The two primary goals of our previous proposal were to determine the contribution of multiple Ras-mediated effector signaling pathways to transformation and to determine whether different Ras signaling events are important for oncogenic Ras transformation of fibroblasts versus epithelial cells. In our continuation of these studies, we extend these analyses to emphasize two additional themes. First, we will evaluate the role of a novel effector pathway in Ras transformation We have identified Tiami as a novel effector of Ras. Tiami in turn can activation the Rac small GTPase. Rac has been shown to mediate Ras regulation of the p38 and JNK MAPK pathways. Therefore, we will assess the roles of these MAPK cascades as key components of Raf-independent signaling pathways important for Ras transformation. Second, the least understood aspect of Ras signaling involves the gene targets important for Ras-mediated oncogenesis. We describe studies to begin a delineation of the genes whose expression is deregulated by oncogenic Ras.
Four specific aims are proposed to accomplish these goals. First, we will characterize the role of Tiami as a critical effector of Ras transformation, and whether Tiami links Ras with Rac and p38/JNK signaling. Second, we will evaluate the distinct and opposing roles of the p38 and JNK MAPK cascades in Ras transformation, and determine whether these MAPKs mediate Ras regulation of gene expression. Third, we will determine the role of specific Ras-mediated signaling pathways involved in transformation of ROSE rat ovarian epithelial cells and define the gene targets that promote Ras transformation. Finally, we will employ microarray analyses to delineate the gene targets important for Ras transformation (in cooperation with telomerase and SV4O T antigen) of primary human embryonic kidney and mammary epithelial cells. These studies will evaluate our hypothesis that oncogenic Ras transformation of epithelial cells is critically dependent on activation of Raf-independent effector signaling pathways, in part, by deregulating the expression and function of key target genes.
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