FIG 9.

The YV motif of claudin-1 is critical for determining the cell surface level of claudin-1 in and HCV entry. (A) Huh7.5 cells were transfected with wild-type Flag–claudin-1 or Flag–claudin-1 with a mutation in the C terminus (²¹⁰YV²¹¹/²¹⁰AA²¹¹) and fixed by PFA, followed by staining with Flag antibody. DAPI staining indicates the nucleus (blue). Scale bar, 15 μm. (B) Immunoblots showing the protein expression of wild-type or mutant siRNA-resistant claudin-1. Endo, endogenous. (C and D) Huh7.5.1 cells were treated with siRNA targeting the claudin-1 5′ UTR or a control siRNA for 24 h and transfected with a construct expressing wild-type or mutant siRNA-resistant claudin-1 for 48 h. Then the cells were infected with genotype 1b HCVpp (Con1) for another 72 h (C) or infected with Jc1 for another 24 h (D). The infection activity and ATP levels were measured. The relative luciferase activity was normalized to the cellular ATP levels. Data are presented as the means ± SD. ***, P < 0.001; NS, not significant.