INTEGRATION OF ONCOGENIC SIGNALS BY THE NUCLEOLAR ARF-NPM NETWORK

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ABSTRACT Initially described as an organelle contained within specific chromosomal regions, the nucleolus has advanced through the past century imparting much knowledge about cell and cancer biology along the way. Once hindered with the label as being the static center of ribosome biogenesis, a newfound appreciation for this visible organelle has been established in recent years. One of the most dynamic occupants of the mammalian nucleolus is the ARF tumor suppressor protein. Since its discovery, the mechanism behind ARF's tumor suppressive function has been under intense investigation. Initially touted as a bona fide inhibitor of the p53 negative regulator, Mdm2, more recent studies have identified a p53-independent role for ARF in suppressing tumor formation in mice and humans. Much of the more recent work has focused on the novel localization of ARF in the nucleolus, a seemingly benign organelle in terms of transformation properties. One of the least understood aspects of ARF biology is what is ARF doing in the nucleolus. Numerous labs, including ours, have identified the nucleophosmin (NPM) proto-oncogene as a nucleolar binding partner of ARF. Our preliminary data demonstrates that hypergrowth signals emanating from the loss of the Tsc1 tumor suppressor stimulate the protein expression of both NPM and ARF in an effort to influence ribosome synthesis rates. Additionally, basal nucleolar ARF proteins interact with a pool of NPM in the nucleolus and acute loss of these basal ARF molecules results in dysregulated NPM function and increased ribosome biogenesis. Loss of nucleolar ARF alters the landscape of the nucleolus, allowing for proteins, such as the p68 RNA helicase, to interact more readily with NPM to potentially promote ribosome biogenesis and cell transformation. We hypothesize that ARF resides in the nucleolus to tightly regulate ribosome biogenesis through the sensing of hypergrowth signals and to halt unwarranted NPM-p68 activities. Based on our preliminary data and our stated hypothesis, three specific aims are proposed: 1) Determine how oncogenic growth signals are interpreted by the nucleolar ARF-NPM complex, 2) Establish the physical and functional interaction of the p68 RNA helicase with NPM, and 3) Determine the in vitro and in vivo ability of ARF to act as a nucleolar checkpoint protein. PROJECT NARRATIVE The ARF tumor suppressor is the second most commonly mutated gene in human cancers, second only to p53. We are just beginning to appreciate how this critical tumor suppressor functions to prevent unwarranted cell growth and proliferation. We seek to understand the mechanism behind ARF's ability to regulate p53-independent growth arrest in vivo and to move these findings into a more clinical setting where novel ARF-targeted therapeutics might affect a broad spectrum of cancer patients. Thus, basic research into ARF biology is appropriate given its mutational prevalence in human cancers.