Activated α2-macroglobulin binding to cell surface GRP78 induces T-loop phosphorylation of Akt1 by PDK1 in association with Raptor.

PDK1 phosphorylates multiple substrates including Akt by PIP3-dependent mechanisms. In this report we provide evidence that in prostate cancer cells stimulated with activated α2-macroglobulin (α2M*) PDK1 phosphorylates Akt in the T-loop at Thr(308) by using Raptor in the mTORC1 complex as a scaffold protein. First we demonstrate that PDK1, Raptor, ...
and mTOR co-immunoprecipitate. Silencing the expression, not only of PDK1, but also Raptor by RNAi nearly abolished Akt phosphorylation at Akt(Thr308) in Raptor-immunoprecipitates of α2M*-stimulated prostate cancer cells. Immunodepleting Raptor or PDK from cell lysates of cells treated with α2M* drastically reduced Akt phosphorylation at Thr(308), which was recovered by adding the supernatant of Raptor- or PDK1-depleted cell lysates, respectively. Studies of insulin binding to its receptor on prostate cancer cells yielded similar results. We thus demonstrate that phosphorylating the T-loop Akt residue Thr(308) by PDK1 requires Raptor of the mTORC1 complex as a platform or scaffold protein.
Mesh Terms:
Adaptor Proteins, Signal Transducing, Animals, Cell Extracts, Cell Line, Tumor, Cell Membrane, Heat-Shock Proteins, Humans, Immunoprecipitation, Male, Mice, Nude, Models, Biological, Phosphorylation, Phosphothreonine, Protein Binding, Protein-Serine-Threonine Kinases, Proto-Oncogene Proteins c-akt, RNA, Double-Stranded, Transfection, alpha-Macroglobulins
PLoS ONE
Date: Feb. 12, 2014
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