Novel Functions for Ras Family GTPases

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DESCRIPTION (provided by applicant): Project Summary: Ras proteins play a central role in many aspects of biology. These monomeric GTPases cycle between an inactivate GDP-bound state and an active GTP-bound state. Mutational activation of Ras occurs in approximately 30% of human tumors and traps Ras in the GTP-bound state, leading to oncogenic transformation. Thus, RasGTP has been considered the only biologically important form of the molecule. However, accumulating evidence suggests this may not be the case. We have discovered that nucleotide-free Ras (nf-Ras), formed as an intermediate in the transition from RasGDP to RasGTP, negatively regulates phosphatidyl-inositol 3-kinase, class IIbeta (PI3KC2¿). These finding have profound implications for the understanding of Ras-mediated signaling and transformation. We propose that nf-Ras binds targets such as PI3KC2¿ and that interaction with these targets leads to the mutual inhibition of both Ras and the associated target. Our model further predicts that oncogenic activation of Ras leads to loss of repression of these targets resulting in their activation withou binding RasGTP. Thus, our findings point to a new class of molecules important for Ras-driven tumorigenesis, yet do not bind activated Ras. The goal of this proposal is to characterize this novel role of nf-Ras in cellular signaling and transformation through the following three aims. In Aim 1, we will determine the molecular basis for the interaction of nf-Ras with PI3KC2¿ using a combination of molecular and biochemical approaches to map the interaction of Ras with PI3KC2¿. In addition, we will determine the role of the intersectin scaffold in regulation of the n-Ras: PI3KC2¿ complex. In Aim 2, we will determine the fraction of Ras that resides in the nucleotide-free state in vivo. Finally, in Aim 3 we propose two approaches to identify additional targets of nf-Ras and test the importance of these targets in models of oncogenesis. Given the prevalence of Ras mutations in human cancers, it is critical to understand the mechanisms through which Ras contributes to tumorigenesis. Our work represents an entirely new concept in Ras biology that defines a new class of targets that participate in Ras-mediated signaling and transformation. These studies will provide new insights into the mechanisms of Ras-mediated tumorigenesis and are therefore of high translational significance. This work will be beneficial to Veterans as well as the general population, both of which suffer from cancer. However, the incidence for certain cancers, such as lung and pancreatic cancer the latter of which has a high incidence of Ras mutations (<90%), has been reported to be up to 5-7x higher in Veteran populations making these studies particularly relevant to Veterans.