Defining the logic and function of the AMPK-mTORC1 signaling axis

项目摘要 在后生动物中，器官的稳态需要增殖信号和能量状态的协调， 然而，分子反馈系统如何平衡单个细胞内的能量消耗， 明白本研究的目的是确定细胞生长是如何调节代谢反馈控制在单一的， 细胞水平。为了实现这一目标，我将结合时间推移成像和代谢组学方法来构建 信号动力学和代谢通量之间的定量关系。一个完善的机制 其协调细胞生长和体内平衡是AMPK信号传导轴。在能量有限的状态下， AMPK直接抑制能量昂贵的生长过程，同时促进分解代谢 途径。AMPK活性的净效应是增加还原剂的可用性， 前体和ATP。这项工作将建立在最近的发现，即在正常生长条件下，AMPK 活性是动态的，并且与个体中主要增殖途径的活性强烈反相关。 细胞为了确定AMPK活性如何直接限制mTORC 1的信号整合，mTORC 1是合成代谢的主要调节因子， 过程中，我将多重荧光报告，同时测量AMPK和mTORC 1活动 活细胞内。活细胞成像方法将用于定义动态输入/输出关系 AMPK活性、mTORC 1信号传导和下游过程（包括蛋白质合成速率）之间的关系 翻译和DNA合成。接下来，代谢组学表征将定义精确的适应函数 以及由AMPK-mTORC 1控制回路支持的代谢配置。最终，这项研究将确定 如何整合基于信号和基于代谢的控制机制以协调代谢 在增殖细胞中的稳态。对AMPK-mTORC 1控制回路的动态理解对于以下方面至关重要： 了解能量平衡是如何在单细胞水平上实现的。这项研究的结果可能 支持开发针对许多人类疾病的新型治疗方法， 生长和能量过程的失调。

PROJECT SUMMARY Across metazoans, organ homeostasis requires the coordination of proliferative signals and energetic states, yet how molecular feedback systems balance energy expenditure within an individual cell is not well understood. This study aims to define how cell growth is regulated by metabolic feedback control at the single- cell level. To achieve this goal, I will couple time-lapse imaging and metabolomic approaches to construct quantitative relationships between signaling dynamics and metabolic flux. One well-established mechanism which coordinates cellular growth and homeostasis is the AMPK signaling axis. In energetically limited states, AMPK directly inhibits energetically expensive growth processes and simultaneously promotes catabolic pathways. The net effect of AMPK activity is to increase the availability of reducing agents, biosynthetic precursors, and ATP. This work will build upon the recent finding that under normal growth conditions, AMPK activity is dynamic and strongly anti-correlated with the activities of major proliferative pathways in individual cells. To define how AMPK activity directly limits signal integration by mTORC1, a master regulator of anabolic processes, I will multiplex fluorescent reporters to simultaneously measure AMPK and mTORC1 activities within living cells. Live cell imaging approaches will then be used to define dynamic input/output relationships between AMPK activity, mTORC1 signaling, and downstream processes including the rates of protein translation and DNA synthesis. Next, metabolomic characterization will define the precise adaptive function and metabolic configurations supported by the AMPK-mTORC1 control loop. Ultimately, this study will define how signaling-based and metabolite-based control mechanisms are integrated to coordinate metabolic homeostasis in proliferating cells. A dynamic understanding of the AMPK-mTORC1 control loop is essential for understanding how energetic homeostasis is achieved at a single cell level. Findings from this study may support the development of novel therapeutic approaches that target the many human diseases characterized by the dysregulation of growth and energetic processes.