The overall goals of this project are to elucidate regulatory mechanisms of ErbB signaling components and to explore these molecules as molecular targets for cancer therapy. SHP2 protein tyrosine phosphatase (PTPase) mediates ErbB-induced Erk mitogen-activated protein (MAP) kinase activation and is involved in cytoskeletal reorganization and cell motility. However, how SHP2 mediates these responses is not well understood. It has been shown that Gab1 is a key SHP2 regulator in epidermal growth factor (EGF)-stimulated cells and that Src and Ras are activated by SHP2, but the mechanisms by which SHP2 activates Src and Ras have not been defined. Two models for SHP2-mediated Src activation are postulated and will be evaluated in Specific Aim I. The involvement of Src in EGF-induced Ras and Erk activation also will be analyzed.
In Specific Aim II, the novel small interfering RNA (siRNA) technique for gene knockdown will be developed and used to analyze the role of Gab1, Gab2, and SHP2 in EGF-induced cellular responses. These include Src and Akt/PKB activation, temporal regulation of Ras-MAP kinase activation, gene expression, paxillin dephosphorylation, and cell growth and metastatic properties. Based on our preliminary observations, we postulate that Gab1 and related pleckstrin-homology (PH) domains are novel molecular targets for antagonizing ErbB signaling and PTEN mutation in human cancers. This hypothesis will be evaluated in Specific Aim III using doxycycline-inducible decoy constructs in stable cancer cell lines in vitro and in tumor xenografts. These studies will define a major ErbB and SHP2 signaling mechanism, greatly advance our understanding of the function of Gab proteins, uncover novel targets for molecular intervention of malignant signaling, and stimulate further development of new inhibitors targeting these signaling components for cancer treatment. ? ? ?
