Autophagy is an essential degradation mechanism that is highly conserved throughout eukaryotes. Autophagy recycles cellular components, generating essential precursors needed for cell survival during stress or starvation, and removing harmful materials. Deficiencies in autophagy are linked to aging, cancer, and neurodegenerative diseases. Targeted components are engulfed within a double membrane structure, termed the autophagosome, and shuttled to the lysosome for degradation. Starvation is the primordial signal for autophagy induction. The proposed research seeks to elucidate, at the molecular and atomic levels, how starvation initiates autophagy. Atg1 is the earliest acting complex in autophagy, where it serves as a vesicle tether. The central hypothesis to be tested is that assembly of the Atg1 complex is regulated by the TORC1 via phosphoregulation of the interaction of the Atg13 and Atg17 subunits.
The specific aims of the project are to (1) determine the structure of Atg13 in complex with Atg17 using X-ray crystallography complemented by solution studies using analytical ultracentrifugation, (2) determine the in vivo effects of mutatios at the interface of Atg13 and Atg17 by conducting three well-characterized yeast autophagy assays: GFP-Atg8 puncta, GFP-Atg8 processing, and Pho8-delta-60 activity, and (3) to determine how phosphorylation sites in Atg13 perturb binding to Atg17 using structural analysis and biophysical analysis using isothermal titration calorimetry (ITC). Together, these aims will provide mechanistic understanding into the role of the TORC1 kinase complex in regulating starvation-induced autophagy.
The proposed research seeks to elucidate the fundamental mechanisms of autophagy that is essential for cell survival under starvation and stress. This ancient and conserved mechanism has profound connections to aging, cancer, and neurodegenerative diseases. Understanding autophagy at its most basic level could lead to a better understanding of the most complex diseases that hinder our society.
|Stjepanovic, Goran; Davies, Christopher W; Stanley, Robin E et al. (2014) Assembly and dynamics of the autophagy-initiating Atg1 complex. Proc Natl Acad Sci U S A 111:12793-8|