Metabolic homeostasis is impaired in Setdb1-deficient HSPCs. (A) Results of metabolome analyses of GMPs freshly isolated from Setdb1^Δ/Δ mice 2 weeks after the first injection of tamoxifen. Relative values (WT values were set as 1) are shown as the mean ± SD (glucose 6-phosphate, F6P, F1,6BP, n = 4-6; pyruvic acid, n = 2; ribose-5-P, reduced glutathione/oxidized glutathione, n = 3; ATP, n = 9). (B) F1,6BP levels relative to F6P in GMPs (n = 4), MLL-AF9-transformed GMPs 48 hours after the tamoxifen treatment (n = 3), and MLL-AF9-transformed GMPs overexpressing Fbp2 (n = 1). (C) ATP levels in LSK cells and GMPs freshly isolated from Setdb1^Δ/Δ mice 2 weeks after the first injection of tamoxifen and MLL-AF9-transformed GMPs 48 hours after the tamoxifen treatment. Data are shown as the mean ± SD (n = 3). (D) ATP levels in LSK cells transduced with an Fbp2 retrovirus in culture (left) and their growth in culture supplemented with SCF (10 ng/mL) and thrombopoietin (10 ng/mL) without or with IL-3 (10 ng/mL). mRNA levels of exogenous Setdb1 in LSK cells detected by RT-PCR were normalized to Hprt1 expression. (Right) Relative expression levels. Data are shown as the mean ± SD (n = 3). (E) Repopulating capacity of Fbp2-overexpressing HSCs in vivo. Fifty CD45.2 CD34⁻LSK HSCs were transduced either with a control or Fbp2 retrovirus, and then transplanted into lethally irradiated CD45.1 mice (n = 6) with 2 × 10⁵ CD45.1 competitor cells. The chimerism of CD45.2⁺GFP⁺ cells derived from transduced HSCs as well as CD45.2⁺GFP⁻ cells derived from nontransduced HSCs was monitored. At 16 weeks after primary transplantation, all BM cells from recipients were pooled and 1 × 10⁷ cells were infused into the secondary recipients. The proportions of GFP⁺ cells in donor-derived cells are plotted in blue (control) and red (Fbp2) lines. *P < .05; **P < .01; ***P < .001.