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. 2020 Oct 20;6:107. doi: 10.1038/s41420-020-00342-7

Fig. 3. The MEKK3-MEK5-ERK5 kinase cascade prevents accumulation of excess mitochondria.

Fig. 3

a U2OS mito-mCherry cells were transfected with the indicated siRNA oligos. Seventy-two hours later, mitochondrial accumulation was analyzed by flow cytometry. Mean ± SD of n = 3 independent experiments is shown. b, c Mitochondrial accumulation in U2OS mito-mCherry cells treated overnight with vehicle (DMSO) or XMD8–92 was measured by flow cytometry or western blot, respectively. Mean ± SD of n = 2 independent experiments is shown b. d, e Mitochondrial levels of three independent MAPK7/ERK5-knockout U2OS clones were assessed by western blotting analysis of mitochondrial marker mtCOX2 (d) or by MitoTracker Green FM staining and flow cytometry (e). f Protein levels of p62 from U2OS cells that were treated with ERK5 siRNA for 72 h were detected by western blotting analysis. XPB was used as a loading control. g, h U2OS cells were either transfected with the indicated siRNAs for 72 h (g) or treated with the indicated drug at 10 µM overnight (h). Cells were fixed and stained with p62 antibody and then imaged. See Supplementary Fig. 1f, g for representative images.