Image analysis

For confocal images, the viral M clusters area for different conditions were measured using segmentation (ImageJ/Fiji). A schematic representation for details of the segmentation methods is given in Figure S1. Viral particles size from STED images were also analysed using segmentation (ImageJ). The mean fluorescence intensity of F-actin, alpha-Actinins and ER-GRP78 labelling were measured using ImageJ. For Mean intensity analysis, images were acquired using a confocal microscope. To ensure reasonable quantification among conditions, all images were taken with the same objective and microscope settings. For post-processing, individual cells, which are well separated from each other were chosen. Z-stacks with 0.3 micrometer slices per cell were chosen to ensure the whole cell slicing in focus. The resulting Z-projection images of individual cells were used to calculated the mean fluorescence intensity of each cell for each labelling. In addition, we calculated the cell area, volume and height using 3D viewer plugin from ImageJ. From STED and Airy scan images the diameter sizes of F-actin, Rab7, Lamp1 and viral rings were determined “manually” for each ring using ImageJ software. Mander’s colocalization analysis done by using JACoP plugin of Image J software.