A variety of human diseases result from improper regulation of cell growth (an increase in cell mass and size), proliferation, survival and death, differentiation and development. Phosphatidylinositol 3-kinase (PI3K), a mediator of mitogenic signals via production of lipid second messengers, plays a critical role in regulating these processes. Indeed, approximately 30% of human cancers have loss of function mutations in PTEN, a critical suppressor of PI3K signaling. The mammalian target of rapamycin, mTOR, integrates both mitogenic and nutrient signals, and its disregulation leads to the formation of benign tumors associated with the disease, tuberous sclerosis, mTOR is a protein kinase, a scaffold, and a critical mediator of p70-S6 kinase (S6K1) activation and suppressor of the translational inhibitor, 4E-BPI. Activation of S6K1 is very complex, requiring multiple distinct PI3K- and mTOR-dependent inputs. Indeed, S6K1 was the first protein kinase shown to act downstream of PI3K and mTOR and is completely inhibited by the immunosuppressant drug rapamycin. Underscoring the importance of S6K1 in tumor formation, the oncogene products Dbl, TIAM-1 and Akt have been shown to contribute to its activation. In addition to being a potent immunosuppressant, rapamycin also prevents restenosis after angioplasty and is in clinical trials as an anticancer drug due to its anti-proliferative effects. Thus, it is clear that understanding how the PI3K/S6K1 and mTOR/S6K1 pathways are regulated and signal to downstream effectors to regulate cell growth and proliferation is of significant importance with regards to human disease. The first objective is to define further the nature of the S6K1 signaling complex. Part of the focus will be on how mTOR regulates S6K1, but the screens described should lead to the identification and characterization of all S6Kl-associated proteins. The second objective will focus on how Rheb, a GTPase recently shown to be a positive modulator of mTOR, is regulated and how it regulates mTOR signaling to S6K1, and 4E-BPI. The third objective of this proposal is to characterize a newly identified S6Kl-specific interactor and target, SKAR, and determine how it contributes to S6K1 signaling and function. The final objective will continue our characterization of how the mTOR targets, S6K1 and 4EBPI/ elF4E, regulate cell growth and cell cycle progression.
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