The guanine nucleotide binding protein Ras is an important regulatory-switch in cell growth control, and Ras is often mutated in tumors. Its activation is achieved by the exchange of bound GDP for GTP, catalyzed by Sos. Targeting of Sos to the membranes for Ras activation is mediated through the adaptor protein Grb2, which is recruited to the membranes upon growth factor stimulation. Crb2 binds the prolinerich sequences of Sos via its SH3 domains. Additional studies indicate that complex mechanisms exist for Ras regulation. It is reasoned that when these control mechanisms are lost due to mutation or inactivation of regulatory molecules, even without an intrinsic Ras mutation, cancers arise frog uncontrolled cell proliferation. Based on our preliminary studies, we propose that Dab2 is such a Ras regulator. Dab2 is a 96 kd, mitogen-responsive phosphoprotein that we have recently identified and doned. We have found that Dab2 associates in vivo and in vitro with Grb2 and competes with Sos. Additionally, we found that Dab2 exhibits growth regulatory/suppressive activity. These preliminary studies have led us to hypothesize:that Dab2 stimulates the dissociation of the Grb2/Sos complex and sequesters Grb2 from Sos. In doing so, Dab2 may negatively regulate Ras activation, therefore restrain cell growth. The relevance of Dab2 in cancer is also suggested by the finding that the expression of Dab2 is lost in many breast and ovarian carcinoma cells and tissues examined to date, but is present in normal cells and tissues. We propose to study the cellular functions and growth regulatory mechanisms of Dab2 based on our hypothesis. Specifically, we will analyze the binding of Dab2 to Grb2 and its competition with Sos (Aim 1), and the effects of Dab2 on Ras activation, cell growth and transformation (Aim 2). We will also investigate the functional sigttificance of Dab2 phosphorylation (Aim 3). - Moreover, we will identify additional proteins that physically interact with Dab2 (Aim 4) in order to understand further its function and regulation. These mechanistic studies of Dab2 will enhance our understanding of its role in cell growth regulation and the possible involvement in breast and ovary tumorigenicity.
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