Figure 2

The minimal RSE protected the gag stop codon from recognition by the NMD machinery. Co-transfection of wildtype, ΔRSE, 2577-2732 (minimal RSE) and 2788-2732 with wildtype Upf1 or a dominant negative form of Upf1 (RR857GA). Each construct was transiently transfected into CEFs, and RNA steady-state levels were assayed 48 hours later by RNase protection assay. A representative RNase protection assay is shown, with the set of bands below each bar corresponding to the construct indicated directly above on the graph. The top band of the gel (experimental) is a fragment of gag probe protected during the RNase protection assay corresponding to the unspliced viral RNA from the experimental construct. The bottom band (control) is a wild-type viral loading control that protects a different sized fragment of the same gag probe due to a small deletion. Standard deviations are represented on the bar graph. Values represent the average of at least four experiments. A star indicates a significant reduction compared to the corresponding viral construct co-transfected with wild-type Upf1 (*: p < 0.01).