Fig. 4: MTCH1-deficiency reduced mitochondrial NAD⁺ levels, leading to ROS elevation and GPX4 inhibition.

A, B NAD⁺ (A) and NAD⁺/NADH (B) levels in whole cells or mitochondria lysates of MTCH1^WT (n = 2) and MTCH1^KO (n = 3) HeLa clones. C Relative transcriptional levels of SLC25A51, NMNAT1, NMNAT3, NAPRT1 and NRK1 in MTCH1^WT (n = 2), MTCH1^KO (n = 3) HeLa clones. D Relative cell viability in MTCH1^WT (n = 2) and MTCH1^KO (n = 3) HeLa clones treated with different concentrations of NAD⁺ (0, 1, 2, 5, 10 mM) for 24 h. * refers to the compare of NAD⁺ treated and untreated clones. E, F The levels of ROS (E) and mitochondrial MDA (F) in MTCH1^WT (n = 2), MTCH1^KO (n = 3), and MTCH1^KO treated with 5 mM NAD⁺ for 24 h (n = 3) HeLa clones. G Relative transcriptional levels of GPX4 in Parental (n = 1), MTCH1^WT (n = 2), MTCH1^KO (n = 3), and MTCH1^KO treated with 5 mM NAD⁺ for 24 h (n = 3) HeLa clones. Data were presented as mean ± SEM of at least 3 independent replicates (**P < 0.01, ***P < 0.001, ns, no significant) and analyzed by one-way ANOVA with Tukey’s multiple comparisons test or unpaired t-test.