Ectopic expression of FOG-1 in BMMCs represses mast cell phenotype and produces alternate lineage cells. (A) Percentage of viable GFP⁺ primary BMMCs retrovirally transduced with eGFP alone (vector), FOG-1–IRES-eGFP (FOG-1), or m1,5,6,9 FOG-1–IRES-GFP (m1,5,6,9). Viability was determined by Trypan blue dye exclusion. Time refers to days after FACS sorting of eGFP⁺ cells (2 d after retroviral infection). (B) FACS analysis for Annexin V (apoptosis marker) and 7-amino-actinomycin D (necrosis marker) expression of primary BMMCs transduced with each of the retroviral vectors described in A and examined 3 d after FACS sorting of GFP⁺ cells. Percentages of total cells that fall within the boxed FACS gates are shown. (C) May-Grunwald-Giemsa stains, FACS analysis showing side-scatter (SS) and forward scatter (FS), and FcεRI expression of primary BMMCs transduced with each of the retroviral vectors shown in A, sorted for GFP after 2 d, and cultured for 6 d in 2 U/ml EPO, 5 ng/ml TPO, 10 ng/ml IL-3, and 50 ng/ml SCF. (bottom) The green lines represent negative control staining for FcεRI (no IgE added; see Materials and methods). The purple histograms represent staining for FcεRI (IgE added). The percentage in the middle panel represents the proportion of total stained cells present within the negative staining control peak area. Bar, 10 μm. (D) May-Grunwald-Giemsa, o-dianisidine (benzidine), and acetylcholinesterase (AChE) histochemical stains of the cultures in C incubated for an additional 5 d. Benzidine⁺ cells stain black/brown; AChE⁺ cells stain orange. The percentage of positive-staining cells is indicated (±SEM; n = 2; 5,000 cells examined for all samples). Bar, 10 μm.