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. 2017 Nov 9;6:e30292. doi: 10.7554/eLife.30292

Figure 2. Mitochondria undergo DRP1-dependent fission upon AFM-mediated force application.

(A) Scheme of the experimental setup. (B) Quantification of fission events elicited by the application of AFM-induced mechanical forces observed in cells treated either with scrambled or DRP1 siRNA. Successful force application to an individual mitochondrion was defined as visible constriction of the mitochondrial matrix following tip approach (examples shown with blue arrowhead in C and D). (C) U2OS cells stained with Mitotracker Deep Red and transduced with viruses encoding mCherry-FIS1TM and a control shRNA were imaged by time-lapse microscopy. Two examples are shown. At t = 0 s, the cantilever of the AFM approached the cell in Contact mode, with a force set at 15 nN, at the position of the red ring. Green rings mark the time and area of tip retraction. Blue arrowheads indicate mitochondria that are visibly thinned by the pressure but have not yet undergone fission. Fission events are indicated by an orange arrowhead. OMM panels, mCherry-FIS1TM (red). Matrix panel, mtBFP (green). Scale bar, 5 µm. (D) As in (C) except that the cells were treated with a virus encoding DRP1 shRNA. Scale bar, 5 µm.

Figure 2.

Figure 2—figure supplement 1. Mitochondria undergo DRP1-dependent fission upon AFM-mediated force application.

Figure 2—figure supplement 1.

(A) U2OS KERMIT cells stably expressing mtBFP and Sec61β-GFP and treated with DRP1 siRNA were imaged by time-lapse microscopy. Two examples are shown. At t = 0 s, the cantilever of the AFM approached the cell in Contact mode, with a force set at 15 nN, at the position of the red ring. Green rings mark the time and area of tip retraction. Blue arrowheads indicate mitochondria that are visibly thinned by the pressure. Mito panels, mtBFP. Scale bar, 5 µm.