The mitochondrial fission and fusion machinery perform competing roles in MDC formation. (A) Immunoblots of whole-cell protein extracts from the indicated yeast strains probed for Fzo1 using an antibody that targets the N-terminus of Fzo1. Arrows point to protein bands for sfGFP-Fzo1 and Fzo1. Note that sfGFP-Fzo1 is expressed from the NOP1 promoter. Pgk1 is shown as a loading control. (B) Immunoblots of whole-cell protein extracts from the indicated yeast strains at the indicated temperatures, all probed for Fzo1 using an antibody that targets the N-terminus of Fzo1. Arrows point to protein bands for Fzo1 and a non-specific (NS) protein. Pgk1 is shown as a loading control. (C and D) Super-resolution confocal fluorescence microscopy images of wild-type (C) or mgm1∆ (D) yeast cells expressing mgm1-5, Tom70-yEGFP, and Tim50-mCherry treated with 200 nM Rap at the indicated temperatures. MDCs are indicated by white arrows. Scale bar = 1 µm. (E) Quantification of MDC formation in the indicated yeast strains upon treatment with either DMSO or Rap at the indicated temperatures. Error bars show mean ± SE of three replicates, n ≥ 100 cells per replicate. Source data are available for this figure: SourceData FS5.