Samples were subjected to electron microscopy with minor variations from Zischka et al. (2008) (link). Briefly, 3×105 cells were immediately pelleted, fixed in 2.5% glutaraldehyde, fixed with 1% osmium tetroxide, dehydrated with acetone, and embedded in EPON resin. Ultra-thin sections were stained with lead citrate and UranyLess and imaged on a 1200EX electron microscope (JEOL, Tokyo, Japan) at 60 kV. Electron micrographs were recorded with a KeenView II digital camera (Olympus, Munich, Germany) with iTEM Ver. 5.0 (analySIS FIVE, Olympus, Munich, Germany).
Mitochondrial roundness was quantified as described in Tobias et al. (2018) (link) with ImageJ (V1.52i). Briefly, at least 150 mitochondria per condition were encircled with the polygonal section tool and fitted to an ellipse. Outliers were identified and removed with GraphPad Prism by the ROUT method at Q=1%. Mitochondrial cristae density was quantified by encircling 100 mitochondria per condition and manually counting the cristae, which were then divided by mitochondrial area. Lysosomes were counted manually in 100 µm2 rectangular sections, each composed from four 25 µm2 adjacent sections from the same slice. At least eight composite sections per condition were analyzed.
Mitochondrial roundness was quantified as described in Tobias et al. (2018) (link) with ImageJ (V1.52i). Briefly, at least 150 mitochondria per condition were encircled with the polygonal section tool and fitted to an ellipse. Outliers were identified and removed with GraphPad Prism by the ROUT method at Q=1%. Mitochondrial cristae density was quantified by encircling 100 mitochondria per condition and manually counting the cristae, which were then divided by mitochondrial area. Lysosomes were counted manually in 100 µm2 rectangular sections, each composed from four 25 µm2 adjacent sections from the same slice. At least eight composite sections per condition were analyzed.