Fig. 3: Senescence, aberrant myogenesis and Q deficiency are associated with ETFDH deletion.
a,b, Lactate production (a) and energy map (b) in CRL and ETFDH-ko myoblasts. n = 4. c, PI staining of the cell cycle in CRL and ETFDH-ko myoblasts. n = 3 independent repeats. Cell percentages in the G1, S and G2/M phases are indicated. d, Representative western blot of proteins related to the cell cycle and senescence in two clones of CRL and ETFDH-ko myoblasts. β-Actin is shown as a loading control. e, RNA relative expression of cyclins, CDKs and other proteins related to the cell cycle in CRL and ETFDH-ko myoblasts. The upper scheme illustrates Skm cell-cycle regulators. n = 3. f, RNA relative expression of proteins related to myogenesis (MyoD, Myf5, MyoG, MyHC1 and MyHC7) in CRL and ETFDH-ko myoblasts and myocytes. n = 3. g, PI staining of the cell cycle in CRL and patient-derived fibroblasts. n = 4, three independent repeats. h, Relation between total Q10 and mitochondrial ROS in CRL and patient-derived fibroblasts. n = 4, three independent repeats. i, Representative immunocapture (IP) of COQ2 blotted with anti-ETFDH antibody in mouse Skm. Tubulin is shown as a loading control. j, Left, representative images of PLA between COQ2 and ETFDH in CRL and ETFDH-ko myoblasts. Right, quantification histogram. Five images per condition, n = 3. k, Mitochondrial ROS in the presence or absence of the COQ2 inhibitor 4-CBA in ETFDH-ko and double-knockout (COQ2-ko, ETFDH-ko) cells. n = 3. l, Left, representative respiratory profile of CRL (grey traces) and ETFDH-ko (red traces) myocytes treated or not with 0.5 mM 4-CBA. Glucose was used as a substrate. Right, histogram of the quantification of maximal respiration. n = 8 replicates per condition. Results are shown as the mean ± s.e.m. of the indicated n. *, **, ***, ****P < 0.05, 0.01, 0.001 and 0.0001 when compared with CRL by two-tailed Student’s t test (a, j), one-way ANOVA with Dunnett’s test (k) and two-way ANOVA with Šidák’s test (f, l).
