PAX5 is organized in an open chromatin conformation in t(8;21) AML and non-t(8;21) myeloid precursors. (A) Quantitative real-time PCR analysis showing PAX5 expression relative to TBP expression in Ramos (B-cell line), Nalm-6 (pre-B cell line), Kasumi-1, and SKNO-1 (t(8;21) AML cell lines), primary cells from 2 t(8;21) AML patients [t(8;21)#1-CD34⁺ cells from the peripheral blood of a t(8;21) patient, t(8;21)#2-CD34⁺ cells from the bone marrow of a t(8;21) AML patient after relapse], KG1, HL-60 [myeloid cell lines without t(8;21) translocation], HL-60+PMA (HL-60 cell line differentiated to macrophage like cells by treatment with PMA) and HeLa (epithelial cell line). The y-axis represents average relative PAX5 expression from 3 independent experiments; the error bars represent SD between 3 independent experiments. For the patient samples, the error bars represent variability between qPCR measurements. (B-C) Schematic diagram showing the mammalian sequence conservation from the UCSC genome browser. The black bars represent the positions of the DHSs identified at the (B) PAX5 promoter region and (C) enhancer, respectively, in Nalm-6, Kasumi-1, SKNO-1, HL-60, KG-1, and HeLa cell lines. (D-E) DNase I footprinting experiments examining the distal promoter element HS 8 showing that the transcription factor binding pattern and the chromatin fine structure at HS 8 is identical in Nalm-6, t(8;21) AML cells and HL-60. In panel D, samples from the different cell lines digested with different concentrations of DNase I were separated by electrophoresis on the same gel; thereafter, the samples showing equal DNAse I digestion (as shown by the digestion pattern at the TBP locus) were digitally cut and realigned to prepare the figure. PMA, phorbol myristate acetate.