ME is preferentially expressed in HSCs. (A) Whole BM cells were loaded with FDG and then stained with a cocktail of Cy-chrome-labeled lineage markers (B220, CD3, CD11b, Ly6G, and Ter119). Shown is a 2-dimensional scatterplot of FL3 (y-axis; Cy-chrome fluorescence) compared with FL1 (FDG fluorescence). Enumeration of FL1 channel fluorescence in BM from WT mice reveals minimal staining (< 0.1%), whereas ME^m1/+ BM shows that 2% of the cells are positive; the majority (> 90%) are Lin⁻. (B) Whole BM cells from ME^m1/+ mice were loaded with FDG and stained with Cy-chrome–labeled lineage markers as above, as well as with PE-labeled Sca-1 (FL2) and APC-labeled c-Kit (FL4). Gating was performed first on Lin⁻ cells, and then on c-Kit/Sca-1 double-positive cells. Within this population, 97% of the cells were FDG positive. These studies and those in panel A were repeated at least twice with identical results. (C) FDG staining reveals low-level ME expression in committed progenitors. Shown is the percentage of HSCs and committed progenitors that stained positive for β-galactosidase activity using the fluorogenic substrate FDG. HSCs were identified as LSKs, whereas committed progenitors were identified as described by Tothova et al.¹³  The error bars represent SD based on 3 independent flow cytometry readings of different mice. (D) Quantitative Taqman PCR analysis of RNA from subpopulations of mouse BM cells including LT-HSCs, ST-HSCs, CMPs, GMPs, granulocytes (Gr), monocytes (Mo), erythroids (Er), CLPs, and T and B lymphocytes. Cell purity, as assessed by analytical flow cytometry, was > 97%. Data were normalized first to β-actin, then to expression level in LT-HSCs and are shown ± SD. (E) Quantitative Taqman PCR in isolated LSK hematopoietic progenitor cells demonstrating specific ablation of the ME transcript in LSKs in ME^m1/m1 mice (light gray) versus WT (dark gray) mice ± SD. Primers/probe sets detected the nonoverlapping exon 1 of ME and exon 1 of Evi1, respectively.