Akt interacts with Bcl-w.

<p>(A) Co-immunoprecipitation of endogenous Akt with Bcl-w. Wt HeLa cells were lysed and 1 mg of protein extract immunoprecipitated using an anti-Bcl-w antibody. Immunoprecipitates and total lysates (50 µg) were separated on 12%SDS polyacrilamide gel and blotted with an anti-Akt antibody. As negative control, proteins were incubated with beads without antibody (B) Co-immunoprecipitation of transfected Akt with FLAG-Bcl-w or EE-BAD. HEK-293 cells were tansfected with 2 µg of HA-Akt and 2 µg of FLAG-Bcl-w or EE-BAD cDNAs, as indicated. After 48 hr, cells were lysed, and 1 mg of protein extract was immunoprecipitated using an anti-HA antibody. Immunoprecipitates were subsequently blotted with anti-HA, anti-Flag or anti-EE antibodies, as indicated. (C) HEK-293 cells were transfected with 2 µg of either wt-Bcl-w cDNA or the deletion mutants, Bcl-w/BH4 or Bcl-w/CT, as indicated. Protein extracts were immunoprecipitated using an anti-Akt antibody. Immunoprecipitates and total lysates were resolved on 12%SDS-PAGE and transferred to Hybond-C nitrocellulose. Membranes were incubated with an anti-FLAG antibody. Both deletion mutants, Bclw/BH4 and Bclw/CT, immunoprecipitated with Akt.