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. 2016 Apr 6;12(4):689–702. doi: 10.1080/15548627.2016.1151580

Figure 1.

Figure 1.

DNM1L is required for FUNDC1-induced mitochondrial fragmentation and mitophagy. (A) Scrambled shRNA-treated and DNM1L knockdown cells were transfected with FUNDC1-MYC and GFP-LC3 for 24 h. The cells were then fixed and immunostained to detect HSP60 (red) and MYC (purple). Scale bar: 10 µm. (B) Cells treated as in (A), mitochondria fragmentation was quantified by counting numbers of cells with fragmented mitochondria versus all counted cells (mean+/−SEM; n = 100 cells from 3 independent experiments; **, P < 0 .01). (C) The GFP-LC3 aggregates in cells treated as in (A) were quantified with imageJ. The GFP-LC3 aggregation area vs. whole cell area was used to indicate the GFP-LC3 aggregation ratio (mean+/−SEM; n = 100 cells from 3 independent experiments; *, P < 0 .05). (D) Scrambled shRNA-treated and DNM1L knockdown cells were transfected with vector or FUNDC1-MYC for 24 h. The cells were then analyzed via western blotting. (E) HeLa cells were transfected with DNM1LK38A-Flag, FUNDC1-MYC and Mito-DsRed for 24 h. The cells were then fixed and immunostained to detect Flag (Green) and MYC (purple). Scale bar: 10 µm. (F) Cells treated as in (E), % mitochondria fragmentation was quantified as in (B) (mean+/−SEM; n = 100 cells from 3 independent experiments; *, P < 0 .05). (G) HeLa cells were transfected with DNM1LK38A-Flag, FUNDC1-MYC and GFP-LC3 for 24 h. The cells were then fixed and immunostained to detect MYC (purple) and Flag (red). Scale bar: 10 µm. (H) The GFP-LC3 aggregates in cells treated as in (G) were quantified with imageJ. The GFP-LC3 aggregation area versus whole cell area was used to indicate the GFP-LC3 aggregation ratio (mean+/−SEM; n = 100 cells from 3 independent experiments; *, P < 0 .05). (I) Scrambled shRNA-treated and DNM1L knockdown cells were transfected with DNM1LK38A and FUNDC1-MYC for 24 h. The cells were then analyzed via western blotting.