Morphology of receptor-mediated mitophagy revealed by CLEM. (A) Representative CLEM images (left) and enlarged EM images (right) of WT and DRP1 KO HeLa cells expressing mito-mCherry (Mt-mChe) and GFP-LC3B and treated with DFP for 16 h. Boxed areas in the main (upper) CLEM images are shown below as single-color images. Further enlargement of the boxed areas in the EM images are shown at the far right. Arrows, IM; asterisks, ER; mt, mitochondrion. Scale bars: 1 µm (CLEM), 500 nm (EM, middle), and 100 nm (EM, right). See Fig. S1 for an additional CLEM image of DRP1 KO HeLa cells. (B–F) Representative CLEM-FIB–scanning EM images of DRP1 KO HeLa cells expressing the same markers as in A. (B) Confocal image of a cell that was processed for FIB-scanning EM, with the boxed area enlarged and shown below. Arrows indicate the direction of milling. Scale bars: 10 μm (top) and 1 μm (bottom). (C) Serial EM images (intervals of 20 sections) taken from the bracketed region in B, represented in the x–z planes. An IM with high electron density (arrows) can be seen in section numbers 258, 278, and 298. Scale bar: 500 nm. (D) 3D image reconstruction from serial sections 209–408, illustrating a mitochondrion (mt, pink), ER (blue), and IM (green). (E) In views 1 and 2, the image from D is rotated as indicated by arrows, and the entire IM is shown with the ER (left) or IM-ER contact sites (right, red). See also Videos 1 and 2. (F) Two representative EM sections illustrating some of the contact sites in E (colored arrowheads). Scale bars: 500 nm.