CD45 is down-regulated in AE9a leukemia mice and t(8;21) AML patients. (A) A schematic showing part of the mouse Cd45 gene. The positions of the predicted transcription initiation site (arrow) and consensus AML1 sites (asterisks) are indicated. Numbers indicate the positions relative to the transcription initiation site (+1). E4: exon 4. ChIP(a), (b), (c), and (d) are the regions analyzed by qPCR in panel B. (B) Quantitative PCR analysis of the regions containing the consensus AML1 sites indicated in panel A after anti-HA and anti-AML1 ChIP assays. The y-axis shows the relative enrichment level over an arbitrary negative control region (see “ChIP and ChIP-chip assays” and the confirmation section of the supplemental Methods). Input DNA was used for normalization. (C) Flow cytometric analysis of CD45 in EML cells expressing AE9a, AE9a-R174Q, and MIP. (D) Flow cytometric analysis of CD45 within the LK population of wild-type bone marrow and AE9a leukemia blood, spleen, and bone marrow. CD45 expression of blood, spleen, and bone marrow samples from AE9a leukemia mice were compared with that of bone marrow cells from wild-type mice. Representative results from 3 leukemia and 3 wild-type mice are shown. (E) RT-qPCR analysis of CD45 expression relative to GAPDH of each individual t(8;21) AML-M2 (n = 17, dot) and non-t(8,21) AML-M2 (n = 15, square) patient. The bar indicates the mean expression level of each patient group. The P value was determined using Student t test. (F) Flow cytometric analysis of CD45 within the primitive stem/progenitor (CD33⁻/CD38⁻/CD71⁻/CD34⁺) population of t(8;21) AML-M2 blood samples (n = 5), non-t(8,21) AML-M2 blood samples (n = 2), and normal bone marrow samples (n = 2). (Left panel) Representative histograms from each patient group are shown. (Right panel) A dot plot showing the CD45 MFI of each patient. The bar indicates the mean value of each patient group. The P values were determined using Student t test.