Circular oligomerization is an intrinsic property of synaptotagmin.

Jing Wang,James E Rothman,Feng Li,Frédéric Pincet,Oscar D Bello,Shyam S Krishnakumar,Charles Vaughn Sindelar

doi:10.7554/elife.27441

Abstract

Previously, we showed that synaptotagmin1 (Syt1) forms Ca2+-sensitive ring-like oligomers on membranes containing acidic lipids and proposed a potential role in regulating neurotransmitter release (Zanetti et al., 2016). Here, we report that Syt1 assembles into similar ring-like oligomers in solution when triggered by naturally occurring polyphosphates (PIP2 and ATP) and magnesium ions (Mg2+). These soluble Syt1 rings were observed by electron microscopy and independently demonstrated and quantified using fluorescence correlation spectroscopy. Oligomerization is triggered when polyphosphates bind to the polylysine patch in C2B domain and is stabilized by Mg2+, which neutralizes the Ca2+-binding aspartic acids that likely contribute to the C2B interface in the oligomer. Overall, our data show that ring-like polymerization is an intrinsic property of Syt1 with reasonable affinity that can be triggered by the vesicle docking C2B-PIP2 interaction and raise the possibility that Syt1 rings could pre-form on the synaptic vesicle to facilitate docking.

Highlights

IntroductionSynaptic vesicle (SV) fusion at the neuronal synapse is mediated by the SNARE (soluble NSF attachment protein receptor) complex, whose assembly is chaperoned by several other proteins to achieve precision and synchronicity of neurotransmitter release (Sudhof, 2013; Sollner et al, 1993a, 1993b)
Synaptic vesicle (SV) fusion at the neuronal synapse is mediated by the SNARE complex, whose assembly is chaperoned by several other proteins to achieve precision and synchronicity of neurotransmitter release (Sudhof, 2013; Sollner et al, 1993a, 1993b)
The minimal C2AB domains of Syt1 form circular oligomers on lipid surface and the electrostatic interaction between the conserved lysine residues (K326/K327) within the polybasic patch/motif on the C2B domain and the negatively-charged lipids, like PIP2 and PS, on the lipid surface was required for its assembly (Wang et al, 2014; Zanetti et al, 2016)

Summary

Introduction

Synaptic vesicle (SV) fusion at the neuronal synapse is mediated by the SNARE (soluble NSF attachment protein receptor) complex, whose assembly is chaperoned by several other proteins to achieve precision and synchronicity of neurotransmitter release (Sudhof, 2013; Sollner et al, 1993a, 1993b). SV-associated protein Syt is of special interest because it is the primary calcium (Ca2+) sensor that triggers the rapid, synchronous neurotransmitter release after Ca2+ are admitted into the pre-synaptic terminal following an action potential (Brose et al, 1992; Fernandez-Chacon et al, 2001; Geppert et al, 1994) It is well-established that the Ca2+-induced reorientation of the cytoplasmic C2 domains (C2A and C2B) of Syt into the membrane is driving force behind this process (Rhee et al, 2005; Paddock et al, 2011; Hui et al, 2006; Krishnakumar et al, 2013), but the precise molecular details are still unclear.

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