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. Author manuscript; available in PMC: 2023 Jan 20.
Published in final edited form as: Mol Cell. 2021 Dec 1;82(2):447–462.e6. doi: 10.1016/j.molcel.2021.11.006

Figure 2: SILEC-SF reveals compartment-specific acyl-CoA profiles.

Figure 2:

A) Differential centrifugation method for mitochondria and cytosol isolation. B-E) SILEC-SF acyl-CoA quantitation in whole cell lysate (WCL), mitochondria, cytosol and high-density debris B) Brown adipocytes in cell culture (n=4 replicate samples) C) Acly−/− mouse embryonic fibroblasts (MEFs) were incubated in DMEM supplemented with 10% dialyzed FBS and acetate (1 mM) for 4 h before cell harvest (n=4 replicate samples) D) Mouse liver tissue (n=6 mice) E) Transmural left ventricle of human heart (n=5 replicate samples from a single heart). F) Mito-IP protocol with SILEC-SF G) SILEC-SF using Mito-IP was applied to HepG2 cells incubated in serum free DMEM for 2 h before harvest (n=3 replicate samples). All panels display mean with error bars representing standard deviation. Data for all acyl-CoA species that were quantified in each fraction are displayed. Those that were not quantified showed insufficient signal intensity for the analyte, the internal standard or both. Some metabolites indicated in the legend for C-G were not quantified in the mitochondrial fraction. These were B) HMG-CoA, D) Malonyl-CoA and E) CoASH, G) all except for acetyl-CoA succinyl-CoA and CoASH. (iso)Butyryl-CoA = Butyryl-CoA + Isobutyryl-CoA, (iso)Valeryl-CoA = Valeryl-CoA + Isovaleryl-CoA (isomers are not distinguished in analysis). HMG-CoA = 3-hydroxy-3-methylglutaryl-CoA.