We are interested in developing a better understanding of the biology of the non-dividing resting states that eukaryotic cells enter when conditions are not conducive to continued growth. One of our primary goals is to define how the processes that are induced during these periods of quiescence are regulated and how they contribute to general cell survival. This proposal is focused upon a set of related pathways, known collectively as autophagy, that are required for this survival. Autophagy pathways are responsible for the turnover of cytoplasmic material, including bulk protein and damaged or superfluous organelles. This autophagy-mediated degradation has been linked to a variety of processes relevant to human health, including tumor suppression, innate immunity and neurological disorders, such as Huntington's disease. In many of these conditions, the autophagy pathway is being considered as a major point of therapeutic intervention. It is therefore critical that we develop a thorough understanding of the mechanisms normally controlling autophagy in eukaryotic cells. This proposal will examine two key aspects of the regulation of autophagy in the yeast, Saccharomyces cerevisiae. Studies with this organism have contributed tremendously to our basic understanding of autophagy in all eukaryotes, including humans. First, a combination of approaches will be used to characterize the autophagy process induced by the inactivation of the cAMP-dependent protein kinase (PKA) signaling pathway. Our preliminary work indicates that this PKA-regulated process may be similar to an alternative form of macroautophagy recently identified in mammals. The experiments here will explore this possibility and define the molecular machineries governing this PKA-regulated pathway. These studies will also characterize a potential role for the phosphoinositide, PtdIns (3,5)P2, in this PKA-regulated macroautophagy. The second major goal of this proposal is the identification of the substrates of the Atg1 protein kinase. Atg1 is a key regulatory target within the autophagy machinery and identifying the targets of this enzyme represents one of the major goals in the autophagy field today. In all, we feel that the completion of this work will provide important insights into the manner in which autophagy is controlled in eukaryotic cells, insights that should facilitate efforts to manipulate this pathway in clinically beneficial ways.
The specific aims i n this proposal are: (1) to characterize the autophagy pathway induced upon inactivation of PKA signaling;(2) to characterize the link between PKA signaling, phosphoinositide metabolism and the regulation of autophagy;and (3) to identify and characterize substrates of the Atg1 protein kinase.

Public Health Relevance

This proposal aims to further our understanding of a process known as autophagy that has been linked to a number of serious human ailments, including breast and ovarian cancer, Crohn's disease and neurological disorders, such as Huntington's disease. Interestingly, drugs that target autophagy are being developed as potential therapeutics for many of these conditions. By increasing our understanding of the normal control of the autophagy process, the work here would provide potentially novel avenues for this drug discovery process.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM065227-10
Application #
8309101
Study Section
Membrane Biology and Protein Processing (MBPP)
Program Officer
Maas, Stefan
Project Start
2002-04-01
Project End
2015-05-31
Budget Start
2012-06-01
Budget End
2013-05-31
Support Year
10
Fiscal Year
2012
Total Cost
$308,813
Indirect Cost
$106,313
Name
Ohio State University
Department
Genetics
Type
Schools of Arts and Sciences
DUNS #
832127323
City
Columbus
State
OH
Country
United States
Zip Code
43210
Shah, Khyati H; Zhang, Bo; Ramachandran, Vidhya et al. (2013) Processing body and stress granule assembly occur by independent and differentially regulated pathways in Saccharomyces cerevisiae. Genetics 193:109-23
Mousley, Carl J; Yuan, Peihua; Gaur, Naseem A et al. (2012) A sterol-binding protein integrates endosomal lipid metabolism with TOR signaling and nitrogen sensing. Cell 148:702-15
Ramachandran, Vidhya; Herman, Paul K (2011) Antagonistic interactions between the cAMP-dependent protein kinase and Tor signaling pathways modulate cell growth in Saccharomyces cerevisiae. Genetics 187:441-54
Yeh, Yuh-Ying; Shah, Khyati H; Chou, Chi-Chi et al. (2011) The identification and analysis of phosphorylation sites on the Atg1 protein kinase. Autophagy 7:716-26
Yeh, Yuh-Ying; Shah, Khyati H; Herman, Paul K (2011) An Atg13 protein-mediated self-association of the Atg1 protein kinase is important for the induction of autophagy. J Biol Chem 286:28931-9
Ramachandran, Vidhya; Shah, Khyati H; Herman, Paul K (2011) The cAMP-dependent protein kinase signaling pathway is a key regulator of P body foci formation. Mol Cell 43:973-81
Stephan, Joseph S; Yeh, Yuh-Ying; Ramachandran, Vidhya et al. (2010) The Tor and cAMP-dependent protein kinase signaling pathways coordinately control autophagy in Saccharomyces cerevisiae. Autophagy 6:294-5
Yeh, Yuh-Ying; Wrasman, Kristie; Herman, Paul K (2010) Autophosphorylation within the Atg1 activation loop is required for both kinase activity and the induction of autophagy in Saccharomyces cerevisiae. Genetics 185:871-82
Stephan, Joseph S; Yeh, Yuh-Ying; Ramachandran, Vidhya et al. (2009) The Tor and PKA signaling pathways independently target the Atg1/Atg13 protein kinase complex to control autophagy. Proc Natl Acad Sci U S A 106:17049-54
Deminoff, Stephen J; Ramachandran, Vidhya; Herman, Paul K (2009) Distal recognition sites in substrates are required for efficient phosphorylation by the cAMP-dependent protein kinase. Genetics 182:529-39

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