The Target of Rapamycin kinase Complex I (TORC1) is a key regulator of cell growth and metabolism in eukaryotes. Work carried out over the last 15 years has shed light on the mechanisms underlying hormone and amino acid signaling to TORC1, but it is still unclear how other key signals, such as glucose starvation, are transmitted to this highly conserved complex. In the last grant period, we examined TORC1 signaling in budding yeast and found that glucose starvation leads to rapid inactivation of the TORC1 pathway. Then, in a slower reaction (?=10min), TORC1 falls apart and the key regulatory component Kog1/Raptor moves into a single body on the edge of the vacuole/lysosome. The latter event is driven (in part) by AMPK/Snf1 dependent phosphorylation of Kog1 at Ser 491/494, and two nearby prion-like motifs. Kog1-bodies then serve to increase the threshold of TORC1 activation and ensure that cells remain committed to a quiescent state in suboptimal conditions. More recently, we found that the EGO complex (EGOC; Rag/Ragulator in humans)?known to activate TORC1 in the presence of amino acids?also moves into Kog1-bodies. Following up on these data we will now determine: (A) How glucose starvation triggers inhibition of the TORC1 pathway; (B) What proteins are in Kog1/EGOC-bodies; (C) What drives assembly of the Kog1/EGOC-bodies; and, (D) What role Kog1/EGOC-bodies play in cell signaling. To do this we will: (1) Dissect the structure and function of Kog1- bodies. In preliminary experiments we have shown that the Kog1/EGOC bodies remain intact on purified vacuoles. Taking advantage of this, we will now map the composition of the Kog1/EGOC-body and its assembly pathway. Then, using mutants that have defects in Kog/EGOC-body assembly (including some we have already isolated), we will test our prediction that Kog1-bodies play a role in (a) bet hedging, (b) noise dampening, and (c) the coordination of stress and starvation responses. We will also: (2) Identify the proteins that regulate the TORC1 pathway and Kog1/EGOC-body formation, and dissect their mechanism of action. In this aim, we will test the predication that glucose starvation and AMPK/Snf1, trigger changes in EGOC to inactivate TORC1 and drive the formation of Kog1/EGOC?bodies. To do this we will map the circuit that regulates TORC1 signaling and Kog1-body formation, and then study the function of key regulators (including two we recently identified) in detail. Our proposal is innovative in that we study new and unexplored aspects of TORC1 signaling using state-of-the-art systems, genomic, and biochemical approaches. The proposed research is significant in that it promises to shed light on the mechanisms underlying cell growth control in humans and/or lower eukaryotes?with implications for (a) understanding TORC1 related diseases such as cancer, diabetes and obesity, and (b) developing drugs that selectively block the growth of pathogenic fungi. In addition, our work promises to shed light on the role that protein bodies?formed by hundreds of proteins in stress and starvation conditions, and in organisms ranging from yeast to man?play in a healthy cell.

Public Health Relevance

The proposed research is relevant to public health since it focuses on dissecting the mechanisms underlying the regulation of a highly conserved signaling complex (TORC1) known to play an important role in cancer development, diabetes, obesity and depression. The work may also reveal differences between the pathways that control cell growth in lower eukaryotes and humans--opening the door to creating drugs that can selectively block the growth of fungal pathogens.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM097329-08
Application #
9644539
Study Section
Cellular Signaling and Regulatory Systems Study Section (CSRS)
Program Officer
Maas, Stefan
Project Start
2011-09-30
Project End
2021-02-28
Budget Start
2019-03-01
Budget End
2020-02-29
Support Year
8
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Arizona
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
806345617
City
Tucson
State
AZ
Country
United States
Zip Code
85721
Sullivan, Arron; Wallace, Ryan L; Wellington, Rachel et al. (2018) Multi-layered regulation of TORC1-body formation in budding yeast. Mol Biol Cell :mbcE18050297
Hughes Hallett, James E; Luo, Xiangxia; Capaldi, Andrew P (2015) Snf1/AMPK promotes the formation of Kog1/Raptor-bodies to increase the activation threshold of TORC1 in budding yeast. Elife 4:
Worley, Jeremy; Sullivan, Arron; Luo, Xiangxia et al. (2015) Genome-Wide Analysis of the TORC1 and Osmotic Stress Signaling Network in Saccharomyces cerevisiae. G3 (Bethesda) 6:463-74
Hughes Hallett, James E; Luo, Xiangxia; Capaldi, Andrew P (2014) State transitions in the TORC1 signaling pathway and information processing in Saccharomyces cerevisiae. Genetics 198:773-86
Worley, Jeremy; Luo, Xiangxia; Capaldi, Andrew P (2013) Inositol pyrophosphates regulate cell growth and the environmental stress response by activating the HDAC Rpd3L. Cell Rep 3:1476-82
Buchan, J Ross; Capaldi, Andrew P; Parker, Roy (2012) TOR-tured yeast find a new way to stand the heat. Mol Cell 47:155-7