Fig. 4.

G3BP granules form concomitantly with viral RNA replication and NSP accumulation. Vero (replication non-permissive) or C-Vero (replication-permissive) cells were transfected with RUBrep/P150-GFP in vitro RNA transcripts and analysed for the percentage of GFP-positive (i.e. successfully transfected) cells that contained G3BP granules by immunofluorescence using antibodies against G3BP. (a) Western blotting of Vero and C-Vero cell lysates for FLAG-tagged CP (approx. 30 kDa) is shown at top, whilst below are micrographs showing Vero or C-Vero cells at 48 h post-transfection with RUBrep/P150-GFP transcripts and stained for dsRNA (red) (GFP-tagged P150 is green in these micrographs and nuclei were counterstained with Hoechst 33342). (b) Quantification by ImageQuant of P150 levels on Western blots from 48 h post-transfection lysates (Vero or C-Vero) using GFP antibodies (mean of two independent experiments). (c) Quantification of GFP-positive cells containing G3BP granules. At least 100 cells from 15 different fields of view were analysed from at least two different experiments. Error bars represent sd.