Fig. 5

Bag-1 forms complexes with Akt and C-Raf in breast cells. a. MCF-7, MDA-MB-231 and MCF-10A cell lysates were immunoprecipitated (IP) with anti-Bag-1 antibody, and immunocomplexes were analyzed by immunoblots (IB) using Bag-1, C-Raf, phospho-C-Raf^Ser338, phospho-B-Raf^Ser445 Akt and phospho-Akt^Ser473 antibodies. Phospho-Bad^Ser136 and A-Raf antibodies were used as negative controls. b. Normal (N) and tumor (T) tissue extracts were IPed with anti-Bag-1 antibody, and immunoblots were performed using p-Akt, p-C-Raf and p-B-Raf antibodies. c. PRISM predictions for the complexes between Bag-1 and its interaction patners, and models for their cell survival promotion mechanism. Binding energy scores (BES) for BAG/C-Raf, BAG/C-Raf/B-Raf and BAG/Akt complexes were predicted as −30.11, −20.84, − 14.08, respectively. Bag-1 binding to C-Raf and/or B-Raf kinases activates them, which in turn phosphorylate Bad (at S112) and other downstream effectors, such as MAPK. Bag-1 binding to Akt is essential for Bad phosphorylation at S136 by the activated Akt kinase. Phosphorylated Bad inhibits apoptosis