Fig. 4: Local re-organization of the actin cytoskeleton at the virus-cell interface.

a DiD-labeled IAVs were immobilized on reactive glass surfaces and A549 cells were cultivated on top. The cells were fixed, and the actin cytoskeleton was labeled with Alexa647-conjugated Phalloidin. The sample was imaged using TIRF to visualize actin structures close to the plasma membrane. We observed that some viruses show a distinct accumulation of the phalloidin signal as visualized by a line plot measurement (white line, upper right panel). b To further investigate the nanoscale organization of F-actin at the virus binding site, we performed STORM using the same samples as shown in (a). We identified a diverse F-actin organization with many filamentous structures as well as some smaller local accumulations. Closer inspection of the A647-phalloidin signal in the proximity of immobilized viruses revealed different structural patterns that we classify as arcs/rings, holes, or local amorphous accumulations (blobs) (b, right side). c We performed live-cell TIRF imaging of A549 cells expressing Lifeact-GFP and cultivated on top of immobilized DiD-labeled IAVs. While we observed a similar diverse structural organization of Lifeact-GFP, we could now follow its signal at the viral periphery over time (d). Images shown in (a–c) are representative of 2 experiments with similar results. d shows snapshots of a 5 min movie of the cell shown in (c) zoomed at the viral particle shown in the boxed inset. The four snapshots and the corresponding line profiles below show the structural transitions of the local Lifeact-GFP signal. Scale bars: A, 10 µm; B, 5 µm; B, zoom boxes, 500 nm; C, 10 µm; D, 1 µm. Source data are provided as a Source Data file.