FIG 7 .

AUF1 inhibits poliovirus and CVB3 IRES-driven translation during infection. (A) Schematic of luciferase reporter RNAs. Viral IRES-driven translation was measured using in vitro-transcribed RNA encoding the poliovirus or CVB3 5′ noncoding region (5′ NCR) upstream of firefly luciferase (FLuc). Cap-dependent translation was measured using a Renilla luciferase (RLuc) construct in vitro transcribed in the presence of m⁷G(5′)ppp(5′)G cap analog. (B) Viral IRES- and cap-dependent translation measured in uninfected cells 8 h after cotransfection of 293-shCtrl or -shAUF1 cells. FLuc and RLuc activities were measured using a dual-luciferase assay and represented as relative light units per second (RLU/s) normalized to cell count. The means from three individual experiments ± SEM are represented. (C) Schematic of infection followed by cotransfection experiment. 293-shCtrl or -shAUF1 cells were infected with poliovirus (MOI of 1) or CVB3 (MOI of 20). Immediately following virus adsorption, cells were cotransfected with the 5′ NCR-FLuc and capped RLuc RNAs. At 4 h and 8 h posttransfection, cells were lysed and dual luciferase activity was measured. (D) 293-shCtrl or -shAUF1 cells were infected with poliovirus (MOI of 1) prior to cotransfection with luciferase reporter RNAs. FLuc and RLuc activities were measured at 4 h and 8 h posttransfection and represented as relative light units per second (RLU/s) normalized to cell count. Data from cotransfections were graphed separately due to differences in scale and represent the means from three individual experiments ± SEM. The ratios of IRES- to cap-dependent translation from these experiments are also presented. (E) 293-shCtrl or -shAUF1 cells were infected with CVB3 (MOI of 20) prior to cotransfection with luciferase reporter RNAs. Results are presented the same as in panel D. *, P < 0.05; ***, P < 0.001.