FIG 3.

ISG20 expression does not accelerate degradation of CHIKV RNA. (A) MEFs were infected with CHIKV expressing nLuc in frame with the nsP3 nonstructural protein at an MOI of 5 and then treated with 0 or 10 μM cycloheximide (Cyc) to block translation and, ultimately, replication of CHIKV RNA. (B) MEFs were infected with CHIKV-LR (MOI = 5) and then treated with 10 μM cycloheximide. RNA was collected at the indicated time p.i. and analyzed by qRT-PCR for the CHIKV 3′ terminus. (B) Data represent results of 2-way ANOVA with Holm-Sidak post hoc analysis against eGFP control. (n = 6, 2 independent experiments; data were not statistically significant). (C) RNA isolated from ISG20 MEFs is functional. The CHIKV-translation reporter was isolated from tet-off MEFs overexpressing GFP, ISG20, or ExoII mutant, retransfected into MEFs expressing either GFP or ISG20 for 3 h, and assayed for firefly luciferase reporter activity, and luciferase activity data from second transfections are given as ratios of firefly luciferase signals from h 3 versus h 0 samples taken from first transfection. Data represent results of ANOVA with Dunnett’s post hoc analysis against GFP control (n = 6, 2 independent experiments; data were not statistically significant).