FIG. 6.

CBP and p300 modulate the HDV replication. (A) CBP and p300 enhance SHDAg expression. For each experiment, approximately 2 × 10⁶ HuH-7 cells were transfected with 10 μg of empty vector (−) or expression construct of FLAG-tagged CBP or Myc-tagged p300 by Lipofectamine 2000 transfection reagent (Invitrogen). At 12 h posttransfection, cells were then transfected with HDV genomic RNA (15 μg) and SHDAg mRNA (5 μg) by DMRIE-C transfection reagent (Invitrogen). The expression levels of CBP, p300 (1 day posttransfection), and HDAg (4 days posttransfection) were analyzed by SDS-PAGE followed by immunoblotting with anti-FLAG tag, anti-Myc tag, or anti-HDAg antibodies. (B) CBP and p300 enhance the replication of HDV RNA. The expression levels of HDV genomic (G) and antigenomic (AG) RNA in the transfected cells as described in panel A were determined by Northern blotting at 4 days posttransfection. (C) Acetyltransferase-defective mutant of p300 inhibits HDV replication. This experiment is similar to that shown in panel A, except that 10 μg of empty vector (−) or expression construct of Myc-tagged p300 (p300) or its mutant D1399Y (D1399Y) was used for the transfection experiment. At 12 h posttransfection, cells were then transfected with HDV genomic RNA (10 μg) and SHDAg mRNA (10 μg) by DMRIE-C transfection reagent (Invitrogen). The expression levels of p300 and HDAg (1 day posttransfection) were analyzed by SDS-PAGE followed by immunoblotting with anti-Myc tag or anti-HDAg antibodies at 24 h posttransfection. (D) The expression levels of HDV RNAs and SHDAg in the transfected cells as described in panel C were determined by Northern blotting and Western blotting at 4 days posttransfection. Mock, HuH-7 cells without transfection. P, SHDAg or HDV RNAs as positive controls; WB, Western blotting; α, anti.