Regulation of Cellular Survival, Size and Metabolism

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DESCRIPTION (provided by applicant): Cancer develops as a result of uncontrolled cell proliferation coupled with defects in the induction of programmed cell death. Two classes of genes that have been shown to prevent cell death are the pro-survival Bcl-2 family members, and the serine-threonine kinase Akt. Previous work with Akt has demonstrated that cells surviving in an Akt-dependent manner are characterized by increased cell size and a reliance on increased cell metabolism. The molecular mechanisms by which Akt controls cell size and metabolism are unclear. The tuberous sclerosis gene TSC2 is a potential substrate of Akt, and has been shown to regulate cell size when expressed together with its binding partner, TSC1. In this proposal, experiments will be performed to determine the molecular targets for Akt-dependent increases in cell size, metabolism, and survival. The effects of the expression of hamartin and tuberin on these same processes will be assessed, as well as the possibility that TSC2 is an Akt substrate. Specific aims: 1. Identify the major downstream targets of Akt in the control of cell size and survival. A. Determine the major phosphoproteins induced in hematopoietic cells by activation of Akt. B. Determine which of the major downstream targets of Akt transduce signals that maintain cellular viability, metabolism and size in the absence of growth factor. C. Characterize Akt-induced degradation of its substrates. 2. Test the role of the tuberous sclerosis genes in regulating cell size and survival in hematopoietic cells. A. Characterize changes in TSC1 and TSC2 expression in hematopoietic cells upon growth factor withdrawal. B. Express TSC1 and TSC2 in control, Bcl-xL- and activated Akt-expressing cells, and measure changes in cell cycle, size, viability and metabolism in the presence and absence of growth factor. C. Determine if TSC1 and TSC2 contribute to Akt effects on cell size or survival. Test the importance of the potential Akt phosphorylation sites in tuberin. Together, these studies will help define the roles of the proto-oncogene Akt and the tumor suppressor genes TSC1 and TSC2 in the deregulation of cell metabolism in cancers associated with these genes.