Fig. 5: Inhibition of Ringo formation correlates with decreased respiration.

a Total protein extracts prepared from WT, dnm1Δ or ADH1-DNM1 cells in glycerol (SG) media and analyzed by immunoblotting as indicated. MW (kDa) indicated on the right. Right graph: Dnm1 levels normalized to Pgk1 in dnm1Δ and ADH1-DNM1 relative to the WT strains in SG. Mean ± s.d. from n = 3 independent experiments (blue circles). ****p < 0.0001 (One-way Anova followed by Tukey’s multiple comparisons test). b SIM acquisitions (z-projections) of WT and ADH1-DNM1 cells labeled with mt-mCherry and Tom70-GFP in fermentation (top) or respiration (bottom). Scale bar, 2 µm. Right graphs: percentage of WT (blue) and ADH1-DNM1 (red) cells with Tubular, Ringo, Fragmented, Aggregated, Hyperfused or HFR mitochondria during fermentation (Top) or respiration (Bottom). Mean ± s.d. from > 95 cells in n = 3 independent experiments (colored circles). ****p < 0,0001, ***p = 0.0007 (Two-way ANOVA multiple comparisons) (c) Dextrose and glycerol serial dilutions of WT, dnm1Δ or ADH1-DNM1 strains at 37 °C. Right graph: Quantification of indicated cells grown at 37 °C on glycerol media relative to the WT strain. Mean ± s.d. from n = 3 independent experiments. ****p < 0.0001, **p = 0.0014 (Two-tailed unpaired t-test) (d) Survival curves derived from single-cell analysis of WT (blue) and ADH-DNM1 (red) cells in SD (top) or SG (bottom) media (Supplementary Fig. 9c), were generated using the Kaplan–Meier estimator (WT n = 32 cells; ADH1-DNM1 n = 32 cells for SD; WT n = 34 cells; ADH1-DNM1 n = 34 cells for SG). e Oxygen Consumption Rates (OCRs) of WT, dnm1Δ and ADH-DNM1 cells grown in 2% Ethanol media (SE) at 30 °C. Mean ± s.d. from n = 8 independent experiments (green circles). ****p < 0.0001, ns p = 0,9299 (Two-tailed unpaired t-test). ns, not significant.