Regulation of Ras signaling by the ubiquitin pathway

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DESCRIPTION (provided by applicant): Signaling through the well-known oncogene Ras can promote proliferation, cell survival, and differentiation and is implicated in cancer. Mutations that increase signaling through this pathway are found frequently in cancer and also occur in developmental disorders such as Noonan's Syndrome. Thus, it is vital to understand how Ras is regulated. Much attention has focused on understanding activation of this pathway at upstream steps, such as the binding of ligands to Receptor Tyrosine Kinases (RTKs), which act through other proteins to recruit and activate Ras. Recently, modification of Ras by a process called ubiquitination has been reported in mammalian cells. My research group now has data confirming the ubiquitination of Ras using Drosophila (fruit flies). This suggests that this newly discovered regulation of Ras is highly conserved. Moreover, using Drosophila (fruit flies) we generated compelling evidence that the ubiquitination of Ras has profound effects on growth, proliferation and cell survival. The powerful system of Drosophila genetics and the well-established paradigm of studying Ras in Drosophila make this an excellent system to address and characterize the significance of Ras ubiquitination in a multi-cellular context. Furthermore, an in vivo Drosophila approach has the potential to make significant and unique discoveries that would not be possible in other systems. The goal of this proposal is to use Drosophila to understand the biological role of Ras ubiquitination. Our hypothesis is that ubiquitination of Ras restricts its ability to promote growth and proliferation. Our hypothesis is based on our Drosophila findings that (1) Drosophila Ras is ubiquitinated, (2) impairing ubiquitination promotes increased signaling through Ras, and (3) impairing ubiquitination promotes Ras-dependent overgrowth, increased, proliferation, and cell death resistance that appear independent of steps upstream of Ras. The novelty of our findings is that Ras may be a distinct step in the pathway whose ubiquitination is crucial to ensure proper control of growth and proliferation. My research group will exploit our expertise in studying the ubiquitin pathway and in utilizing Drosophila to most efficiently attack this problem. The proposed studies perfectly complement ongoing molecular and tissue culture studies being performed elsewhere. In the short term, our studies could establish the biological significance of this novel regulation of Ras. Over the longer term, our work could contribute tremendously towards a better understanding of the Ras oncogene and may even identify therapeutic targets. Therefore, we believe that these studies could have profound implications for cancer research. We propose research according to the following Aims: Specific Aim 1: Establish the biological significance of Ras ubiquitination. Specific Aim 2: Examine Ras ubiquitination in vivo. Specific Aim 3: Identify and characterize the Ras E3.