Dissection of the TORC1 Signaling Network in Yeast

University Of Arizona, Tucson AZ

Investigators

Abstract

Summary The Target of Rapamycin kinase Complex I (TORC1) is a central hub in the cell growth and metabolic control network of eukaryotes. Studies carried out over the last twenty years have identified numerous components in the TORC1 pathway, and uncovered the important role that TORC1 plays in diseases and disorders such as cancer, epilepsy, diabetes, depression, and aging. However, even as our understanding of TORC1 signaling matures, it remains unclear how TORC1 functions at the systems level. Why are there so many TORC1 regulators--some with seemingly identical functions? And how are the signals transmitted through individual TORC1 regulators integrated to ensure that cell growth and metabolism remain balanced across conditions? Over the past 12 years we have addressed these and related questions by building up a detailed, network level, model of TORC1 signaling in the simple model organism Saccharomyces cerevisiae. Our work has led to the identification of several new proteins involved in TORC1 regulation, but perhaps most importantly has (i) shown that TORC1 is driven into different signaling states in different conditions to optimize the activity of hundreds of downstream targets and (ii) uncovered that the cAMP dependent protein kinase (PKA) pathway acts in parallel with TORC1 to tune the threshold and timing of the cell growth response. Most recently, we discovered that TORC1 is pushed into a unique, intermediate signaling state in poor nutrient conditions, where it activates targets involved in amino acid transport and metabolism, and slows growth via a single transcriptional repressor, but other TORC1 outputs match those seen in ideal growth conditions. Building on this framework, we now propose an ambitious and integrated set of experiments to: (1) Determine how TORC1 is driven into the intermediate signaling state in poor nutrient conditions; (2) Determine how TORC1 signaling is altered, as conditions get worse, to slow growth further and tune metabolism; and (3) Identify the missing layers of TORC1 regulation that push cells into quiescence/arrest during complete starvation. To do this, we will map the impact that known and putative TORC1 regulators have on signaling across a wide range of conditions using several reporters of TORC1 activity, live cell microscopy, and phosphoproteomics. We will also study the impact that over 100 newly identified TORC1 pathway interactors have on TORC1 and global signaling across conditions, using high- throughput microscopy, phosphoproteomics, transcriptomics, and detailed follow up experiments. The resulting model should serve as a paradigm for future systems level studies of human TORC1 signaling and will shed light on differences between yeast and human TORC1 signaling network, opening the door to creating new (and sorely needed) antifungal compounds.