HNF4A overexpression rescues Tifab deletion–induced LSPC dysfunction. (A-B) HNF4A overexpression increases LSPC proliferation (n = 5) (A) and enhances colony formation (n = 5) (B). (C) Glucose uptake assessed by NBDG incorporation in WT or Tifab KO LSPCs transfected with either vector- (Vec) or HNF4A-expressing viruses (n = 5). (D-F) Mitochondrial mass (n = 5) (D), mitochondrial membrane potential (n = 5) (E), and ROS levels (n = 10) (F) in WT or Tifab KO LSPCs transfected with either Vec- or HNF4A-expressing viruses, measured using MitoTracker, TMRE, and CellROX, respectively. (G-H) OCR (G) and ECAR (H) in WT or Tifab KO LSPCs transfected with either Vec- or HNF4A-expressing viruses (n = 5). (I) Western blot analysis of key components of ETC complexes I, II, III, IV, and V, as well as TIFAB and HNF4A, in WT and Tifab KO LSPCs transfected with either Vec- or HNF4A-expressing viruses (n = 3). ACTIN served as a loading control. (J) Proliferation of WT or Tifab KO LSPCs transfected with either Vec- or HNF4A-expressing viruses (n = 8). (K) Survival curve of sublethally irradiated recipient mice transplanted with Vec- or HNF4A-transduced WT or Tifab KO KMT3A::MLLT3 LSPCs (n = 8 in each group). ∗P < .05; ∗∗P < .01. Statistical tests used in panels A-B, Mann-Whitney U test; in panels C-F and J, 1-way analysis of variance and Tukey multiple comparisons test; in panel K, log-rank test.