p27 levels are regulated by SKP2 in BCR-ABL-positive cell lines. (A) Mo7ep210^BCR-ABL cells were infected with lentivirus-producing SKP2 shRNA-1 and -2 or control (scramble) shRNA construct for stable knockdown of SKP2. The effect on cell-cycle distribution (PI staining) as well as on SKP2, p27, and pT187p27 expression was analyzed by immunoblot analysis. The differences in cell-cycle progression after knockdown of SKP2 are significant compared with control with P values less than .001 from 3 independent experiments. (B) SKP2 mRNA levels were assessed in Mo7e and Mo7ep210^BCR-ABL cells by quantitative RT-PCR after treatment with 2.5 μM imatinib for 16 and 24 hours. (C) Ba/F3 cells expressing p210^BCR-ABL were stably transduced with a Skp2 expression vector and treated with 2.5 μM imatinib for 16 hours. Cell-cycle distribution was analyzed by PI staining in 3 independent experiments, and (D) p27 expression was analyzed by immunoblot. (E) Effect of BCR-ABL kinase inhibition on the expression of SKP2 substrates was analyzed. Cells were treated with 2.5 μM imatinib and the expression of p130, p57, Tob1, and p21 was measured by immunoblot analysis in Mo7ep210^BCR-ABL and Ba/F3p210^BCR-ABL cells. Error bars represent SE.