Amino acids are essential activators of mTORC1 via a complex containing RAG GTPases, RAGULATOR and the vacuolar ATPase. Sensing of amino acids causes translocation of mTORC1 to lysosomes, an obligate step for activation. To examine the spatial and temporal dynamics of this translocation, we used live imaging of the mTORC1 component RAPTOR and a cell permeant fluorescent analogue of di-leucine methyl ester. Translocation to lysosomes is a transient event, occurring within 2 min of aa addition and peaking within 5 min. It is temporally coupled with fluorescent leucine appearance in lysosomes and is sustained in comparison to aa stimulation. Sestrin2 and the vacuolar ATPase are negative and positive regulators of mTORC1 activity in our experimental system. Of note, phosphorylation of canonical mTORC1 targets is delayed compared to lysosomal translocation suggesting a dynamic and transient passage of mTORC1 from the lysosomal surface before targetting its substrates elsewhere.
Cells in all organisms must constantly measure the amount of nutrients available to them in order to be healthy and grow properly. For example, cells use a complex sensing system to measure how many amino acids – the building blocks of proteins – are available to them. One enzyme called mTOR alerts the cell to amino acid levels. When amino acids are available, mTOR springs into action and turns on the production of proteins in the cell. However, when amino acids are scarce, mTOR turns off, which slows down protein production and causes the cell to begin scavenging amino acids by digesting parts of itself.
Studies of mTOR have shown that the protein cannot turn on until it visits the surface of small sacks in the cell called lysosomes. These are the major sites within cell where proteins and other molecules are broken down. Scientists know how mTOR gets to the lysosomes, but not how quickly the process occurs.
Now, Manifava, Smith et al. have used microscopes to record live video of the mTOR enzyme as it interacts with amino acids revealing the whole process takes place in just a few minutes. In the experiments, a fluorescent tag was added to part of mTOR to make the protein visible under a microscope. The video showed that, in human cells supplied with amino acids, mTOR reaches the lysosomes within 2 minutes of the amino acids becoming available. Then, within 3-4 minutes the mTOR turns on and leaves the lysosome. Even though the mTOR has left the lysosome, it somehow remembers that amino acids are available and stays active.
The experiments show that mTOR’s brief interaction with the lysosome switches it on and keeps it on even after mTOR leaves. Future studies will be needed to determine exactly how mTOR remembers its interaction with the lysosome and stays active afterwards.