Identification of the potential downstream effectors of the mir-17–92 cluster in erythroleukemia cells. (A) The expression of phospho-Akt and phospho-Erk was determined by Western blot in HB60–5 cells infected with either Fli-3-mir-17–92 or empty vector in response to Epo stimulation. Cells were grown without growth factors for 6 hours and then stimulated with Epo (1 U/mL) for 10 minutes. The controls are HB60–5 cells cultured in media containing Epo and SCF. The intensity of HB60–5 band was used as control (1.0). (B) Expression analysis of Spi-1, Fli-1, and p27 protein in a panel of erythroleukemia cells. The indicated cells were lysed and Western blotted with Spi-1–, Fli-1–, or p27-specific antibodies. The intensity of HB60–5 band was used as control (1.0). (C) Expression analysis of p27 protein in HB60–5 cells overexpressing Fli-1. The blot was stripped and rehybridized with β-actin antibody to demonstrate the equal loading. The intensity of HB60–5 band was used as control (1.0). Mean fold changes are shown at the right side.