The Ubiquitin-Specific Protease 18 Promotes Hepatitis C Virus Production by Increasing Viral Infectivity

Figure 1

Human USP18 expression and catalytic activity in Huh7.5 cells. (a) Huh7.5 cells were seeded at , 2 ml per well in 6-well plates in antibiotic-free medium for 24 hours before 4 μg empty vector pcDNA-DEST53, 2 μg USP18 WT, or 4 μg USP18 WT was transfected into each well. 48 hours posttransfection, total protein was extracted to detect GFP tag by western blot. (b) Cleavage of ISG15-GST fusion in vitro. Huh7.5 cells were transfected with various USP18 plasmids (wild-type USP18, WT; USP18 C64S, C64S; USP18 C65S, C65S; or USP18 C64/65S, C64/65S) in combination with ISG15-GST. 48 hours posttransfection, total protein was extracted to detect ISG15 and USP18 by western blot. (c) Cleavage of ISG15 conjugates in IFNα-treated Huh7.5 cells. Huh7.5 cells were transfected with an empty vector (vector), USP18 WT, or USP18 C64S. 24 hrs later, the cells were treated with IFNα (0-100 U/ml) for 24 hours, after which western bot was performed to analyze expressions of ISG15 conjugates and USP18. Untreated: untreated control; mock: transfected with 4 μg empty vector pcDNA-DEST53; pcDNA-DEST53-USP18: transfected with wild-type USP18 (USP18 WT).

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