FIG 1.

Ubiquitination is important for HCV replication. (A) HCV replicon (9-13) cells were treated with PR-619, a nonselective DUB inhibitor, or dimethyl sulfoxide (DMSO) for 24 h, and cell viability and intracellular HCV RNA levels were determined by PI staining and qRT-PCR, respectively. (B) Expression of FLAG-tagged OTUD7B, OTUB1, and OTUD1 in Huh7 cells was detected by Western blotting using anti-FLAG antibody. (C) Subcellular localization of OTUD7B, OTUB1, or OTUD1 overexpression in Huh7 cells was observed by confocal microscopy. Each DUB (green) or nucleus (blue) was stained with anti-FLAG antibody and DAPI, respectively. (D) Huh7 cells expressing the indicated DUBs were infected with HCV at an MOI of 3. After 4 days postinfection, the culture supernatants were collected, and infectious HCV titers in the culture supernatants were determined by a focus-forming assay. WT, wild type; FFU, focus-forming units. (E) HCV was inoculated into Huh7 cells expressing the indicated DUBs at an MOI of 3 and stained with anti-NS5A antibody at 4 days postinfection. *, P < 0.05; **, P < 0.01.