Figure 1
From: GBF1 and Arf1 interact with Miro and regulate mitochondrial positioning within cells
The inhibition of GBF1 induces the condensation of the mitochondrial network in the pericentriolar region without affecting cell shape. RPE1 cells were incubated with either GCA (10 μM) or DMSO alone (CT) for 1h30, were loaded with MitoTracker Orange for an additional 30 min prior to fixation and imaged using confocal microscopy. Images are maximum intensity Z-projections. All bars = 10 µm. Nuclei appear in blue (DAPI staining) and mitochondrial network in white or red. (A) Cells were stained for α-tubulin after fixation (green signal). (B) RPE1 cells were transfected with the ER marker Sec61β-GFP. (C) RPE1 cells were stained for γ−tubulin after fixation (green signal). In cells treated with GCA, the mitochondrial network is frequently condensed in the juxta-nuclear region next to the centrosome (white arrows) but not in control cells (green arrows). (D) The mitochondria area was segmented using ImageJ software. The mitochondrial network was converted into masks after the cells were manually contoured. (E) Quantification of image analysis. Left: the length of the mitochondrial network in close vicinity of the nucleus was quantified from confocal images using ImageJ (see D) in thirteen cells for each cell treatment and normalized by the nucleus perimeter. Bars represent the mean values (0.66 ± 0.153 and 0.37 ± 0.192 for CT and GCA treated cells, respectively). Right: the mitochondria surface area was quantified in 96 (CT) or 81 cells (GCA). Graph shows the distribution of the mitochondria area depending on the cell treatment. Lower: Likewise, the mitochondria surface area was quantified in 62 to 96 cells depending on the cell treatment. Results are the mean ± S.D. of three independent experiments, *p ≤ 0.05 versus control cells.
