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. 2021 May 6;12:641059. doi: 10.3389/fmicb.2021.641059

FIGURE 3.

FIGURE 3

R31E mutation significantly impairs NS1 induced ER remodeling and attenuates viral replication. (A) ER morphology with the expression of WT and R31E mutant NS1. Wild type NS1 or R31E mutant with C-terminal Myc-tag was expressed in HeLa cells and then labeled with anti-Myc and anti-Calreticulin antibodies. WT NS1 expression induced apparent ER remodeling, whereas R31E mutant could rarely remodel the ER. Scale bar, 20 μm. (B) The percentage of ER-remodeled cells induced by expression of WT NS1 or R31E mutant. The ratio of ER-remodeled cells to total NS1 positive cells is defined as the percentage of ER-remodeled cells. (C) NS1 R31E mutation impairs viral replication. NS1 R31E mutation was introduced into the ZIKV replicon. Luciferase activity was measured at 10 h and 36 h post-transfection to validate the viral replication ability. (D) NS1 R31E mutation significantly impairs the production of infectious viruses. NS1 R31E mutation was introduced into ZIKV infectious clone. At about 72 h post-transfection, the culture medium was collected, and viral RNA was detected by RT-qPCR. (E) Plaque assay was performed using the recombinant ZIKV to infect Vero cells. The data are presented as the mean ± SEM. The p-values were obtained from a two-tailed t-test. ***P < 0.001.