Metabolism and Phosphatase Regulation of the TOR Pathway

TOR 通路的代谢和磷酸酶调节

• 批准号: 7017038
• 负责人: David M. Sabatini
• 金额: $ 31.7万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7017038
• 关键词: Drosophilidae; RNA interference; acetyl coA carboxylase; active sites; aminoacid; biological signal transduction; cell growth regulation; cell line; cell morphology; clinical research; enzyme mechanism; fatty acid biosynthesis; fatty acid synthase; gene mutation; glucose; high throughput technology; microarray technology; phosphoprotein phosphatase; protein protein interaction; serine threonine protein kinase; sirolimus; tissue /cell culture

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.

描述（由申请人提供）：生长过程（物质积累）是细胞、器官和身体大小的关键决定因素，并且在癌症和糖尿病等疾病中常常不受控制。我们正在研究 TOR（哺乳动物中的 mTOR；果蝇中的 dTOR）通路，这是一种保守的信号系统，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 通路是 FDA 批准的免疫抑制剂雷帕霉素的靶标，雷帕霉素也用于预防血管成形术后血管再狭窄，并正在进行治疗癌症和自身免疫性疾病的试验。在过去的几年里，我们一直在研究人类组织培养细胞中 mTOR 通路的生物化学。尽管这项工作卓有成效，并发现了几种与 mTOR 相互作用的蛋白质（例如 raptor 和 GbetaL），但我们已经意识到，有关 TOR 信号传导的许多重要问题在使用以下模型的模型系统中更容易得到解答：果蝇而不是人类组织培养细胞。人类和果蝇 TOR (dTOR) 通路高度保守，我们的初步数据表明，这两种通路对营养代谢的反应相似，都含有一种未识别的 TOR 调节的磷酸酶，并且对细胞大小具有相似的影响。相比之下，酵母TOR通路缺少几个重要的成分，包括S6激酶（S6K）、TSC1和TSC2，感知不同的营养物质并且没有生长因子输入。 研究果蝇而不是人类组织培养细胞中的 TOR 信号传导有几个优点。果蝇途径的冗余度比人类途径的冗余少，功能丧失突变的制造非常容易且高效，而且我们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。为了利用果蝇系统研究 TOR 信号传导的潜力，我们建议：（1）识别并表征控制 TOR 途径的代谢调节机制； (2) 识别并了解抑制 TOR 效应子的 TOR 控制磷酸酶的调节； (3) 使用 RNAi 细胞微阵列进行大规模功能丧失筛选，以发现 TOR 途径的新成分。一般来说，我们将在果蝇系统中开始实验，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。我们的研究不仅将使我们对调节真核生物生长的机制的理解取得根本性进展，而且还将发现新的信号传导机制，这些机制可能作为药物开发目标有价值。