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. 2021 Apr 16;12:2290. doi: 10.1038/s41467-021-22341-7

Fig. 2. Zika virus DVG-A relies on WT-encoded NS1 for its genome replication.

Fig. 2

a Schematic diagram of reporter constructs. For the wild-type replicon, C38 and E30 represent the N-terminal 38 aminoacids of C and the C-terminal 30 aminoacids of E proteins, respectively. For the DVG-A replicon, Pr38 and NS198 represent the N-terminal 38 aminoacids and the C-terminal 98 aminoacids of Pr and NS1 proteins, respectively. NanoLuc2A represents the NanoLuc reporter sequence followed by the foot-and-mouse disease virus 2A protease. An inactive replicon (with a mutated polymerase active site) was also constructed. b Replicon assays in infected or uninfected cells. Equal amounts of replicon or reporter RNA was transfected in naïve or infected Vero cells (MOI 1 PFU/cell), and relative light units (RLU) measured at the indicated times post-transfection. WT and inactive replicons were used as controls. ****p ≤ 0.0001. c DVG-A RNA reporter activity in HEK-293T cells pre-transfected with control DNA, E30-NS, E30-NSStopNS1 or E30-NSStopNS2A. WT replicon was used as a control. RLU were measured at the indicated times p.t. ****p ≤ 0.0001. d WT and DVG-A reporter assays in HEK-293T cells pre-transfected with control DNA or E30-NS1. RLU were measured at the indicated times p.t. ****p ≤ 0.0001. In b, c, and d, reporter activity was compared to that at 4 h by two-way ANOVA with Dunnet’s multiple comparison. e Cells transfected with a V5-tagged version of the E30-NS1 were harvested 24 h p.t. for western blotting, a representative blot of two is shown. Corresponding uncropped images of the western blot are shown in Supplementary Fig. 12. All graphs show the mean and SD; n = 3 per group for a representative experiment of three. Control DNA: GFP-expressing plasmid. Source data are provided as a Source Data file.