Probing a novel signaling complex that sustains AKT activation to support stress survival in cancer

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PROJECT SUMMARY Abnormal metabolism is a hallmark of cancer that helps cancer cells to grow, undergo malignant transformation, and survive under stressful conditions such as nutrient deprivation. Cancer cells are exposed to many cellular stresses during tumorigenesis, which must be overcome for the propagation of malignancy. In cancer, the abnormal activation of many signaling networks serves to disconnect the control of growth, metabolism, and survival, and recent efforts have sought to therapeutically target cancer metabolism. The phosphatidylinositol 3- kinase (PI3K)-AKT (protein kinase B) signaling pathway is the most activated in human cancer and has a wide range of effects on cellular metabolism. We have recently identified the Cdc42/Rac guanine nucleotide exchange factor (GEF) dedicator of cytokinesis 7 (Dock7) as a novel signaling node that supports sustained basal AKT activation and mechanistic target of rapamycin (mTOR) activity as determined by its downstream target S6 kinase (S6K) during stressful conditions to maintain signaling activity required for cell survival and transformation. We find that Dock7 is required for multiple cancer cell lines to resist anoikis and exhibit anchorage-independent growth. While we observe relatively low levels of AKT phosphorylation compared to stimulation by growth factors, Dock7-dependent signaling is critical for the survival of cancer cells during nutrient deprivation. I hypothesize that under cellular stress Dock7 serves as a scaffold for AKT, sustaining its phosphorylation and organizing signaling partners for mTOR signaling required for stress survival. This project will investigate the role of this novel Dock7/AKT/mTOR signaling activity in providing a survival benefit to cancer cells under cellular stress. I propose to study the impact of Dock7-dependent signaling activity on AKT/mTOR signaling, cell survival under stress, and critical characteristics of malignant progression and aggression. In Aim 1, I will investigate the functional activities of the Dock-homology region 2 (DHR2) domain of Dock7, which is responsible for GEF activity, in basal AKT phosphorylation for cancer cell stress survival and malignant transformation. In Aim 2, I will next identify the novel role of the DHR1 domain in Dock7-dependent AKT phosphorylation, cancer cell stress survival, and malignant transformation. Then, in Aim 3, I will identify the subcellular location of this Dock7 signaling complex under stress conditions and determine the individual roles of DHR1, DHR2, and activated Cdc42 in Dock7 localization. The work in this proposal will provide biochemical characterization of Dock7 signaling activity that will lead to a mechanistic understanding of Dock7-dependent AKT/mTOR activation in cancer cell stress survival. These findings will not only contribute to the understanding of cancer aggression and metabolism but may also identify new therapeutic targets for cancer treatment.