Figure 6.
Endogenous Bim forms high-MW complexes together with Mcl-1 but not with Bcl-2 or Bcl-XL. (A–C) Complexes of endogenous Bcl-2 family proteins on mitochondria. BN-PAGE was performed as described earlier with solubilized mitochondrial fractions of the indicated cell lines. In order to achieve good detection, the amounts of proteins loaded and exposure times were sometimes different for different cell lines. Bcl-XL (except in 293FT, where a small amount of Bcl-XL ran at high-MW complexes) and Bcl-2 ran as low-MW proteins or small complexes, while Mcl-1 ran at high MW. Data are representative of at least three independent experiments with each cell line. We were unable to detect Bcl-2 in the HCC827 and Caco2 cell lines. (D) The majority of endogenous Bim is bound to MCL-1. Co-IP experiments were performed with whole-cell lysates from MCF-7, HeLa, and 293FT cells using anti-Bim antibody. The Bim-interacting partners were detected by Western blotting. (Isotype) Immunoprecipitation control with an isotype antibody of unrelated specificity; (*) antibody light chain. Data are representative of three independent experiments.