarget shRNA (Scram) or an shRNA to Arl8b assessed by Western blotting. B, cells were infected with JFH-1 and treated with 100 nm bafilomycin A1 (BafA) for 4 h as in Fig. 1. C, densitometry analysis of the changes in LC3 levels after bafilomycin treatment (n = 3). D, nontarget (Scram) or shArl8b Huh-7.5 cells were transfected with RFP–GFP–LC3 plasmid, infected with JFH-1, and treated with EBSS. Fusion was assessed as stated previously. Scale bars represent 10 μm. E, Huh-7.5 cells stably expressing nontarget shRNA (Scram) or shArl8b were infected with the JC1-Luc strain of HCV. Viral replication was assessed in total lysates 3 days postinfection using the Dual-Glo luciferase assay system (n = 4). F, the secretion of infectious virus is decreased when Arl8b is silenced. The cells were infected as in E, and 3 days postinfection, the presence of infectious virus in the culture medium was measured by inoculating virus naïve cells as described under “Experimental procedures” (n = 4). Cont, control.