title: Structure-guided mutagenesis reveals a hierarchical mechanism of Parkin activation.
abstract: Parkin and PINK1 function in a common pathway to clear damaged mitochondria. Parkin exists in an auto-inhibited conformation stabilized by multiple interdomain interactions. The binding of PINK1-generated phospho-ubiquitin and the phosphorylation of the ubiquitin-like (Ubl) domain of Parkin at Ser65 release its auto-inhibition, but how and when these events take place in cells remain to be defined. Here we show that mutations that we designed to activate Parkin by releasing the Repressor Element of Parkin (REP) domain, or by disrupting the interface between the RING0:RING2 domains, can completely rescue mutations in the Parkin Ubl that are defective in mitochondrial autophagy. Using a FRET reporter assay we show that Parkin undergoes a conformational change upon phosphorylation that can be mimicked by mutating Trp403 in the REP. We propose a hierarchical model whereby pUb binding on mitochondria enables Parkin phosphorylation, which, in turn, leads to REP removal, E3 ligase activation and mitophagy.
authors.affiliation: McGill Parkinson Program, Neurodegenerative Diseases Group, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montréal, Québec, Canada H3A 2B4.
authors.forename: Matthew Y
authors.initials: MY
authors.lastname: Tang
authors.affiliation: Groupe de Recherche Axé sur la Structure des Protéines, Department of Pharmacology and Therapeutics, McGill University, Montréal, Québec, Canada H3G 1Y6.
authors.forename: Marta
authors.identifier: 0000-0002-4296-9018
authors.initials: M
authors.lastname: Vranas
authors.affiliation: McGill Parkinson Program, Neurodegenerative Diseases Group, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montréal, Québec, Canada H3A 2B4.
authors.forename: Andrea I
authors.initials: AI
authors.lastname: Krahn
authors.affiliation: McGill Parkinson Program, Neurodegenerative Diseases Group, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montréal, Québec, Canada H3A 2B4.
authors.forename: Shayal
authors.initials: S
authors.lastname: Pundlik
authors.affiliation: Groupe de Recherche Axé sur la Structure des Protéines, Department of Pharmacology and Therapeutics, McGill University, Montréal, Québec, Canada H3G 1Y6.
authors.forename: Jean-François
authors.initials: JF
authors.lastname: Trempe
authors.affiliation: McGill Parkinson Program, Neurodegenerative Diseases Group, Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montréal, Québec, Canada H3A 2B4.
authors.forename: Edward A
authors.initials: EA
authors.lastname: Fon
mesh_terms: D045744:Cell Line, Tumor; D006367:HeLa Cells; D006801:Humans; D008856:Microscopy, Fluorescence; D063306:Mitophagy; D016296:Mutagenesis; D009154:Mutation; D010766:Phosphorylation; D011487:Protein Conformation; D011494:Protein Kinases / Q000378:metabolism; D012694:Serine / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D059008:Time-Lapse Imaging / Q000379:methods; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination; D000099202:PTEN-Induced Putative Kinase
chemical_list: D011494:Protein Kinases; D012694:Serine; D044767:Ubiquitin-Protein Ligases; D000099202:PTEN-Induced Putative Kinase; C111567:parkin protein