CSB-deficient cells show increased mitochondrial content and decreased mitochondrial biogenesis. (A) Citrate synthase activity in CS1AN cells expressing WT CSB or an empty vector or in MDF from young or old CSBm/m or WT mice or in HSMM expressing scrambled or CSB-specific shRNA (n = 3–5; data are represented as mean ± SEM). (B) Representative confocal image of CS1AN cells expressing WT CSB or an empty vector transfected with the mitochondrial RFP-tagged protein using organelle lights. (C) Quantification of the mitochondrial RFP signal normalized to cell number (n = 3; data are represented as mean ± SEM). (D) Representative flow cytometry experiment of CS1AN cells expressing WT CSB or empty vector treated with 50 nM MitoTracker green. Inset: quantification of the flow cytometry data (n = 3; data are represented as mean ± SEM). (E–H) Quantitative PCR analysis of genes involved in mitochondrial biogenesis in CS1AN cells expressing WT CSB or empty vector (E), MDFs from young (F) and old (G) mice, and HSMM subjected to lentiviral CSB knockdown (H). The inset in H shows efficiency of lentiviral shRNA knockdown of CSB (n = 3; data are represented as mean ± SEM; *, P < 0.05; **, P < 0.01).
