The Ca2+ sensor synaptotagmin-1 and the SNARE complex cooperate to trigger neurotransmitter release. Structural studies elucidated three distinct synaptotagmin-1-SNARE complex binding modes involving polybasic, primary and tripartite interfaces of synaptotagmin-1. We investigated these interactions using NMR and fluorescence spectroscopy. Synaptotagmin-1 binds to the SNARE complex through the polybasic and primary interfaces in solution. Ca2+-free synaptotagmin-1 binds to SNARE complexes anchored on PIP2-containing nanodiscs. R398Q/R399Q and E295A/Y338W mutations at the primary interface, which strongly impair neurotransmitter release, disrupt and enhance synaptotagmin-1-SNARE complex binding, respectively. Ca2+ induces tight binding of synaptotagmin-1 to PIP2-containing nanodiscs, disrupting synaptotagmin-1-SNARE interactions. Specific effects of mutations in the polybasic region on Ca2+-dependent synaptotagmin-1-PIP2-membrane interactions correlate with their effects on release. Our data suggest that synaptotagmin-1 binds to the SNARE complex through the primary interface and that Ca2+ releases this interaction, inducing PIP2/membrane binding and allowing cooperation between synaptotagmin-1 and the SNAREs in membrane fusion to trigger release.
The NMR data corresponding to Figs. 1-4 and corresponding figure supplements are publicly available at https://doi.org/10.5061/dryad.0zpc866vt. Source data are provided for all the FRET experiments shown in Figs. 5-9 and corresponding figure supplements.
NMR data in 'Ca2+-dependent release of Synaptotagmin-1 from the SNARE complex on phosphatidylinositol 4,5-bisphosphatecontaining membranes' by Voleti, Jaczynska and Rizo, eLife 2020Dryad Digital Repository, doi:10.5061/dryad.0zpc866vt.
- Josep Rizo
- Josep Rizo
- Rashmi Voleti
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
- Nils Brose, Max Planck Institute of Experimental Medicine, Germany
© 2020, Voleti et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.