Fig. 3.

Increased association of p150glued and Trak1 with mitochondria isolated from mitotic Myo19-KO cells and implications of inhibiting dynein. (A) Representative images of cells in interphase and during mitosis stably expressing Halo-Myo19. Cells were transfected with Mito-EGFP and stained with TMR-ligand and Hoechst 34580. Scale bar: 20 µm. See also Movie 4. (B,C) Quantification of the correlation between the localization of the TMR-ligand (Halo-Myo19 construct), Mito-eGFP and Hoechst 34580. Myo19 remains associated with mitochondria at all phases of the cell cycle. (D) Representative immunoblots with indicated antibodies after cell fractionation into cytosol and mitochondria. Grey arrowhead indicates Halo-Myo19. (E) Quantification of proteins associated with purified mitochondria from synchronized cells in G1/S phase and prometaphase. Data are from 3–7 independent experiments; error bars represent ±s.e.m. *P≤0.05 (paired two-tailed Student's t-test). (F,I) Quantification of mitochondria distribution at cell poles versus the spindle at anaphase for cells of the indicated cell lines that were treated with DMSO or Dynarrestin and non-targeting (n-t) or Actr10 siRNA, respectively. (G,H,J,K) Quantification of the uneven mitochondrial distribution between the two halves of a cell at anaphase and telophase. Cells were treated with DMSO or Dynarrestin and n-t or Actr10 siRNA, respectively. Data are displayed as box plots with the corresponding 75th and 25th percentile. The whiskers show the 10th–90th percentiles. The minimum and maximum values are highlighted with dashes. n≥6 independent experiments, N≥30 cells. ****P≤0.0005; ***P≤0.001; **P≤0.01; *P≤0.05; n.s., not significant (one-way ANOVA with Bonferroni post-hoc test).