Extended Data Fig. 3: Tomographic analysis of SARS-CoV-2 infected A549ACE2+TMPRSS2 cells (10 h p.i.).
From: A coronavirus assembly inhibitor that targets the viral membrane protein
a, Example electron micrograph of a high-pressure frozen and freeze substituted DMSO-treated infected cell. Yellow arrowheads indicate assembling virions and black stars (*) indicate DMVs. b,c, Detail of the regions boxed in panel a, showing a strong contrast of the assembling viral ribonucleoprotein at progressive stages of assembly. The tomographic slice in panel b corresponds to a diagonally resliced tomographic volume capturing a view of three adjacent assembly sites. Three-dimensional segmentations of the progressively assembling virions are shown on the right of the tomographic sections. Light blue volumes represent DMVs, light green the ERGIC/Golgi membranes containing the virions. The virions’ color coding indicates stages of budding: early (yellow), intermediate (orange) and complete (red). Viral RNPs are represented in purple. d,e, Further examples of virions in very early and late stages, respectively taken from the tomographic volume in a at a z height not visible in the overview. f, Tomographic slice through an infected and CIM-834 (1 µM)-treated cell showing complete absence of virions and assembly events, despite a high abundance of DMVs of similar size and distribution as observed in DMSO-treated cells. Darkened, elongated mitochondria (m) are present in the area surrounding the DMVs. g, Segmentation of the volume displayed in f, showing the spatial relationship of DMVs (light blue) and mitochondria (pink), confirming the absence of assembly sites throughout the volume. h, i Magnified tomographic sections of the boxed regions in panel f. Tomographic section thickness: 0.78 nm (a,f), average of 5 × 0.78 nm thick slices (b,c,h,i).
