Forced expression of Gfi1 partially restores the function of Mysm1-deficient HSC. (A-F) 1 × 10³ LSKFlt3⁻ cells sorted from WT or Mysm1^−/− mice (CD45.2) and Mysm1^−/− LSKFlt3⁻ cells transduced with pMIG-Gfi1 were transplanted into lethally irradiated recipients (CD45.1) together with 2 × 10⁵ competitor BM cells (CD45.1). (A) Flow cytometric analyses of WT, Mysm1^−/− CD45.2 cells, and donor-derived chimerism (CD45.2) 6 weeks after transplantation in the recipient mice transplanted with WT, Mysm1^−/− cells, and Mysm1^−/− cells expressed with Gfi1 (pMIG-Gfi1 Mysm1^−/−). (B) Percentages of donor-derived cells (CD45.2) in BM. Each dot indicates an individual recipient mouse. (C-D) Flow cytometric analyses and percentages of donor-derived LSKFlt3⁺ cells in the recipient mice 6 weeks after transplantation. Each dot indicates an individual recipient mouse. (E-F) Flow cytometric analyses and percentages of donor-derived CD45.2 or CD45.1 cells gated on B220⁺ B cells, CD3⁺ T cells, and CD11b⁺Gr1⁺ myeloid cells in the recipient mice 6 weeks after transplantation. Each dot indicates an individual recipient mouse. (A-F) Data are representative of two independent experiments with n = 5 mice per group in the first experiment and n = 3 mice per group in the second experiment. Shown are means ± SD of 1st experiment with n = 5 mice per group, ***P < .001. (G-J) Sorted Mysm1^−/− LSK cells were infected with retroviral vectors encoding either Gfi1 and GFP (pCDH-Gfi1-GFP) or GFP alone (pCDH-GFP) in vitro. Hoechst 33258/Pyronin Y staining (G), annexin V staining (H), BrdU incorporation assay (I) or cell-cycle analysis (J) were performed 48 hours after infection to monitor G₀ phase, apoptosis, proliferation, and cell-cycle profile, respectively. (G-J) Data are representative of three independent experiments, *P < .05.