Effect of PIM1, PIM2 siRNAs on PIM1, PIM2 and MCL1 mRNA and Pim kinase targets in MCL cell lines. Effects of PIM1, PIM2 siRNAs and their combination on PIM1, PIM2 and MCL1 mRNA expressions in JeKo-1 (A) and Mino (B) cells. Cells were treated with 150nM scramble, siGLO, PIM1, PIM2 siRNA as well as PIM1 and PIM2 siRNA combination (150nM for each) for 24 hours. Then, RNA was extracted and analyzed using RT-PCR. (C) Effects of PIM1, PIM2 siRNA and their combination on total and phospho–c-Myc (Ser62), total and phospho–4E-BP1 (Thr37/46), as well as Mcl-1 protein expression were measured by immunoblots. (D) Quantitation of immunoblots in cells with siRNA treatments. Phospho- and total protein ratios or Mcl-1-to-GAPDH ratios were calculated, and data are presented as means of 3 independent experiments ± SEM. Percentage of cell death is marked under each lane of immunoblot measured by annexin V and PI positivity.