Figure 1. Viperin inhibits translation of Zika virus RNA.

(A) Immunoblot analysis of viperin induction in a doxycycline-inducible 293T variant (293T.iVip). Cells were treated with doxycycline (Dox) for 24 h. Cell lysates were analyzed by immunoblotting with anti-viperin and anti-GRP94 antibodies.

(B)Quantification of Zika virus (ZIKV) replication in 293T.iVip cells. Cells were treated with Dox for 24 h and then infected with ZIKV at an MOI of 1 for 24 h, and viral titers were determined by plaque assay. PFU, plaque-forming unit.

(C) Immunoblot analysis of ZIKV envelope (E) protein in 293T.iVip cells. Cells were treated with Dox for 24 h and then infected with ZIKV at an MOI of 1 for 24 h. Cell lysates were analyzed by immunoblotting with anti-flavivirus envelope (E) protein, antiviperin and anti-GAPDH antibodies. Quantification of ZIKV E protein from immunoblot analysis was shown on the right.

(D) qRT-PCR analysis of ZIKV viral RNA in 293T.iVip cells. Cells were treated with Dox and then infected with ZIKV at an MOI of 1 for 24 h. Viral RNA was determined by qRT-PCR targeting the 5’ and 3’ ends or in the middle of the viral RNA and normalized to GAPDH.

(E) Translation analysis of ZIKV RNA in 293T.iVip cells. Cells were treated with Dox for 24 h and then infected with ZIKV at an MOI of 1 for 24 h. Cell lysates were fractionated by sucrose density gradient velocity sedimentation. Viral RNA in sucrose density gradient fractions was determined by qRT-PCR.

(F) Quantification of ZIKV RNA translation in (E). Data are represented as the percentages of ZIKV RNA in monosome and polysome pools.

For (B-D) and (F), data are shown as mean ± SD of three biological repeats (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001 by unpaired Student’s t test. See also Figures S1 and S2.