407a Oncogenic AKT1 Regulation by Tumor Suppressor KLF11: Implications in Growth Regulation During Neoplastic Transformation in Barrett's Esophagus and Interference by Carcinogenic Bile Acid Dependent Erk2 and PKCa Activation

Abstract

Pancreatic adenocarcinoma is the fifth leading cause of cancer death, with an abysmal 5year survival rate of less than 5%, due in large part to the high incidence of metastasis. Metastatic invasion represents a complex synergy of cell motility, adhesion, and matrix remodeling. How these multiple processes are coordinated during dissemination is a central focus of tumor cell biology. The small oncogenic G protein Rac1 is a major regulator of cell motility, and it is activated through the action of guanine nucleotide exchange factors (GEFs). A critical GEF in tumor cell biology is the proto-oncogene Vav1, which is ectopically expressed in most pancreatic tumors to cause poor prognosis in patients and enhanced tumor growth In Vitro. Vav1 interacts directly with the large GTPase Dynamin 2 (Dyn2), though the function of this interaction is not known. Importantly, Dyn2 is also markedly upregulated in pancreatic cancer, is a potent activator of metastatic migration, and is required for Rac1-mediated formation of lamellipodia. We have recently found that Dyn2 binds Vav1 directly in pancreatic tumor cells. Therefore, the GOAL of this study was to test if the Dyn2Vav1 interaction could promote the activation of Rac1 and the invasive migration of pancreatic tumor cells. RESULTS: We found that disruption of the Dyn2-Vav1 interaction impairs Rac1 activation and subsequent lamellipodia formation and cell migration, indicating that Dyn2-Vav1 binding regulates tumor cell motility. Surprisingly, RNAi-mediated suppression of Dyn2 or disruption of Dyn2-Vav1 binding results in a dramatic reduction in Vav1 protein stability, resulting in diminished levels of this proto-oncogene and decreased Rac1 activation and migration. Remarkably, this degradative reduction is a result of Vav1 targeted transport to the lysosome, as treatment of these cells with the protease inhibitor chloroquine prevents Vav1 loss, and immunofluorescent cell staining shows a dramatic localization of Vav1 to the lysosomal lumen when it cannot bind Dyn2.CONCLUSION: These findings demonstrate a novel regulatory role for direct Dyn2 binding in Rac1 activation, lamellipod protrusion and cellular migration through the stabilization of Vav1, and demonstrate a potent mechanism by which metastatic migration is upregulated in pancreatic tumor cells. This study was funded by NCI CA104125 to MAM.