Metabolism and Phosphatase Regulation of the TOR Pathway

描述(由申请人提供)：生长过程(大量积累)是细胞、器官和身体大小的关键决定因素，在癌症和糖尿病等疾病中经常被解除管制。我们正在研究TOR(哺乳动物中的mTOR；果蝇中的dtor)途径，这是一个保守的信号系统，正在成为真核生物生长的关键调节因子，并受到新陈代谢、生长因子和应激的调节。TOR途径是FDA批准的免疫抑制剂雷帕霉素的靶点，雷帕霉素也用于预防血管成形术后的血管再狭窄，目前正处于治疗癌症和自身免疫性疾病的试验中。在过去的几年里，我们一直在研究人类组织培养细胞中mTOR途径的生物化学。尽管这项工作取得了丰硕的成果，并导致了几种与mTOR相互作用的蛋白质(例如Raptor和GbetaL)的发现，但我们已经意识到，许多关于TOR信号转导的重要问题在使用果蝇而不是人类组织培养细胞的模型系统中更容易回答。人类和果蝇的TOR(Dtor)途径是高度保守的，我们的初步数据表明，这两条途径对营养代谢的反应相似，都含有一种未知的TOR调节的磷酸酶，并且对细胞大小有相似的影响。相比之下，酵母TOR途径缺少几个重要的组成部分，包括S6激酶(S6K)、TSC1和TSC2，它们感知不同的营养物质，并且没有生长因子输入。 在果蝇而不是人类组织培养细胞中研究TOR信号有几个优点。苍蝇途径比人类途径的冗余少，功能突变的丢失非常容易和高效，我们已经开发了一种超高通量技术(RNAi-细胞微阵列)，用于在果蝇细胞中进行基因组规模的RNAi筛选。为了开发果蝇系统研究TOR信号的潜力，我们建议：(1)识别和表征控制TOR通路的代谢调节机制；(2)识别和了解抑制TOR效应的TOR控制的磷酸酶的调节；以及(3)使用RNAi-细胞微阵列来进行大规模的功能丧失，以发现TOR通路的新成分。总的来说，我们将在果蝇系统中开始我们的实验，随着我们对特定问题的了解的增加，我们的工作将扩展到人类细胞和蛋白质。我们的研究不仅将从根本上促进我们对真核生物生长调控机制的理解，还将发现可能具有药物开发靶点价值的新的信号机制。

DESCRIPTION (provided by applicant): The process of growth (mass accumulation) is a critical determinant of cell, organ, and body size and is often deregulated in diseases such as cancer and diabetes. We are studying the TOR (mTOR in mammals; dTOR in drosophila) pathway, a conserved signaling system that is emerging as the critical regulator of growth in eukaryotes and is regulated by metabolism, growth factors and stress. The TOR pathway is the target of the FDA-approved immuno suppressant rapamycin that is also used to prevent vessel restenosis after angioplasty and is in trials for the treatment of cancer and autoimmune diseases. Over the last few years we have been studying the biochemistry of the mTOR pathway in human tissue culture cells. Although this work has been fruitful and has lead to the discovery of several proteins that interact with mTOR (e.g. raptor and GbetaL), we have come to realize that many of the important questions about TOR signaling are more easily answered in a model system that uses drosophila rather than human tissue culture cells. The human and drosophila TOR (dTOR) pathways are highly conserved and our preliminary data suggests that both pathways respond alike to nutrient metabolism, contain an unidentified TOR-regulated phosphatase, and have similar effects on cell size. In contrast, the yeast TOR pathway is missing several important components, including S6 kinase (S6K), TSC1, and TSC2, senses different nutrients and does not have growth factor inputs. There are several advantages to studying TOR signaling in drosophila rather than human tissue culture cells. The fly pathway has less redundancy than the human version, loss of function mutations are remarkably easy and efficient to make, and we have developed an ultra high-throughput technology (RNAi-cell microarrays) for undertaking genome-scale RNAi screens in drosophila cells. To exploit the potential of the drosophila system to study TOR signaling we propose to: (1) identify and characterize the metabolism-regulated mechanisms that control the TOR pathway; (2) identify and understand the regulation of a TOR controlled phosphatase that inhibits TOR effectors; and (3) use RNAi-cell microarrays to undertake large scale loss of function screens to discover new components of the TOR pathway. In general, we will begin our experiments in the drosophila system and, as our knowledge of a particular problem increases, extend our work to human cells and proteins. Our research will not only lead to a fundamental advance in our understanding of the mechanisms that regulate growth in eukaryotes, but also to the discovery of novel signaling mechanisms that will likely be of value as drug development targets.