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

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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.