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. Author manuscript; available in PMC: 2023 May 5.
Published in final edited form as: Mol Cell. 2022 Mar 21;82(9):1631–1642.e6. doi: 10.1016/j.molcel.2022.02.031

Figure 7. GCN2 and ZAK are required for the inhibition of viral protein synthesis and viral replication by viperin.

Figure 7

(A) 293T.iVip cells were treated with Dox and infected with West Nile virus Kunjin strain (WNVKUNV) at an MOI of 1. Viral titers were determined by plaque assay. PFU, plaque-forming unit. hpi, hours post-infection. ND, not detected.

(B)Immunoblot analysis of WNVKUNV E proteins in 293T.iVip cells. 293T.iVip cells were treated with Dox for 24 h, infected with WNVKUNV at an MOI of 1. Cell lysates were analyzed by immunoblotting with anti-flavivirus envelope (E) protein, anti-viperin and anti-GRP94 antibodies. hpi, hours post-infection.

(C) 293T.iVip cells were treated with Dox and then infected with WNVKUNV at an MOI of 1 for 24 h. Cells were harvested for RNA extraction. Viral RNA was determined by qRT-PCR and normalized to GAPDH.

(D-E) NT, GCN2 KO and ZAK KO cells were treated with ddhC for 24 h and infected with WNVKUNV at an MOI of 1 for 24 h. Viral titers were determined by plaque assay (D). PFU, plaque-forming unit. Cell lysates were analyzed by immunoblotting with anti-anti-flavivirus E protein and anti-GAPDH antibodies.

For (A), (C) and (D), data are shown as mean ± SD of three biological repeats (n = 3). ***p < 0.001 by unpaired Student’s t test. ns, not significant.