Bcl-xL and Bcl-w overexpression protects against ABT-737 induced cell death. (A) WT fetal liver cells retrovirally transduced with pMIG–Bcl-2, pMIG–Bcl-xL, pMIG–Bcl-w, pMIG-Mcl1, or control pMIG vectors were transplanted into lethally irradiated recipient mice (n = 5 per group). Lymphocytes were harvested 8 weeks later and treated in culture with ABT-737. The reminder of the results are shown as a heat map in supplemental Figure 3A. (B) Overexpression of Bcl-2, Bcl-xL, Bcl-w, or Mcl-1 in lymphocytes is accompanied by an increase in Bim as assessed by Western blotting. For a reason that is unclear, the Flag-tagged version of Mcl-1 is very difficult to detect with our anti-Flag antibody, but is detected with the anti–Mcl-1 antibody as a slower moving band. (C) Murine DO11.10 T hybridoma cells stably transfected with pEF–mBcl-2, pEF–mBcl-xL or control pEF vectors were challenged with increasing concentrations of ABT-737 for 24 hours. (D) Increase in Bim levels on overexpression of Bcl-2 or Bcl-xL in DO11.10 cells as revealed by Western blotting. (E) Mcl-1fl/fl/Eμ-Myc lymphoma cells retrovirally transduced with pMIG–Bcl-2, pMIG–Bcl-xL, pMIG–Mcl-1 or control pMIG vectors were treated for 3 days with 4-OHT to remove endogenous Mcl-1 before being exposed to ABT-737. These experiments have been reproduced using an independent Mcl-1fl/fl/Eμ-Myc lymphoma line. (F) The corresponding levels of Mcl-1, Bcl-2, Bcl-xL, and Bim were determined by Western blotting.
