Mechanistic Studies of Autophagy Initiation in Mammalian Cells

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Mechanistic Studies of Autophagy Initiation in Mammalian Cells

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dc.contributor.advisor Wang, Xiaodong en
dc.creator Shang, Libin en
dc.date.accessioned 2011-08-26T17:35:06Z en
dc.date.available 2011-08-26T17:35:06Z en
dc.date.issued 2011-08-26T17:35:06Z en
dc.identifier.other 759107106 en
dc.identifier.uri http://hdl.handle.net/2152.5/886 en
dc.description.abstract Macroautophagy (herein referred to as autophagy) is an evolutionarily conserved self-digestive process cells use to adapt to starvation and other stresses. During autophagy, portions of cytoplasmic materials are engulfed into specialized double-membrane structures to form autophagosomes, which then fuse with lysosomes to degrade their cargos and regenerate nutrients. Initiation of autophagy has been extensively studied in budding yeast Saccharomyces cerevisiae. However, various significant differences exist between yeast and mammals. To pinpoint how mammalian autophagy is initiated, I first adopted proteomic approaches to identify associating partners of Unc-51-like kinase 1 (Ulk1), key initiator for mammalian autophagy. Two novel proteins, mAtg13 and Atg101, were found to interact with Ulk1 stoichiometrically. Knockdown of either mAtg13 or Atg101 led to decreased autophagy, and autophagy could be rescued with exogenous expression, suggesting the two proteins were critical for mammalian autophagy initiation. I then observed Ulk1 undergoes dramatic dephosphorylation upon starvation, particularly at serine 638 and serine 758. I found phosphorylations of Ulk1 are mediated by mammalian target-of-rapamycin (mTOR) kinase and AMP-activated protein kinase (AMPK). AMPK interacts with Ulk1 in a nutrient-dependent manner, and proper phosphorylations on Ulk1 are crucial for Ulk1/AMPK association, as a single serine-to-alanine mutation (S758A) at Ulk1 impairs this interaction. Compared to its wild-type counterpart, this Ulk1-S758A mutant initiates starvation-induced autophagy faster at early time points, but does not alter the maximum capacity of autophagy when starvation prolongs. With this layer of regulation, mammalian autophagy is capable of responding to environmental changes more promptly than previously considered. en
dc.language.iso en en
dc.subject TOR-Serine-Threonine Kinases en
dc.subject Protein-Serine-Threonine Kinases en
dc.subject Autophagy en
dc.title Mechanistic Studies of Autophagy Initiation in Mammalian Cells en
dc.type Thesis en
thesis.degree.grantor Graduate School of Biomedical Sciences en
thesis.degree.name Doctor of Philosophy en
thesis.degree.level Ph.D. en

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