title: Leucine-rich repeat kinase 2 binds to neuronal vesicles through protein interactions mediated by its C-terminal WD40 domain.
abstract: Mutations in the leucine-rich repeat kinase 2 gene (LRRK2) are associated with familial and sporadic Parkinson's disease (PD). LRRK2 is a complex protein that consists of multiple domains, including predicted C-terminal WD40 repeats. In this study, we analyzed functional and molecular features conferred by the WD40 domain. Electron microscopic analysis of the purified LRRK2 C-terminal domain revealed doughnut-shaped particles, providing experimental evidence for its WD40 fold. We demonstrate that LRRK2 WD40 binds and sequesters synaptic vesicles via interaction with vesicle-associated proteins. In fact, a domain-based pulldown approach combined with mass spectrometric analysis identified LRRK2 as being part of a highly specific protein network involved in synaptic vesicle trafficking. In addition, we found that a C-terminal sequence variant associated with an increased risk of developing PD, G2385R, correlates with a reduced binding affinity of LRRK2 WD40 to synaptic vesicles. Our data demonstrate a critical role of the WD40 domain within LRRK2 function.
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Research Unit Protein Science, Neuherberg, Germany Institute of Neuroscience, National Research Council, Milan, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Piccoli
authors.affiliation: Department of Experimental Medicine, University of Genoa, Genoa, Italy.
authors.forename: Franco
authors.initials: F
authors.lastname: Onofri
authors.affiliation: Institute of Neuroscience, National Research Council, Milan, Italy.
authors.forename: Maria Daniela
authors.initials: MD
authors.lastname: Cirnaru
authors.affiliation: Center for Integrated Protein Science Munich and Technische Universität München, Department of Chemistry, Garching, Germany.
authors.forename: Christoph J O
authors.initials: CJ
authors.lastname: Kaiser
authors.affiliation: Center for Integrated Protein Science Munich and Technische Universität München, Department of Chemistry, Garching, Germany Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Structural Biology, Neuherberg, Germany.
authors.forename: Pravinkumar
authors.initials: P
authors.lastname: Jagtap
authors.affiliation: Center for Integrated Protein Science Munich and Technische Universität München, Department of Chemistry, Garching, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Kastenmüller
authors.affiliation: Institute of Neuroscience, National Research Council, Milan, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Pischedda
authors.affiliation: Department of Experimental Medicine, University of Genoa, Genoa, Italy.
authors.forename: Antonella
authors.initials: A
authors.lastname: Marte
authors.affiliation: Eberhard Karls University Tübingen, Institute for Ophthalmic Research, Medical Proteome Center, Tübingen, Germany.
authors.forename: Felix
authors.initials: F
authors.lastname: von Zweydorf
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Research Unit Protein Science, Neuherberg, Germany Eberhard Karls University Tübingen, Institute for Ophthalmic Research, Medical Proteome Center, Tübingen, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Vogt
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Developmental Genetics, Neuherberg, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Giesert
authors.affiliation: Division of Life Science, Centre of Systems Biology and Human Health, Institute for Advanced Study and School of Science, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
authors.forename: Lifeng
authors.initials: L
authors.lastname: Pan
authors.affiliation: Department of Biotechnology and Translational Medicine, University of Milan and Humanitas Clinical and Research Center, Rozzano, Italy.
authors.forename: Flavia
authors.initials: F
authors.lastname: Antonucci
authors.affiliation: CRG-EMBL System Biology Program, Centre de Regulació Genòmica UPF, Barcelona, Spain.
authors.forename: Christina
authors.initials: C
authors.lastname: Kiel
authors.affiliation: Division of Life Science, Centre of Systems Biology and Human Health, Institute for Advanced Study and School of Science, Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong.
authors.forename: Mingjie
authors.initials: M
authors.lastname: Zhang
authors.affiliation: Center for Integrated Protein Science Munich and Technische Universität München, Department of Chemistry, Garching, Germany.
authors.forename: Sevil
authors.initials: S
authors.lastname: Weinkauf
authors.affiliation: Center for Integrated Protein Science Munich and Technische Universität München, Department of Chemistry, Garching, Germany Helmholtz Zentrum München, German Research Center for Environmental Health, Institute of Structural Biology, Neuherberg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Sattler
authors.affiliation: Institute of Neuroscience, National Research Council, Milan, Italy.
authors.forename: Carlo
authors.initials: C
authors.lastname: Sala
authors.affiliation: Department of Biotechnology and Translational Medicine, University of Milan and Humanitas Clinical and Research Center, Rozzano, Italy.
authors.forename: Michela
authors.initials: M
authors.lastname: Matteoli
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Research Unit Protein Science, Neuherberg, Germany Eberhard Karls University Tübingen, Institute for Ophthalmic Research, Medical Proteome Center, Tübingen, Germany.
authors.forename: Marius
authors.initials: M
authors.lastname: Ueffing
authors.affiliation: Helmholtz Zentrum München, German Research Center for Environmental Health, Research Unit Protein Science, Neuherberg, Germany Eberhard Karls University Tübingen, Institute for Ophthalmic Research, Medical Proteome Center, Tübingen, Germany j.gloeckner@helmholtz-muenchen.de.
authors.forename: Christian Johannes
authors.initials: CJ
authors.lastname: Gloeckner
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D006801:Humans; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D051379:Mice; D008810:Mice, Inbred C57BL; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D009474:Neurons / Q000378:metabolism*; D009479:Neuropeptides / Q000378:metabolism; D009498:Neurotoxins / Q000633:toxicity; D010300:Parkinson Disease / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D025941:Protein Interaction Mapping; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry* / Q000378:metabolism* / Q000648:ultrastructure; D000075370:Receptors for Activated C Kinase; D013329:Structure-Activity Relationship; D013569:Synapses / Q000378:metabolism; D013572:Synaptic Vesicles / Q000378:metabolism*
chemical_list: D050505:Mutant Proteins; D009479:Neuropeptides; D009498:Neurotoxins; C090294:RACK1 protein, mouse; D000075370:Receptors for Activated C Kinase; C495280:LRRK2 protein, human; D000071158:Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; D017346:Protein Serine-Threonine Kinases