Fig. 7: SLC25A16 (A16) and SLC25A42 (A42) are required for mitochondrial import of free CoASH.
a, Schematic of the mitochondrial uptake assay of the CoA pool using immuno-isolated mitochondria from K562 cells expressing HA-MITO tag. The substrate pool [13C315N1]-acyl-CoAs, including CoASH (M+4, stable isotope labelling at the CoA moiety marked in red), was prepared from K562 cells after [13C315N1]-pantothenate labelling for 3 days. b, Labelled CoASH (M+4) from the mitochondrial uptake assay of CoA pool using A42 KO and A42 KO re-expressing A42WT K562 cells. The input level of CoASH (M+4) from the 5× diluted substrate pool is shown. c, Labelled CoASH (M+4) (left) and labelled acetyl-CoA (M+4) (right) from the mitochondrial uptake assay of CoA pool using Ctrl and A16/A42 DKO mitochondria. The input levels of CoASH (M+4) and acetyl-CoA (M+4) from the 5× diluted substrate pool are shown. d, Schematic of mitochondria acetyl-CoA uptake assay of 13C2-acetyl-CoA (M+2) at 10 μM using immuno-isolated mitochondria from K562 cells expressing HA-MITO tag. The substrate 13C2-acetyl-CoA (M+2) contained labelling at the acetyl moiety, marked in red. e, Labelled acetyl-CoA (M+2) from the mitochondrial acetyl-CoA uptake assay using Ctrl mitochondria. The input level of 13C2-acetyl-CoA (M+2) from the 5× diluted input is shown. f, Summary diagram highlighting the physiological role of A16 and A42 in mitochondrial CoASH uptake and in regulating mitochondrial CoA metabolism. All data are expressed as mean ± s.d.; n = 3 unless otherwise indicated. Statistical significance was calculated using a two-tailed Student’s t-test. ***P < 0.001; **P < 0.01; *P < 0.05.
