RUNX1 is ubiquitously expressed in human T-ALL cells, and RUNX1 or CBFβ knockdown results in apoptosis. (A) Protein was isolated from human T-ALL cell lines and RUNX1, RUNX3, CBFβ, TAL1, MYB, NOTCH1, and MYC protein levels were determined by immunoblotting. Extracellular signal–regulated kinase 1/2 (ERK1/2) was used as a loading control. (B) The human T-ALL cell line Jurkat was infected with lentiviruses expressing a control shRNA or 2 shRNAs specific for RUNX1. RUNX1 mRNA and protein levels were examined by qRT-PCR and immunoblotting. (C) RUNX1 knockdown results in leukemic cell apoptosis. Control (GFP) and RUNX1 shRNA-transduced Jurkat cells were stained with Annexin V-FITC and 7AAD and analyzed by flow cytometry 6 days after infection. A representative flow profile is shown (left). The percentage of apoptotic cells was determined by Annexin V/7AAD staining and analyzed by flow cytometry. Four independent experiments were performed, and data are shown as means ± SD (right). (D) CBFβ knockdown also induces apoptosis. Control (GFP) or CBFβ shRNA-transduced Jurkat cells were stained with Annexin V-FITC and 7AAD and analyzed by flow cytometry. Four independent experiments were performed, and data are shown as means ± SD (right). (E) CBFβ protein levels in control and knockdown cells were analyzed by immunoblotting. **P < .005; ***P < .0005; ****P < .0001, one-way ANOVA multiple comparisons test.