Regulation of Cellular Survival, Size and Metabolism

描述(申请人提供)：癌症的发展是由于不受控制的细胞增殖和程序性细胞死亡诱导的缺陷。已被证明可以防止细胞死亡的两类基因是促进生存的Bcl-2家族成员和丝氨酸-苏氨酸激酶Akt。之前对Akt的研究表明，以Akt依赖的方式生存的细胞的特征是细胞大小增加，依赖于细胞代谢的增加。Akt控制细胞大小和新陈代谢的分子机制尚不清楚。结节性硬化症基因TSC2是Akt的潜在底物，当与其结合伙伴TSC1一起表达时，已被证明调节细胞大小。在这项提案中，将进行实验以确定Akt依赖的细胞大小、新陈代谢和存活增加的分子靶点。将评估Hamartin和Tuberin的表达对这些相同过程的影响，以及TSC2是Akt底物的可能性。 具体目标： 1.确定Akt在控制细胞大小和存活方面的主要下游靶点。 A.确定Akt激活后在造血细胞中诱导的主要磷蛋白。 B.确定Akt的哪些主要下游靶点在缺乏生长因子的情况下传递维持细胞活力、新陈代谢和大小的信号。 C.表征Akt诱导的底物降解。 2.检测结节性硬化症基因在调节细胞大小和造血细胞存活中的作用。 A.表征生长因子停用后造血细胞中TSC1和TSC2表达的变化。 B.表达TSC1和TSC2在对照组、表达Bclxl和激活Akt的细胞中，测量细胞周期、大小、活力和代谢在有或没有生长因子的情况下的变化。 C.确定TSC1和TSC2是否参与Akt对细胞大小或存活的影响。测试可能的Akt磷酸化位点在tuberin中的重要性。 总之，这些研究将有助于确定原癌基因Akt和肿瘤抑制基因TSC1和TSC2在与这些基因相关的癌症细胞代谢放松调控中的作用。

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.