VACV infection increases the levels of H3K9me3.

(A) BSC-40 cells were grown on coverslips and infected at a MOI 5.0 with VACV strain WR. At various time points post infection, the coverslips were fixed and stained to detect the VACV I3 protein and H3K9me3. DNA was counterstained with DAPI. Images were acquired using an Olympus IX-71 inverted microscope at 60x magnification and deconvolved using Softworx software (GE Healthcare) (scale bar = 25 μm). The nuclear H3K9me3 signal intensities from (B) microscopy images and (C) flow cytometry were quantified using FIJI imaging analysis software and normalized to mock-infected cells. At least five images were analyzed per samples within an independent experiment. Data represent the standard error of the mean (SEM) of three independent experiments. GraphPad was used to determine significant differences in H3K9me3 levels following VACV infection. Statistically significant differences are noted, relative to time zero (*P<0.05; **P<0.01).