Molecular Basis of Coupling Calcium Sensing to Fast Membrane Fusion by Synaptotagmin-1 in Neurotransmitter Release

Neuronal communication depends on the rapid release of neurotransmitters through Ca2+-triggered synaptic vesicle exocytosis. Synaptotagmin-1 (Syt1) acts as the Ca2+ sensor for fast, synchronous neurotransmitter release. Despite decades of research, the precise molecular mechanism of Syt1 action and the coupling of Ca2+ sensing to membrane fusion remains elusive. Addressing these questions requires an understanding of the cooperation between Syt1 and the SNARE complex, which drives membrane fusion. The SNARE proteins anchored on opposing membranes bring them together by forming a tight four-helix bundle, a process referred to as zippering. A recent molecular model of SNARE-mediated membrane fusion has revealed that the zippering of the SNARE motifs into the juxta-membrane linkers, which catalyzes encounters of acyl chains from opposing bilayers, is the key event leading to membrane fusion. The work presented in this thesis provides a thorough analysis of the interactions between Syt1 and the SNARE complex in solution and on membranes. Leveraging NMR and fluorescence spectroscopy, the included studies systematize the often-contradictory data and present a comprehensive model illustrating how Ca2+ signaling is coupled to SNARE-mediated fusion. In our model, in the primed state of synaptic vesicles Syt1 binds the SNARE complex through a primary interface and the plasma membrane through a polybasic region. In the absence of Ca2+, Syt1 both enables the formation of a partially assembled SNARE complex yet strongly inhibits complete helical zippering and hence membrane fusion. Upon Ca2+ influx, Ca2+ binding by Syt1 induces its membrane insertion, leading to reorientation of the Syt1-SNARE complex on the plasma membrane. The reorientation generates a lever-like action of Syt1 that pulls the SNARE complex, facilitating the zippering of the linkers that induces fast membrane fusion and subsequent neurotransmitter release.