Reorganization of microtubules, centrosome, and actin stress fibers during infection imaged using a custom-built SIM microscope.A, HFF cells were infected with eYFP-ICP0/gC-mCherry HSV-1, stained with SiR-tubulin, and imaged live. At late stages in infection, microtubules are bundled and excluded from the juxtanuclear region as shown by plotting the normalized intensity profiles from the nucleus to the outer cell boundary for each stage of infection. Bottom row shows enlarged view of the juxtanuclear region as marked with red boxes in the merged images. Arrows highlight the colocalization of ICP0 with the microtubule organizing center (MTOC) during the early phase in replication. B, HFF cells were infected with eYFP-ICP0/gC-mCherry HSV-1, fixed at 3.5, 5.5, 7.5, and 9.5 hpi, and the centrosome labeled with an antibody for pericentrin (indicated by red boxes in the merged images). During stage 1 and 2, colocalization of ICP0 with the centrosome can be detected. C, HFF cells were infected with eYFP-ICP0/gC-mCherry HSV-1, stained with SiR-actin, and imaged live. For classification purposes, three-color images of ICP0, gC, and SiR-actin were taken at a middle section of each cell. Shown are the merged images of all three channels (left). Actin stress fibers are remodeled at the basal side of respective cells. Bottom row shows enlarged view of actin stress fibers (areas marked with red boxes in upper row). Scale bars 10 μm.