Fig 1: MCL-1 targeting induces changes in actomyosin structure organization in a DRP-1-dependent manner.A Mitochondrial fission was detected by TOM20 staining in CAFs treated or not with S63845 500 nM or A1331852 100 nM for 18 h in the presence or not of Mdivi-1 50 µM. Scale bar = 50 µm. B Quantification of mitochondrial network length was performed by measuring the size of TOM-20 labeled structures on deconvolved images in 10 cells/condition in 3 independent experiments. Objects smaller than 0.5 µm were not considered. C Representative co-immunoprecipitation of MCL-1 and DRP-1 and western blotting for MCL-1, DRP1, or NOXA in CAFs treated or not with S63845 in presence or not of Mdivi-1 (n = 3). FT: Flow Through. D qRT-PCR of MYH10, MYH2, ACTA2, and ITGA11 mRNA in CAFs treated or not by S63845 500 nM for 18 h in the presence or not of Mdivi-1. Mean and SEM of three independent experiments are represented as fold-change of mRNA level related to untreated control. Two-way ANOVA, *P < 0.1, **P < 0.01, ***P < 0.001, ****P < 0.0001, ns: not significant. E Quantification (percentage of cells positive for nuclear YAP or nuclear and cytosolic YAP) (Bottom) and confocal image (Top) of YAP staining (red) and FITC-phalloidin (green) in CAFs treated or not with S63845. Randomly generated images set names were randomized for analysis. Around sixty to one-hundred cells were analyzed per condition. Counting events were done manually through NIS-Elements software (Nikon software). Fluorescence intensity and positivity staining were determined by positive and negative control comparison, dependent on the experiment. The nuclei were counterstained with DAPI (blue). Scale bar = 50 µm. Two-way ANOVA, *P < 0.05. F Quantification (Bottom) and confocal image (Top) of Myosin IIB staining (green) and immunofluorescence of TOM-20 (red) in CAFs treated or not with different BH3 mimetics (as indicated) in the presence or not of Mdivi-1. Randomly generated images set names were randomized for analysis. Around sixty to one-hundred cells were analyzed per condition. Counting events were done manually through NIS-Elements software (Nikon software). Fluorescence intensity and positivity staining were determined by positive and negative control comparison, dependent on the experiment. The nuclei were counterstained with DAPI (blue). Scale bar = 50 µm, Two-way ANOVA, ###P < 0.001, ##P < 0.01, #P < 0.05 for myosin IIB-, **P < 0.01; *P < 0.05 for fragmented mitochondria.