Fig. 3: ARF1 localizes to LD-mitochondria contact sites and regulates fatty acid transfer.

A Representative optical section images showing LD-mitochondrion contacts in C2C12 cells immunostained with MitoTracker (mitochondrial labeling, purple), LipidTOX (LD labeling, yellow), and ARF1-specific antibody. Cells were treated for 8 h under glucose-fed or glucose-starved conditions. Scale bar, 10 μm. (B, C) Fluorescence intensity profiles from the indicated line scans in panel A, showing the distribution of MitoTracker LipidTOX, and ARF1 labeling across the contacts between LDs and mitochondria. D Representative optical section images of C2C12 cells incubated with Red C12 for 16 h and then stained with MitoTracker (mitochondrial marker, green) and BODIPY493/503 (LD marker, green). Cells were transfected with ARF1 overexpression plasmid or empty vector and then treated for 8 h under glucose-fed or glucose-starved conditions. Scale bar, 2 μm. E Quantification of Red C12/LD overlap coefficient. n = 18 cells. F Quantification of Red C12/mitochondrial overlap coefficient. n = 18 cells. G Representative optical section images of C2C12 cells incubated with Red C12 for 16 hours and then stained with MitoTracker and BODIPY493/503. Cells were treated with Scrambled or ARF1 siRNA under glucose-fed or glucose-starved treatment conditions. Scale bar, 10 μm. H Quantification of Red C12/LD overlap coefficient. n = 24 cells. I Quantification of Red C12/mitochondrial overlap coefficient. n = 24 cells. E, F, H, I Data are presented as means ± SD of three biologically independent replicates, analyzed by two-way ANOVA followed by Turkey’s post-hoc test.