Inhibition of Translation Initiation by Protein 169: A Vaccinia Virus Strategy to Suppress Innate and Adaptive Immunity and Alter Virus Virulence
Fig 10
Protein 169 inhibits CrPV IRES-dependent translation.
(A, B) HEK 293T cells were transfected with a plasmid encoding a bicistronic RNA expressing renilla luciferase (RLuc, A) by a cap-dependent translation and firefly luciferase (FLuc, B) by a CrPV IRES-dependent translation, together with plasmids for expression of A49.TAP, 169 or EV control in quintuplicate. After 7 h the cells were treated with CHX (1 μg/ml) for 16 h. The relative amount of RLuc and FLuc was determined by luminescence and results are presented as the fold increase in luciferase expression normalized to the EV control ± SD. Data shown are from one representative experiment (n = 5). Statistical analysis was performed using a two-tailed Student’s t-test with Welch’s correction where necessary, *** p < 0.01, **** p < 0.001. (C) Cell lysates from (A) were resolved by SDS-PAGE followed by immunoblotting with the indicated antibodies.