title: Human R1441C LRRK2 regulates the synaptic vesicle proteome and phosphoproteome in a Drosophila model of Parkinson's disease.
abstract: Mutations in leucine-rich repeat kinase 2 (LRRK2) cause late-onset, autosomal dominant familial Parkinson`s disease (PD) and variation at the LRRK2 locus contributes to the risk for idiopathic PD. LRRK2 can function as a protein kinase and mutations lead to increased kinase activity. To elucidate the pathophysiological mechanism of the R1441C mutation in the GTPase domain of LRRK2, we expressed human wild-type or R1441C LRRK2 in dopaminergic neurons of Drosophila and observe reduced locomotor activity, impaired survival and an age-dependent degeneration of dopaminergic neurons thereby creating a new PD-like model. To explore the function of LRRK2 variants in vivo, we performed mass spectrometry and quantified 3,616 proteins in the fly brain. We identify several differentially-expressed cytoskeletal, mitochondrial and synaptic vesicle proteins (SV), including synaptotagmin-1, syntaxin-1A and Rab3, in the brain of this LRRK2 fly model. In addition, a global phosphoproteome analysis reveals the enhanced phosphorylation of several SV proteins, including synaptojanin-1 (pThr1131) and the microtubule-associated protein futsch (pSer4106) in the brain of R1441C hLRRK2 flies. The direct phosphorylation of human synaptojanin-1 by R1441C hLRRK2 could further be confirmed by in vitro kinase assays. A protein-protein interaction screen in the fly brain confirms that LRRK2 robustly interacts with numerous SV proteins, including synaptojanin-1 and EndophilinA. Our proteomic, phosphoproteomic and interactome study in the Drosophila brain provides a systematic analyses of R1441C hLRRK2-induced pathobiological mechanisms in this model. We demonstrate for the first time that the R1441C mutation located within the LRRK2 GTPase domain induces the enhanced phosphorylation of SV proteins in the brain.
authors: Silantes GmbH, Munich, Germany.|Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), Cologne, Germany.|Center for Neurodegenerative Science, Van Andel Research Institute, Grand Rapids, Michigan, USA.
authors: Md Shariful
authors: MS
authors: Islam
authors: Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), Cologne, Germany.
authors: Hendrik
authors: H
authors: Nolte
authors: Biochemistry II, Molecular Neurobiochemistry Faculty for Chemistry and Biochemistry Ruhr-University Bochum, NC 7/174 Universitaetsstraße 150, 44780 Bochum, Germany.
authors: Wright
authors: W
authors: Jacob
authors: CNRS, UMR6265 Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.|INRA, UMR1324 Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.|Université de Bourgogne Franche-Comté, UMR Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.
authors: Anna B
authors: AB
authors: Ziegler
authors: Stefanie
authors: S
authors: Pütz
authors: CNRS, UMR6265 Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.|INRA, UMR1324 Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.|Université de Bourgogne Franche-Comté, UMR Centre des Sciences du Goût et de l'Alimentation, F-21000 Dijon, France.
authors: Yael
authors: Y
authors: Grosjean
authors: Institute for Mitochondrial Diseases and Aging, Medical Faculty, University of Cologne, D-50931 Cologne, Germany.
authors: Karolina
authors: K
authors: Szczepanowska
authors: Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), Cologne, Germany.|Institute for Mitochondrial Diseases and Aging, Medical Faculty, University of Cologne, D-50931 Cologne, Germany.|Center for Molecular Medicine (CMMC), University of Cologne, Germany.
authors: Aleksandra
authors: A
authors: Trifunovic
authors: Max Planck Institute for Heart and Lung Research, Ludwigstr. 43, 61231 Bad Nauheim, Germany.
authors: Thomas
authors: T
authors: Braun
authors: Silantes GmbH, Munich, Germany.
authors: Hermann
authors: H
authors: Heumann
authors: Biochemistry II, Molecular Neurobiochemistry Faculty for Chemistry and Biochemistry Ruhr-University Bochum, NC 7/174 Universitaetsstraße 150, 44780 Bochum, Germany.
authors: Rolf
authors: R
authors: Heumann
authors: Bernhard
authors: B
authors: Hovemann
authors: Center for Neurodegenerative Science, Van Andel Research Institute, Grand Rapids, Michigan, USA.
authors: Darren J
authors: DJ
authors: Moore
authors: Institute for Genetics, Cologne Excellence Cluster on Cellular Stress Responses in Aging-Associated Diseases (CECAD), Cologne, Germany.|Center for Molecular Medicine (CMMC), University of Cologne, Germany.|Max Planck Institute for Heart and Lung Research, Ludwigstr. 43, 61231 Bad Nauheim, Germany.
authors: Marcus
authors: M
authors: Krüger
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001921:Brain / Q000378:metabolism* / Q000473:pathology; D004195:Disease Models, Animal; D059290:Dopaminergic Neurons / Q000378:metabolism* / Q000473:pathology; D029721:Drosophila Proteins / Q000096:biosynthesis / Q000235:genetics; D004331:Drosophila melanogaster / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2 / Q000096:biosynthesis / Q000235:genetics*; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics; D010300:Parkinson Disease / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D010744:Phosphoric Monoester Hydrolases / Q000096:biosynthesis / Q000235:genetics; D010766:Phosphorylation; D060066:Protein Interaction Maps; D020543:Proteome / Q000235:genetics*; D013572:Synaptic Vesicles / Q000235:genetics; D050863:Synaptotagmin I / Q000096:biosynthesis / Q000235:genetics; D050827:Syntaxin 1 / Q000096:biosynthesis / Q000235:genetics; D020692:rab3 GTP-Binding Proteins / Q000096:biosynthesis / Q000235:genetics
chemical_list: D029721:Drosophila Proteins; D009419:Nerve Tissue Proteins; D020543:Proteome; D050863:Synaptotagmin I; D050827:Syntaxin 1; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; C097702:synaptojanin; D010744:Phosphoric Monoester Hydrolases; C000607807:Rab3 protein, Drosophila; D020692:rab3 GTP-Binding Proteins