MICOS subcomplexes independently assemble and localize in proximity to ERMES foci. (A) Representative deconvolved fluorescence microscopy images of Δmic19 cells expressing Mic60-mCherry (magenta), Mdm34-EGFP (ERMES; green), and mito-TagBFP (blue). Examples are shown of each category of spatial relationships between Mic60 assemblies and ERMES complexes and their indicated frequency as in Fig. 1 J. Arrows depict either complete overlap (white) or the relative positioning of MICOS assembly (magenta) compared with ERMES (green). The full overlap example is redisplayed from Fig. 1 H. (B) As in A for Δmic60 cells expressing Mic27-mCherry (magenta). The full overlap example is redisplayed from Fig. 1 I. (C) Maximum-intensity projections of deconvolved fluorescence microscopy images are shown of Δmic19 cells expressing Mic60-EGFP (green) and Mic27-mCherry (magenta) in the presence (ρ⁺) and upon depletion (ρ⁰) of mtDNA. White arrows indicate colocalization of Mic60 and Mic27 assemblies and green arrows indicate Mic60 assemblies with no detectable Mic27 assembly colocalization. (D) Images are shown of Δmic19 (top) and Δmic10 (bottom) cells expressing Mic60-mCherry (magenta) and mito-EGFP (green). Arrows indicate Mic60 assemblies. (E) Images are shown as in D for cells coexpressing mito-TagBFP (blue) and Mdm34-EGFP (green). Arrows mark Mic60 assemblies that localize in proximity to ERMES. (F) Images are shown as in D for cells expressing mito-TagBFP (blue) and Mic60-EGFP (green) and constitutively overexpressing mKate-Vps39 (magenta). Scale bars = 2 µm.