Autophagy is known to be required for eukaryotic cells to withstand nutrient starvation by mediating the degradation of cellular components to supply substrates for ATP generation thereby helping cells survive until extracellular nutrient availability returns. Autophagy also mediates forms of quality control within cells by engulfing and degrading protein aggregates and damaged mitochondria that mitigates neurodegeneration in mice. Autophagy substrates become encapsulated by a double membrane structure that then fuses with lysosomes to mediate degradation of the cargo. How autophagosomes form and how their substrates are recognized for engulfment remain poorly understood. We are exploring the regulation and mechanism of gene products including ATG6, ATG13, ATG14 that are known to be required for autophagy. ATG6 is known to form a complex with VPS34 and ATG14 and its recruitment to membranes is dependent on ATG14. We have found that a novel domain of ATG6 that is required for membrane binding and for autophagy induction. We have confirmed that the autophagy activity and membrane targeting function of this domain is conserved in yeast and mammals. Fusion of this domain to the apoptosis inducing protein Bax spontaneously activates Bax indicating that this domain can target mitochondrial membranes. We have also found that Atg6 is phosphorylated specifically upon autophagy induction and we have identified the amino acids phosphorylated allowing us to mutate these sites and determine the role of phosphorylation in autophagy regulation. To more conclusively understand the molecular mechanisms of autophagy we have knocked out ATG5, ATG6, ATG13, and ATG14 in mammalian cells using new Talen nuclease technology. These knock out cells reveal that ATG6 phosphorylation depends on expression of ATG14, a protein that specifically docks ATG6 to membranes and is crucial for autophagosome induction. We are using these knock out cells to understand how autophagosomes form and how they recognize and engulf specific cargo such as damaged mitochondria. In a collaborative project we found that the chaperone, HSP90 is required for the selective autophagy of mitochondria and accumulation of ATG13 on the mitochondria as a step in this process. We also plan to explore if the loss of dopaminergic neurons that can occur in animal models of Parkinson's disease can be rescued by up regulation of autophagy pathways.

Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Zip Code
Sekine, Shiori; Youle, Richard J (2018) PINK1 import regulation; a fine system to convey mitochondrial stress to the cytosol. BMC Biol 16:2
Sarraf, Shireen A; Youle, Richard J (2018) Parkin mediates mitophagy during beige-to-white fat conversion. Sci Signal 11:
Yamano, Koji; Wang, Chunxin; Sarraf, Shireen A et al. (2018) Endosomal Rab cycles regulate Parkin-mediated mitophagy. Elife 7:
Stolz, Alexandra; Putyrski, Mateusz; Kutle, Ivana et al. (2017) Fluorescence-based ATG8 sensors monitor localization and function of LC3/GABARAP proteins. EMBO J 36:549-564
Yao, Yong; Nisan, Danielle; Fujimoto, Lynn M et al. (2016) Characterization of the membrane-inserted C-terminus of cytoprotective BCL-XL. Protein Expr Purif 122:56-63
Murphy, Elizabeth; Ardehali, Hossein; Balaban, Robert S et al. (2016) Mitochondrial Function, Biology, and Role in Disease: A Scientific Statement From the American Heart Association. Circ Res 118:1960-91
Lazarou, Michael; Sliter, Danielle A; Kane, Lesley A et al. (2015) The ubiquitin kinase PINK1 recruits autophagy receptors to induce mitophagy. Nature 524:309-314
Nezich, Catherine L; Wang, Chunxin; Fogel, Adam I et al. (2015) MiT/TFE transcription factors are activated during mitophagy downstream of Parkin and Atg5. J Cell Biol 210:435-50
Yamano, Koji; Fogel, Adam I; Wang, Chunxin et al. (2014) Mitochondrial Rab GAPs govern autophagosome biogenesis during mitophagy. Elife 3:e01612
Shen, Qinfang; Yamano, Koji; Head, Brian P et al. (2014) Mutations in Fis1 disrupt orderly disposal of defective mitochondria. Mol Biol Cell 25:145-59

Showing the most recent 10 out of 26 publications