The ERG stem cell enhancer is active in human T-cell progenitors and in T-ALL. (A) Relative ERG expression in normal and leukemic cells. RNA was isolated from MACS- or FACS-sorted cord blood CD34⁺ cells, cultured T-cell progenitors and T-ALL xenografts, and ERG expression quantified by SYBR Green qPCR and expressed relative to GAPDH or β actin. ERG is expressed in CD34⁺ stem cells, but is rapidly down-regulated during T-cell differentiation. However, ERG is highly expressed in MOLT4 and xenograft T-ALL cells. (B) Chromatin accessibility at the ERG promoters and +85 enhancer in normal and leukemic cells. Chromatin accessibility measured by histone H3K9/K14 acetylation (AcH3) in CD34⁺ stem/progenitors, developing T cells, and MOLT4 T-ALL cells. AcH3 enrichment at the ERG proximal promoter (PP) and +85 element in the various cell types corresponds with ERG expression. (C) Chromatin accessibility across the ERG locus in NOD/SCID T-ALL xenografts. The top panel shows a VISTA plot of human/mouse sequence conservation > 50% across the ERG locus and flanking genes. The ERG promoters and enhancer are highlighted by gray bars. The +85 enhancer is also marked by an arrow. The bottom graphs show chromatin accessibility as determined by AcH3 ChIP-chip and ChIP-PCR in T-ALL xenografts. ChIP-chip plots show AcH3 enrichment along the ERG locus and flanking genes; clusters of AcH3 enrichment are seen at the ERG PP and +85 element in 6/6 T-ALL xenografts. Graphs at right show these peaks of enrichment confirmed by ChIP-PCR.