Periodic Reporting for period 4 - TENDO (Tension of ENDOmembranes maintained by TORC1)
Período documentado: 2023-11-01 hasta 2024-10-31
TENDO studies the structure, function and regulation of the two Target Of Rapamycin Complexes, TORC1 and TORC2. These complexes are broadly conserved across eukaryote species and serve fundamental roles in cell and organism homeostasis.
TORC1 and TORC2 have protein kinase activity and thus function in signal transduction pathways that help cells maintain their size: TORC1 regulates cell volume while TORC2 appears to regulate cell surface area. Dysregulation of these particular signaling cascades is associated with diseases such as metabolic disorder, cancer and even aging itself. Ultimately, with a better, ideally molecular or eventually atomic, understanding of TOR signaling we hope to be able to understand and thus therapeutically manipulate these pathways for therapeutic gain.
The lab has recently made three major breakthroughs in this field. These were based in the model eukaryote S. cerevisiae (bakers' yeast). The first is that TORC1 is regulated via reversible polymerization into a giant helix. In this helical form, the active site of the enzyme is physically occluded and signaling is thus blocked. The second is that TORC2 is regulated by changes in the tension of the plasma membrane. The third is that membrane tension can be altered using small molecules. In TENDO we wish to determine i) if TORC2, like TORC1 is regulated via reversible polymerization; ii) if TORC1 is regulated downstream of biophysical changes in intracellular membranes; and iii) if we can drug membranes as a means to affect TOR signaling.
TORC1 and TORC2 have protein kinase activity and thus function in signal transduction pathways that help cells maintain their size: TORC1 regulates cell volume while TORC2 appears to regulate cell surface area. Dysregulation of these particular signaling cascades is associated with diseases such as metabolic disorder, cancer and even aging itself. Ultimately, with a better, ideally molecular or eventually atomic, understanding of TOR signaling we hope to be able to understand and thus therapeutically manipulate these pathways for therapeutic gain.
The lab has recently made three major breakthroughs in this field. These were based in the model eukaryote S. cerevisiae (bakers' yeast). The first is that TORC1 is regulated via reversible polymerization into a giant helix. In this helical form, the active site of the enzyme is physically occluded and signaling is thus blocked. The second is that TORC2 is regulated by changes in the tension of the plasma membrane. The third is that membrane tension can be altered using small molecules. In TENDO we wish to determine i) if TORC2, like TORC1 is regulated via reversible polymerization; ii) if TORC1 is regulated downstream of biophysical changes in intracellular membranes; and iii) if we can drug membranes as a means to affect TOR signaling.
Arguably our most important discovery to come from TENDO was the determination of a high-resolution structure of a native membrane microdomain, the eisosome, that functions upstream of TORC2 (10.1038/s41586-024-07720-6). In this work, we could observe how a peripheral protein coat stabilizes specific lipids species in the underlying membrane and how an increase in membrane tension triggers the release of these lipids and likely other signaling molecules. This work provides a major advance in our understanding of how mechanical stress impacting a membrane bilayer can be sensed and transmitted to effect intracellular signaling pathways that enable the cell to respond to these stresses. Furthermore, using a small molecule that we had previously discovered in a high-throughput drug screen for TORC2 inhibitors, we discovered that insults that lead to a reduction in plasma membrane tension trigger an unexpected mobilization of previously cloistered sterol molecules as well as the formation of giant membrane invaginations which appear to be mechanistically coupled to TORC2 inactivation. This work is currently under revision for publication (10.1101/2024.10.18.618785).
A second, far-reaching advance has been our determination of a high-resolution structure of the SEA Complex, a GTPase activating complex that plays a central role in TORC1 regulation in both yeast and humans (10.1038/s41586-022-05370-0). Moreover, we recently determined a structure of the SEA Complex together with its substrate, the EGO Complex (10.1101/2024.10.05.616782). These were important "missing links" in the TOR field important as they help us understand the molecular mechanisms coupling environmental stresses to TORC1 activity regulation. In related work, we discovered that the EGO Complex activates TORC1 by physically removing it from an inhibitory polymer (10.1038/s41594-022-00912-6) bringing critical insight into potentially conserved mechanisms by which TORC1 is regulated in all eukaryotes.
A second, far-reaching advance has been our determination of a high-resolution structure of the SEA Complex, a GTPase activating complex that plays a central role in TORC1 regulation in both yeast and humans (10.1038/s41586-022-05370-0). Moreover, we recently determined a structure of the SEA Complex together with its substrate, the EGO Complex (10.1101/2024.10.05.616782). These were important "missing links" in the TOR field important as they help us understand the molecular mechanisms coupling environmental stresses to TORC1 activity regulation. In related work, we discovered that the EGO Complex activates TORC1 by physically removing it from an inhibitory polymer (10.1038/s41594-022-00912-6) bringing critical insight into potentially conserved mechanisms by which TORC1 is regulated in all eukaryotes.
We have solved the structure, for the first time to our knowledge, of a native membrane microdomain. Mutational analyses based on this structure will now enable us and others to define how eisosomes, and potentially functionally-related caveolae, sense and transduce changes in membrane properties.
We have solved the structure of a major regulator functioning upstream of TORC1. This structures sets the basis for future studies to establish a molecular understanding for how TORC1 is regulated by environmental insults.
Magnaporthe oryzae is a major crop pathogen, destroying each year enough rice to feed 60M people. Presently we are translating our new understanding of TORC1 and TORC2 signaling derived from studies in Saccharomyces cerevisiae (brewers' yeast) to this related fungus. We hope that these future studies will reveal new targets for antifungal agents that could have enormous impact on agriculture.
We have solved the structure of a major regulator functioning upstream of TORC1. This structures sets the basis for future studies to establish a molecular understanding for how TORC1 is regulated by environmental insults.
Magnaporthe oryzae is a major crop pathogen, destroying each year enough rice to feed 60M people. Presently we are translating our new understanding of TORC1 and TORC2 signaling derived from studies in Saccharomyces cerevisiae (brewers' yeast) to this related fungus. We hope that these future studies will reveal new targets for antifungal agents that could have enormous impact on agriculture.