Fig. 4: DKO of SLC25A16 (A16) and SLC25A42 (A42) leads to a depletion of mitochondrial CoA pool and an impairment of TCA cycle and lipid metabolism.
a, Left, cell proliferation rate of Ctrl, SLC25A42 KO, SLC25A16 KO and A16/A42 DKO K562 cells. Right, the proliferation rate on day 9. The dotted line denotes the expected log fold change based on the additive effects from A42 and A16 single KOs, if there were no genetic interactions. b, Oxygen consumption rate (OCR) for Ctrl and A16/A42 DKO K562 cells during a Seahorse Mito Stress Test (n = 4). c, Volcano plot of annotated mitochondrial small polar metabolite profiling from Ctrl and A16/A42 DKO K562 cells, highlighting the changes of the TCA cycle intermediates. Accumulated and depleted metabolites in DKO cells are marked in red and blue, respectively. d, Volcano plot showing the untargeted analysis of mitochondrial small polar metabolite profiling from Ctrl and A16/A42 DKO K562 cells, highlighting a significant depletion of CoASH and succinyl/MMA-CoA, and accumulation of acylcarnitines (see Extended Data Fig. 4). e, LC–MS peak intensities of mitochondrial CoASH, succinyl/MMA-CoA, α-KG and two representative acylcarnitines. f, Basal and FCCP-mediated uncoupled OCR for Ctrl and A16/A42 DKO K562 cells during a Seahorse FAO stress assay using palmitate as the sole fuel source (n = 5). g, Stable isotope tracing of A16/A42 DKO K562 cells using [U-13C5]-glutamine (2 mM for 2 h). The y axis shows the natural abundance-corrected isotopologue fraction. All data are expressed as mean ± s.d.; n = 3 unless otherwise indicated. Statistical significance was calculated using two-tailed Student’s t-test (c, d), one-way ANOVA followed by Tukey’s test (a, f, g) or Fisher’s least significant difference test (e). ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05; ns, P > 0.05.
