Induction of monocyte development by C/EBPα requires the distal enhancer. (A) Marrow mononuclear cells from PU.1^+/+ or PU.1^+/− mice exposed 5 days earlier to 5-fluorouracil were transduced with pBabePuro-C/EBPαER for 3 days and then exposed to 2 μg/mL puromycin for one day. Viable cells were then lineage-depleted and finally cultured with or without E2 for 2 days before FACS analysis. Percentage monocyte (% M) was calculated as (Mac-1⁺Gr-1⁻)/(Mac-1⁺Gr-1⁻ + Mac-1⁺Gr-1⁺) cells × 100%. (B) PU.1^+/kd or PU.1^kd/kd mice were analyzed similarly. (C) Morphology of PU.1^+/kd or PU.1^kd/kd cells transduced with C/EBPα-ER and cultured with or without E2 for 2 days. (D) Percentage of CFU-Gs or CFU-Ms compared with CFU-Gs + CFU-Ms obtained in methylcellulose culture with IL-3/IL-6/SCF without E2 after transduction of C/EBPα-ER into PU.1^+/+, PU.1^+/kd, or PU.1^kd/kd cells and cultured for 24 hours ± E2 prior to plating. Results shown are from 2 independent transduction experiments (top and bottom panels). The graphs represent the mean and SE from 4 determinations within each experiment (P values are from the Student t test).