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Dr Nicholas Ktistakis

Abstract:

The mechanism of amino acid import that determines activation of mammalian target of rapamycin (mTOR) and autophagy. Sensing of amino acids determines whether a cell will grow or will initiate a catabolic pathway of nutrient generation termed autophagy. The master regulator for this is the protein kinase mTOR and much is known about its upstream and downstream signalling network (Cell 149: 274-93). The level of activity of this network is the best characterised predictor of longevity, and manipulations as simple as chronic dampening of mTOR activity lead to lifespan extension (eg Nature: 460: 392-395).

Despite much progress, the mechanism by which amino acids are sensed remains unclear, although lysosomes appear to be critical (Science 334: 678-83). Our recent work has suggested that part of the mechanism involves trafficking steps and is regulated by Vps34 a lipid kinase that generates phosphatidylinositol 3-phosphate. Interestingly, the same lipid has a very important effect on the induction of autophagy and this dichotomy of function has been of wide interest (FEBS Lett 584: 1302-12; Biochem Soc Trans 40: 37-43).    To understand how amino acids are imported we have synthesised a series of amino acid analogues and have examined their ability to activate mTOR and inhibit autophagy. Based on these data, we will synthesise fluorescent versions of these compounds and will use high resolution live imaging to follow their cellular uptake. In parallel experiments we will use FRET to examine mTOR activation in those cells suing a specific mTOR FRET reporter that we have characterised. These experiments will provide a spatial description of amino acid sensing, a fundamental event in cellular physiology.