Fig. 3

Viral inclusions quickly respond to changes in the cellular environment. A549 cells were transfected with a plasmid encoding GFP-NP and infected with PR8 (a) or infected with PA-GFP virus (b) at an MOI of 5. At 10–16 hpi, cells were imaged under time-lapse conditions. The black arrowhead indicates addition of 80% water (hypotonic shock), 5% hexanediol or regular growth medium. White boxes highlight  viral inclusions in the cytoplasm in the individual frames. The dashed white line marks the cell nucleus, whereas the dashed yellow line delineates the cell periphery. Bar = 10 μm. Images were extracted from Supplementary Movies 10 to 15. Experiments were performed at least twice. c A549 cells were infected with PR8, with PA-GFP virus, or transfected with a plasmid encoding GFP-NP and infected with PR8 as above. At 16 hpi, cells were fixed and stained for NP. Percentage of cells with undissolved viral inclusions under the indicated treatments was calculated and plotted. Statistical analysis of data was performed using a non-parametric Kruskal–Wallis test, followed by Dunn’s multiple comparisons test (**p < 0.05; ***p < 0.001). At least 10 cells were analyzed per individual dot and 10 panels per condition