Fig. 4.

itEBLN is incorporated into viral RNPs and affects BDV polymerase activity. (A) Immunoprecipitation analysis of itEBLN in OL/BDV cells. OL cells persistently infected with BDV were transfected with the indicated constructs, lysed 24 h after transfection, and immunoprecipitated with anti-HA antibody. “Empty” indicates cells transfected with empty vector (pcDNA3). BDV P was detected by anti-BDV P antibody (arrow). The asterisk indicates nonspecific bands detected in all cells. BDV genomic RNA was detected by RT-PCR using primers specific for the genome sense RNA within the BDV P region. (B) Expression of itEBLN inhibits BDV polymerase activity in a minireplicon assay. HEK293T cells were cotransfected with a set of BDV minigenome plasmids and the expression constructs for itEBLN, hsEBLN-1, BDV N, and BDV X. Forty-eight hours after transfection, the cells were lysed and subjected to CAT assays. The CAT activities are expressed as the ratio relative to empty vector-cotransfected cells (Empty). The values are the mean ± SE of three independent experiments. Statistical significance was analyzed by the two-tailed t test; **P < 0.01.