Final Report Summary - TORCH (TOR and Cellular Homeostasis)
The Target of the natural product rapamycin, TOR, is a protein kinase that regulates growth of virtually all eukaryote cells. To do this, TOR assembles into two, distinct multiprotein complexes which we named TORC1 and TORC2. The goal of TORCH is to understand the feedback loops in which the two TOR complexes operate to maintain cellular homeostasis in the face of changing environmental conditions.
For these studies we use the model eukaryote Saccharomyces cerevisiae (bakers’ yeast) which is cheap to grow, easy to manipulate genetically and free of ethical concerns. During our investigations, we discovered that changes in glucose availability, a key nutrient for eukaryotes, has a profound and amazing effect on TORC1 – glucose withdrawal causes TORC1 to polymerize into the biggest protein-only filament reported to date. This polymerization causes a steric occlusion of the active site of the kinase and thereby inhibits TORC1 activity, slowing cell growth and enhancing stress responses including self-degradation (autophagy). Furthermore, we found that this regulation is mediated by a conserved family of GTPases (the RAG GTPases) suggesting that similar things might be happening in our own cells. Recently, we have determined a much higher-resolution structure of these filaments (which we named TOROIDs for TORC1 Organized in Inhibited Domains). With this new information we hope to understand how the GTPases regulate TOROID formation and disassembly and, potentially, how TOROIDs could be drugged in diseases in which dysregulation of mammalian TORC1 activity, such as cancer, cachexia and metabolic syndrome, is implicated.
In parallel to these studies, we found that TORC2 is regulated by membrane tension: increased membrane tension triggers another protein, Slm1, to associate with TORC2 and through mechanisms that we are still studying, causes its hyperactivation. Recently, we found that loss of membrane tension provokes a spontaneous phase separation of a certain type of lipid, PI4,5P2 which clusters and inactivates TORC2. Based on our recent results with TORC1, we believe the inactive TORC2 clusters are also some form of higher-order assembly.
In sum, our results suggest that while TORC1 regulates cell mass and cell volume, TORC2 regulates cell surface area – both are important for cell growth to occur. Understanding these pathways is important for both a fundamental understanding of how eukaryotic cells growth and, potentially, for the future design of new therapeutic strategies.
For these studies we use the model eukaryote Saccharomyces cerevisiae (bakers’ yeast) which is cheap to grow, easy to manipulate genetically and free of ethical concerns. During our investigations, we discovered that changes in glucose availability, a key nutrient for eukaryotes, has a profound and amazing effect on TORC1 – glucose withdrawal causes TORC1 to polymerize into the biggest protein-only filament reported to date. This polymerization causes a steric occlusion of the active site of the kinase and thereby inhibits TORC1 activity, slowing cell growth and enhancing stress responses including self-degradation (autophagy). Furthermore, we found that this regulation is mediated by a conserved family of GTPases (the RAG GTPases) suggesting that similar things might be happening in our own cells. Recently, we have determined a much higher-resolution structure of these filaments (which we named TOROIDs for TORC1 Organized in Inhibited Domains). With this new information we hope to understand how the GTPases regulate TOROID formation and disassembly and, potentially, how TOROIDs could be drugged in diseases in which dysregulation of mammalian TORC1 activity, such as cancer, cachexia and metabolic syndrome, is implicated.
In parallel to these studies, we found that TORC2 is regulated by membrane tension: increased membrane tension triggers another protein, Slm1, to associate with TORC2 and through mechanisms that we are still studying, causes its hyperactivation. Recently, we found that loss of membrane tension provokes a spontaneous phase separation of a certain type of lipid, PI4,5P2 which clusters and inactivates TORC2. Based on our recent results with TORC1, we believe the inactive TORC2 clusters are also some form of higher-order assembly.
In sum, our results suggest that while TORC1 regulates cell mass and cell volume, TORC2 regulates cell surface area – both are important for cell growth to occur. Understanding these pathways is important for both a fundamental understanding of how eukaryotic cells growth and, potentially, for the future design of new therapeutic strategies.