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. 2024 Jul 1;223(9):e202311126. doi: 10.1083/jcb.202311126

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© 2024 Li et al.

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Figure 6. — Dynamic regulation of mito-LD contact sites revealed via FAB^mito-LD. (A) Detection of lipid droplets (LDs) and mitochondria (mito)-LD contact sites in oleic acid (OA)-treated HeLa cells producing FAB^mito-LD (top) monitored via confocal microscopy. Representative maximal intensity projected images from two axial slices (0.6 µm in total thickness) are shown in bottom panels. (B) Dynamics of mito-LD contact sites on a LD in OA-treated U2OS cell producing FAB^mito-LD monitored by confocal microscopy over time (top). Relative intensity profiles of mito-LD measured by clockwise circular scanning are shown in the bottom panels. (C) Perilipin 5 (PLIN 5) levels in HeLa cells transfected with scramble (siCtrl) or PLIN 5 siRNA detected by Western blot. * indicates non-specific band. (D) Quantification of relative PLIN 5 level described in C. Data are from three independent experiments (**P ≤ 0.01, unpaired t test, two-tailed). (E) Relative levels of mito-LD contact sites in OA-treated HeLa cells transfected with scramble or PLIN 5 siRNA. Raw data and mean ± SD are shown (61–63 cells from three independent experiments). n.s. = not significant, **P ≤ 0.01, assessed by two-tailed t test. (F) The temporal dynamics of mito-LD contact sites in OA-treated HeLa cells following OA withdrawal in DMEM and in DMEM with 10 µM of isoproterenol or 4 mM 2DG. Mean ± SE are shown (37–58 cells from three or four independent experiments). Statistical significance was compared to time zero by one-way ANOVA. **P ≤ 0.01; ***P ≤ 0.001. Source data are available for this figure: SourceData F6.