PUS1 deficient mice exhibit mitochondrial dysfunction. (A-B) Mitochondrial biomass (A) and MMP (B) of Lin^–/LKS^–/LSK⁺/LT-HSC/ST-HSC/MPP/MEP/CMP/GMP cells were evaluated by flow cytometry. Female mice at 4 weeks: WT, n = 5; S172fs, n = 4. (C-D) Mitochondrial biomass (C) and MMP (D) of BM Ter119⁺ cells were evaluated by flow cytometry. The representative histogram (left) and gMFI (right) are shown. Male mice at 4 weeks: WT, n = 6; S172fs, n = 3. (E-F) Cytoplasmic (E) and mitochondrial (F) ROS levels of BM Ter119⁺ cells evaluated by CellROX and MitoSOX, respectively. The representative histogram (left) and gMFI (right) are shown. Male mice at 4 weeks: WT, n = 6; S172fs, n = 3. (G-H) Measurement of cellular oxygen consumption in BM Ter119⁺ cells of mice. OCRs were monitored by injecting 1 μM Oligo, 2 μM FCCP, and 1μM Rot/AA in order using the Seahorse XFe24 Extracellular Flux Analyzer (G). The average basal and maximum oxygen consumptions were normalized to WT mice (H). n = 6, male mice at 7 to 8 weeks. (I) Activities of mitochondrial respiratory chain complexes in WT and mutant mice. Complex I , II, III, and IV (WT, n = 9-10; S172fs, n = 6, male mice at 7-8 weeks; Complex I , II, and IV: BM cells; Complex III: SP cells) activities were measured according to the manuals of relevant kits. (J) Cellular ATP levels of BM cells between WT and S172fs groups were detected by CellTiter-Glo 2.0 Reagent. WT, n = 10; S172fs, n = 6, male mice at 7 to 8 weeks. Values in all panels denote mean ± SD (∗P < .05; ∗∗P < .01; ∗∗∗P < .001); unpaired Student t test (C-J) or 2-way ANOVA (A-B).