Myc and Pim proteins are required for full transformation of HEL cells. (A) HEL cells were transfected with either control siRNA or c-Myc–, Pim1-, and Pim2-specific siRNA. Relative growth of HEL cells 72 hours after siRNA transfection was calculated as a percentage compared with control siRNA–transfected cells (n = 3, left panel). The change in expression of c-Myc, Pim1, and Pim2 after siRNA treatment was determined in whole-cell lysates (right panel). *Significant differences (P ≤ .05) were observed between treated and control cells. (B) Untreated BaF3 cells with doxycycline-inducible Pim1 and Pim2 were used and compared with cells treated with 1 μg/mL doxycycline (DOX) or BaF3 cells maintained in interleukin-3 (IL-3)–free medium. Cell growth was measured after 72 hours (top panel) or induction of Pim1 and Pim2 after doxycycline treatment (24 hours) by immunoblotting as indicated (bottom panel). (C) Parental BaF3 cells or cells expressing the MycER fusion protein were left untreated or treated with 4-hydroxy-tamoxifen (HT). The expression of the MycER fusion protein was detected by Myc immunoblotting (bottom panel). (D) Untreated BaF3 cells with doxycycline-inducible Pim1 and Pim2 as well as HT-inducible Myc-ER were used and compared with cells treated with 1 μg/mL DOX, HT, or combinations thereof (0.2 × 106 cells/mL). Cell concentrations were measured over a 3-day period (n = 3) and protein expression was assayed after 24 hours of DOX and HT treatment as indicated.
