title: PINK1 rendered temperature sensitive by disease-associated and engineered mutations.
abstract: Mutations in Parkin or PINK1 are the most common cause of recessively inherited parkinsonism. Parkin and PINK1 function in a conserved mitochondrial quality control pathway, in which PINK1, a putative mitochondrial kinase, directs Parkin, a cytosolic E3 ubiquitin ligase, selectively to dysfunctional mitochondria to promote their isolation, immobilization and degradation by macroautophagy (hereafter, mitophagy). As Parkin recruitment to mitochondria is robustly induced by PINK1 expression on the outer mitochondrial membrane, Parkin recruitment to mitochondria was used as an assay for PINK1 function. Unexpectedly, mutation of serine residues within the activation segment of PINK1 uncovered a temperature-sensitive variant of PINK1 (tsPINK1). tsPINK1 allowed for the first time the disassociation of PINK1 activity from its expression and localization. Additionally, extensive mutagenesis identified three disease-associated variants in the activation segment and one in an α-helix N-terminal to kinase domain (Q126P) that are similarly thermally labile, suggesting that their activity could be restored post-translationally (e.g. by reducing the temperature or by a chemical or pharmacologic chaperone). Together, these findings suggest that tsPINK1 may represent a valuable tool for the analysis of the PINK1/Parkin pathway in human cells; additionally, as the serine residue promoting thermal lability is conserved among Mus musculus, Danio rerio, Drosophila melanogaster and Caenorhabditis elegans, it may serve as the basis for developing other temperature-sensitive models for the study of recessive parkinsonism and mitophagy. Finally, these results suggest that PINK1 kinase function could be restored for a subset of patients with PINK1 mutations, and perhaps alter the course of their disease.
authors: Medical Research Council Mitochondrial Biology Unit, Hills Road, Cambridge, UK.
authors: Derek P
authors: DP
authors: Narendra
authors: Chunxin
authors: C
authors: Wang
authors: Richard J
authors: RJ
authors: Youle
authors: John E
authors: JE
authors: Walker
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D004789:Enzyme Activation; D015870:Gene Expression; D056726:Genetic Association Studies; D014644:Genetic Variation; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D020734:Parkinsonian Disorders / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D011487:Protein Conformation; D011494:Protein Kinases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D055550:Protein Stability; D021381:Protein Transport; D016415:Sequence Alignment; D015398:Signal Transduction; D013329:Structure-Activity Relationship; D013696:Temperature; D000099202:PTEN-Induced Putative Kinase
chemical_list: D011494:Protein Kinases; D000099202:PTEN-Induced Putative Kinase