α-Synuclein assembles into higher-order multimers upon membrane binding to promote SNARE complex formation

Physiologically, alpha-synuclein chaperones soluble NSF attachment protein receptor (SNARE) complex assembly and may also perform other functions; pathologically, in contrast, alpha-synuclein misfolds into neurotoxic aggregates that mediate neurodegeneration and propagate between neurons. In neurons, alpha-synuclein exists in an equilibrium between cytosolic and membrane-bound states. Cytosolic alpha-synuclein appears to be natively unfolded, whereas membrane-bound alpha-synuclein adopts an alpha-helical conformation. Although the majority of studies showed that cytosolic alpha-synuclein is monomeric, it is unknown whether membrane-bound alpha-synuclein is also monomeric, and whether chaperoning of SNARE complex assembly by alpha-synuclein involves its cytosolic or membrane-bound state. Here, we show using chemical cross-linking and fluorescence resonance energy transfer (FRET) that alpha-synuclein multimerizes into large homomeric complexes upon membrane binding. The FRET experiments indicated that the multimers of membrane-bound alpha-synuclein exhibit defined intermolecular contacts, suggesting an ordered array. Moreover, we demonstrate that alpha-synuclein promotes SNARE complex assembly at the presynaptic plasma membrane in its multimeric membrane-bound state, but not in its monomeric cytosolic state. Our data delineate a folding pathway for alpha-synuclein that ranges from a monomeric, natively unfolded form in cytosol to a physiologically functional, multimeric form upon membrane binding, and show that only the latter but not the former acts as a SNARE complex chaperone at the presynaptic terminal, and may protect against neurodegeneration.