title: Systematic mutagenesis of α-synuclein reveals distinct sequence requirements for physiological and pathological activities.
abstract: α-Synuclein is an abundant presynaptic protein that binds to phospholipids and synaptic vesicles. Physiologically, α-synuclein functions as a SNARE-protein chaperone that promotes SNARE-complex assembly for neurotransmitter release. Pathologically, α-synuclein mutations and α-synuclein overexpression cause Parkinson's disease, and aggregates of α-synuclein are found as Lewy bodies in multiple neurodegenerative disorders ("synucleinopathies"). The relation of the physiological functions to the pathological effects of α-synuclein remains unclear. As an initial avenue of addressing this question, we here systematically examined the effect of α-synuclein mutations on its physiological and pathological activities. We generated 26 α-synuclein mutants spanning the entire molecule, and analyzed them compared with wild-type α-synuclein in seven assays that range from biochemical studies with purified α-synuclein, to analyses of α-synuclein expression in cultured neurons, to examinations of the effects of virally expressed α-synuclein introduced into the mouse substantia nigra by stereotactic injections. We found that both the N-terminal and C-terminal sequences of α-synuclein were required for its physiological function as SNARE-complex chaperone, but that these sequences were not essential for its neuropathological effects. In contrast, point mutations in the central region of α-synuclein, referred to as nonamyloid β component (residues 61-95), as well as point mutations linked to Parkinson's disease (A30P, E46K, and A53T) increased the neurotoxicity of α-synuclein but did not affect its physiological function in SNARE-complex assembly. Thus, our data show that the physiological function of α-synuclein, although protective of neurodegeneration in some contexts, is fundamentally distinct from its neuropathological effects, thereby dissociating the two activities of α-synuclein.
authors.affiliation: Department of Molecular and Cellular Physiology, Stanford University, Stanford, California 94305-5453, USA. burre@stanford.edu
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Burré
authors.forename: Manu
authors.initials: M
authors.lastname: Sharma
authors.forename: Thomas C
authors.initials: TC
authors.lastname: Südhof
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004195:Disease Models, Animal; D005786:Gene Expression Regulation / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D050356:Lipid Metabolism / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009069:Movement Disorders / Q000235:genetics; D016296:Mutagenesis / Q000235:genetics*; D009154:Mutation / Q000235:genetics*; D009474:Neurons; D010300:Parkinson Disease / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D011597:Psychomotor Performance / Q000502:physiology; D050600:SNARE Proteins / Q000378:metabolism; D013378:Substantia Nigra / Q000378:metabolism / Q000473:pathology; D050825:Synaptosomal-Associated Protein 25 / Q000378:metabolism; D050827:Syntaxin 1 / Q000378:metabolism; D051843:Synucleins / Q000737:chemistry / Q000172:deficiency / Q000235:genetics* / Q000378:metabolism; D014161:Transduction, Genetic; D014162:Transfection; D014446:Tyrosine 3-Monooxygenase / Q000378:metabolism; D050988:Vesicle-Associated Membrane Protein 2 / Q000172:deficiency / Q000378:metabolism
chemical_list: D050600:SNARE Proteins; D050825:Synaptosomal-Associated Protein 25; D050827:Syntaxin 1; D051843:Synucleins; D050988:Vesicle-Associated Membrane Protein 2; C494808:vesicle-associated membrane protein 2, mouse; D049452:Green Fluorescent Proteins; D014446:Tyrosine 3-Monooxygenase; D010751:Phosphopyruvate Hydratase