Signal Transduction to p70 S6 Kinase 1

描述（由申请人提供）：许多人类疾病是由细胞生长（细胞质量和大小增加）、增殖、迁移、存活和死亡的不当调节引起的。这些过程受到包含哺乳动物雷帕霉素靶蛋白（mTOR）、Lst 8和Raptor的复合物（称为mTORC 1复合物）的关键调控。S6蛋白激酶（S6 K）是mTORC 1的主要效应子。mTORC 1/S6 K信号系统是细胞的中心整合点，用于多种稳态输入，感知生长因子可用性，能量水平和氨基酸充足性。mTORC 1/S6 K信号转导的过度活化是几乎所有人类癌症的共同特征。mTORC 1抑制剂，如雷帕霉素及其类似物，目前正在临床上评估用于治疗癌症。虽然抑制剂已经显示出一些前景，但雷帕霉素不敏感的mTOR信号传导也影响肿瘤发生，并且存在反馈回路，其上调雷帕霉素治疗后的存活途径。因此，需要靶向该途径的其他组分的另外的治疗剂。通过采用全系统的方法来定义mTORC 1/S6 K通路调节以及该信号系统调节各种生物学过程的机制，我们希望提供见解，从而确定用于治疗mTORC 1/S6 K依赖性癌症和其他代谢疾病的新型治疗策略。 目的1通过S6 K1特异性相互作用蛋白SKAR，将S6 K1信号传导、基因表达和细胞生长调控联系起来。描述了研究SKAR和S6 K1在mRNA生物合成和蛋白质翻译调节中的作用的方法。这一目标也为我们如何处理与目标2中确定的mTORC 1/S6 K信号系统相关的所有mRNA结合蛋白奠定了基础。 目的2中概述的方法将多种生化纯化方法与质谱分析相结合，以鉴定和验证S6 K信号传导系统的一组常见的近端上游调节因子和下游效应因子。使用多个汇聚的研究路线将把我们的努力集中在该途径的最关键组成部分上，使我们能够剖析S6 K如何调节如此多的不同细胞过程。 目的3利用基于RNAi的遗传学方法阐明mTORC 1/S6 K通路的稳态调控机制。为此，我们开发了一种灵敏的、高通量的、基于图像的筛选策略，用于监测体内S6 K活性。我们建议利用这种独特的检测方法来广泛地询问mTORC 1/S6 K通路中的有丝分裂原和营养调节输入。公共卫生相关性：我们希望本提案中概述的研究将加深我们对mTORC 1/S6 K信号传导缺陷的理解，这些缺陷导致癌症进展和细胞生长相关疾病，例如儿童癌症易感综合征硬化症。这些研究也将影响我们对与S6 K相关的其他代谢疾病的理解，如糖尿病和肥胖症。我们相信这种机制洞察将为识别抑制mTORC 1/S6 K信号网络的生长因子和/或氨基酸传感臂的新型治疗策略打开大门。

DESCRIPTION (provided by applicant): Many human diseases result from improper regulation of cell growth (an increase in cell mass and size), proliferation, migration, survival and death. These processes are critically regulated by a complex containing the mammalian target of rapamycin (mTOR), Lst8 and Raptor, called the mTORC1 complex. The S6 protein kinases (S6K) are major effectors of mTORC1. The mTORC1/S6K signaling system is the cell's central integration point for multiple homeostatic inputs, sensing growth factor availability, energy levels, and amino acid sufficiency. Hyperactivation of mTORC1/S6K signaling is a common feature of nearly all human cancers. mTORC1 inhibitors, such as rapamycin and its analogs, are currently being clinically evaluated for the treatment of cancer. While the inhibitors have exhibited some promise, rapamycin- insensitive mTOR signaling also influences tumorigenesis, and feedback loops exist that up- regulate survival pathways following rapamycin treatment. Thus, additional therapeutic agents targeting other components of this pathway are needed. By taking a systems-wide approach towards defining mTORC1/S6K pathway regulation and the mechanisms by which this signaling system modulates various biological processes, we hope to provide insights that will lead to the identification of novel therapeutic strategies for the treatment of mTORC1/S6K- dependent cancers and other metabolic disorders. Aim 1 will focus on the connection between S6K1 signaling, gene expression and cell growth control through an S6K1-specific interacting protein SKAR. Approaches are described that investigate the role of SKAR and S6K1 in the regulation of mRNA biogenesis and protein translation. This aim also sets the foundation for how we will approach all mRNA binding proteins linked to the mTORC1/S6K signaling system identified in Aim 2. The approach outlined in Aim 2 combines a variety of biochemical purification approaches with mass spectrometry analysis to identify and validate a common set of proximal upstream regulators and downstream effectors of the S6K signaling system. The use of multiple converging lines of investigation will focus our efforts on the most critical components of the pathway, allowing us to dissect how S6K regulates so many disparate cellular processes. Aim 3 utilizes RNAi-based genetic approaches to elucidate the mechanism of homeostatic regulation of the mTORC1/S6K pathway. To this end, we have developed a sensitive, high- throughput, image-based screening strategy for monitoring S6K activity in vivo. We propose to utilize this unique assay to broadly interrogate mitogen- and nutrient-regulated inputs into the mTORC1/S6K pathway. PUBLIC HEALTH RELEVANCE: We hope that the studies outlined in this proposal will deepen our understanding of the defects in mTORC1/S6K signaling that are responsible for cancer progression and cell growth-associated diseases, such as the childhood cancer predisposition syndrome Tuberous Sclerosis. These studies will also impact our understanding of other metabolic diseases linked to S6K, such as diabetes and obesity. We believe such mechanistic insight will open the door to the identification of novel therapeutic strategies for inhibiting the growth factor and/or amino acid sensing arms of the mTORC1/S6K signaling network.