Data includes ET tilt series and reconstructions from samples prepared on 9.6.2010 and 28.6.2010. bEND5 cells were grown on 1.5-mm sapphire discs coated with gelatine. After 1 h treatment with 1.25 µM LtA, the cells were covered with 20% BSA in DMEM and high-pressure frozen. Substitution was carried out in 0.3% uranyl acetate, 2% OsO4, and 10% water in ethanol. Specimens were first kept at −90 °C for 16 h, after which the temperature was increased by 10 °C/h to 0 °C. The specimens were then washed with cold ethanol and acetone, gradually infiltrated into Epon at RT and flat embedded. Sections 90 nm thick were cut parallel to the sapphire disc and imaged using Tecnai FEG 20 microscope operated at 200 kV. Images were collected with a 4k × 4k Ultrascan 4000 CCD camera at 19,000×, providing a 2× binned pixel size of 1.2 nm. For a tilt series, the specimen was tilted at 1° intervals using a high-tilt specimen holder between ±62°. The images were acquired by SerialEM software, and dual-axis tomography was applied. The alignment of tilt series and tomographic reconstruction were done with IMOD software package using 10-nm colloidal gold particles overlaid on the top of the section as fiducial markers. Modelling was done with Amira software. Ju M, Ioannidou S, Munro P, Rämö O, Vihinen H, Jokitalo E, Shima DT. A Na,K-ATPase-Fodrin-Actin Membrane Cytoskeleton Complex is Required for Endothelial Fenestra Biogenesis. Cells. 2020 Jun 3;9(6):1387. doi: 10.3390/cells9061387. PMID: 32503129; PMCID: PMC7349347. Figure 4C Ju et al_Video-S1.avi Ju et al_Video-S2.avi

2024