MDCs form through membrane extension intermediates. (A) Super-resolution time-lapse images of Rap-induced MDC formation in yeast cells expressing Tom70-yEGFP and Tim50-mCherry. Images were acquired over 120 min. White arrows mark MDC. Scale bar = 1 µm. See associated Video 5. (B) Quantification of the initial morphology of Tom70-yEGFP structures during MDC biogenesis. n = 52 events from seven experiments. (C) Super-resolution time-lapse images of Rap-induced MDC formation in yeast cells expressing Tom70-yEGFP. Images were acquired over 120 min. White arrows mark MDC. Scale bar = 1 µm. See associated Video 6. (D) Representative super-resolution confocal fluorescence microscopy images of diverse MDC morphologies observed in Rap-treated haploid yeast expressing Tom70-yEGFP and Tim50-mCherry. MDC structures are indicated by white arrows. Scale bar = 1 µm. The yellow line marks the position of the line-scan fluorescence intensity profile shown to the right. The left and right y axes correspond to Tom70-yEGFP and Tim50-mCherry fluorescence intensity, respectively. Comp = compartment, LS = large spheres, BME = bright membrane extensions, ISC = irregular-shaped clusters. (E) Quantification of the MDC morphologies shown in D as a percent of total MDCs. Error bars show mean ± SE of three replicates, n > 60 MDCs per replicate.