Sac1 overexpression inhibits HCV replication.

<p>(A) Huh 7.5.1 cells transfected with pEGFP-Sac1 or mock and then cell lysates were analyzed by immunoblotting with the indicated antibodies. (B) Huh 7.5.1 cells transfected with pEGFP-Sac1 and infected with JFH1 were stained with antibodies against PI4P (red). (C) Huh 7.5.1 cells transfected with pEGFP-Sac1 and infected with JFH1 were stained with antibodies against HCV core (red). (D) Huh 7.5.1 cells transfected with pEGFP-Sac1 and infected with JFH1 were stained with antibodies against HCV NS5A (red). (E) Huh 7.5.1 cells were transfected with pEGFP-Sac1 and infected with JFH1. Total RNA was isolated and reversely transcribed and then quantitive PCR was performed. (F) Huh7.5.1 cells were transfected with pEGFP-Sac1 and infected with Jc1FLAG2(p7-nsGluc2A) for 3 days. Gaussia luciferase activity and cellular ATP levels were measured. The normalized luciferase activities were then divided by the normalized luciferase activity from mock treatment.</p

GBF1 is required for HCV replication.

<p>(A) Huh7.5.1 cells were treated with different dosages of BFA or GCA as indicated for 1 hour and then infected with JFH1 for 2 days. Total RNA was isolated and reverse transcribed and then quantitive PCR was performed. (B) Huh7.5.1 cells were treated with 20 µM GCA for 1 hour and then infected with JFH1 for 3 days. Cell lysates were analyzed by immunoblotting with the indicated antibodies. (C) Huh7.5.1 cells were treated with siRNA against GBF1 or control siRNA for 3 days and then infected with JFH1 for 3 days. Cell lysates were analyzed by immunoblotting with the indicated antibodies. (D) Huh7.5.1 cells were treated with siRNA against GBF1 or control siRNA for 3 days and then infected with JFH1 for 3 days. Total RNA was isolated and reverse transcribed and then quantitive PCR was performed.</p

PI4P is rearranged during HCV infection.

<p>(A) Huh 7.5.1 cells infected or uninfected with JFH1 were stained with antibodies against PI4P (red) or β-COP (green). (B) Huh 7.5.1 cells infected with JFH1 were treated with 90 IU/ml IFN or mock infection and then stained with antibodies against PI4P (red) or β-COP (green). (C) Huh 7.5.1 cells infected or uninfected with JFH1 were stained with antibodies against PI4P (red) or NS-5A (green).</p

Primers used for quantitative RT-PCR.

a<p>F, forward; R, reverse.</p

ARF1 is required for HCV replication.

<p>(A) Huh7.5.1 cells were treated with siRNA against PI4KIIIβ or control siRNA for 3 days and then infected with JFH1 for 3 days. Cell lysates were analyzed by immunoblotting with the indicated antibodies. (B) Huh7.5.1 cells were treated with siRNA against ARF1 or control siRNA for 3 days and then infected with JFH1 for 3 days. Total RNA was isolated and reverse transcribed and then quantitive PCR was performed. (C) Huh7.5.1 cells were treated with siRNA against ARF1(#6 sequence or #8 sequence) or control siRNA and then infected with Jc1FLAG2(p7-nsGluc2A) for 3 days. Gaussia luciferase activity and cellular ATP levels were measured. The normalized luciferase activities were then divided by the normalized luciferase activity from mock treatment.</p

PI4KIIIβ colocalizes with ARF1 or GBF1.

<p>(A) Huh 7.5.1 cells transfected with a HA-tagged ARF1 construct and infected with JFH1 or mock were stained with antibodies against HA (red) or PI4KIIIβ (green). (B) Huh 7.5.1 cells infected with JFH1 or mock were stained with antibodies against PI4KIIIβ (red) or GBF1 (green).</p

PI4KIIIβ is required for HCV replication.

<p>(A) Huh7.5.1 cells were treated with siRNA against PI4KIIIβ or control siRNA for 3 days and then infected with JFH1 for 3 days. Total RNA was isolated and reversely transcribed and then quantitive PCR was performed. (B) Huh7.5.1 cells were treated with siRNA against PI4KIIIβ or control siRNA for 3 days and then infected with Jc1 for 3 days. Gaussia luciferase activity and cellular ATP levels were measured. The normalized luciferase activities were then divided by the normalized luciferase activity from mock treatment. (C) Huh7.5.1 cells were treated with different dosages of PIK93 as indicated for 1 hour and then infected with JFH1 for 3 days. Total RNA was isolated and reversely transcribed and then quantitive PCR was performed. (D) Huh7.5.1 cells were treated with different dosages of PIK93 as indicated for 1 hour and then infected with Jc1FLAG2(p7-nsGluc2A) for 3 days. Gaussia luciferase activity and cellular ATP levels were measured. The normalized luciferase activities were then divided by the normalized luciferase activity from mock treatment.</p