Our long-term goals are to understand how signaling pathways regulated by oncogenes interact with each other to transform cells. Ras is one of the most common proto-oncogenes found mutated in tumors. When Ras transforms cells, it alters several basic properties of the cell by modifying key transcription factors to promote cell proliferation, rearranging the actin cytoskeleton to stimulate invasion and inhibiting apoptosis to promote survival. Thus, studying the signals from oncogenes to their multiple targets may provide insights into how tumor growth is coordinated. Many effects of Ras, both proliferative and cytoskeletal, are regulated through the coordinated actions of the Rho family of G proteins, including members such as, Rac, Rho and Cdc42. However, the signals beyond these small G proteins that mediate Ras transformation remain to be elucidated. Rac and Cdc42 bind several proteins, including members of a highly conserved family of protein kinases known as Paks. The Pak kinases are candidates for downstream effectors that regulate both transcription and the cytoskeleton. Our preliminary results demonstrate, for the first time, a direct link between Pak and cell transformation. We found that Ras transformation can be inhibited by expression of specific Pak mutants. This type of mutation, usually called a dominant negative mutation, provides a powerful tool to explore the role of Pak in Ras transformation. Furthermore, we traced a signal from Ras through Pak to the downstream kinase, Erk. We also developed several assay systems showing for the first time that Ras activates Pak through a linear pathway requiring PT 3-kinase and a small G protein (Rac or Cdc42). Unexpectedly, one of the key intermediates is the Akt proto-oncogene, which transduces cell survival signals through Pak to a cell survival factor called Bad. This signal inhibits apoptosis. Together, these data demonstrated a new Ras signaling pathway and provided the first evidence for transformation signals through a specific effector of the Rho family. This proposal aims to build on our earlier work by tracing the signals step by step from Ras to Pak and linking Pak to another oncogene, Abl. We hypothesize that Pak is a convergence point for Ras and Abl to transduce signals to transformation and survival pathways. We will: (1) Determine the signals from Ras to Pak. (2) Determine the role of Pak in cell survival signals through Bad. (3) Determine the role of the RasPak module in signaling by the Abl oncogene.
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