Fig. 1.

ESCRT-III is recruited to the nuclear rim in HSV-1-infected cells. a HeLa/CHMP4B-EGFP cells mock-infected or infected with HSV-1 for 22 h were analyzed by confocal microscopy for CHMP4B-EGFP and lamin A/C. Bars, 20 μm. Images are representative of three independent experiments. b Colocalization between CHMP4B-EGFP and lamin A/C in the experiment in (a) was quantified using Mander’s colocalization coefficient. Data are shown as the mean ± SEM (n = 16 for mock-infected and 15 for HSV-1-infected cells representative of three independent experiments). c HeLa or HeLa/CHMP4B KO cells were mock-infected or infected with HSV-1 for 22 h and analyzed by confocal microscopy for CHMP4B and lamin A/C. Bars, 20 μm. Images are representative of three independent experiments. d HeLa/CHMP4B-EGFP cells infected with HSV-1 for 22 h were analyzed by N-SIM super-resolution microscopy for (left) CHMP4B-EGFP and calnexin or (right) CHMP4B-EGFP and lamin A/C. Bars, 1 μm. Fluorescence line scans along the dotted lines of N-SIM images are shown under each image. Images are representative of three independent experiments. e HeLa/CHMP4B-EGFP cells were infected with HSV-1 for 24 h and analyzed by immunoelectron microscopy. C cytoplasm, N nucleus, NM, nuclear membrane. Bars, 200 nm. Arrowheads indicate localization of CHMP4P-EGFP labeled with anti-GFP antibody along the INM and in primary enveloped virions. In the lower panel, straight lines indicate the INM and dotted lines indicate the ONM and the envelope of virions in the perinuclear space. Images are representative of three independent experiments. f Quantification of gold particles on the NM with or without virions in the experiment in (e). Seven areas of each section were analyzed and the data are shown as the mean ± SEM. Data are representative of two independent experiments. The indicated P-values were obtained using the unpaired Student’s t-test (b, f)