Fig. 5: Mip1 variants defective in strand displacement do not support yeast mitochondrial function.

A Schematic representation of the plasmid-shuffling and petite formation frequency assays. Yeast colonies were classified as white (no respiration), sectored (mixed), and red (active respiration). Strains retaining respiratory activity were obtained from the plasmid-shuffling assay and selected by propagation on a glycerol-containing medium. The schematic was created in BioRender. Sokolova, V. (2025) https://BioRender.com/x01j374. B Yeast colonies formed on 5-FOA-containing media. C The plasmid-shuffling assay of the W303a mip1Δ strain expressing either the WT MIP1 or the indicated MIP1 variants. The average fraction of the red, sectored, and white colonies (n = 3) is shown. Data are presented as mean ± SD. An unpaired, two-sided Student’s t-test was used for statistical analysis, with asterisks indicating significant differences between an individual mutant and the wild-type strain. p < 0.05 (*), p < 0.01 (**), p < 0.001 (***). Data are presented as mean ± SD. D Analysis of DAPI staining of mutant yeast strains by fluorescence microscopy (n = 3). The mtDNA and nuclear DNA staining are shown for W303 WT MIP1, W303 mip1Δ, and the MIP1 variants F849A and TETRA, which did not produce colonies containing respiratory active cells in the 5-FOA shuffling procedure. M mitochondria, N nucleus. E Petite formation frequency in mutant MIP1 strains obtained via FOA shuffling, compared to the WT MIP1 strain (n = 3). An unpaired two-sided Student’s t-test was used for statistical analysis, with *** indicating a significant (p < 0.001) difference between an individual mutant and the wild-type strain.