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Munc13 structural transitions and oligomers that may choreograph successive stages in vesicle priming for neurotransmitter release.

ABSTRACT: How can exactly six SNARE complexes be assembled under each synaptic vesicle? Here we report cryo-EM crystal structures of the core domain of Munc13, the key chaperone that initiates SNAREpin assembly. The functional core of Munc13, consisting of C1-C2B-MUN-C2C (Munc13C) spontaneously crystallizes between phosphatidylserine-rich bilayers in two distinct conformations, each in a radically different oligomeric state. In the open conformation (state 1), Munc13C forms upright trimers that link the two bilayers, separating them by ∼21 nm. In the closed conformation, six copies of Munc13C interact to form a lateral hexamer elevated ∼14 nm above the bilayer. Open and closed conformations differ only by a rigid body rotation around a flexible hinge, which when performed cooperatively assembles Munc13 into a lateral hexamer (state 2) in which the key SNARE assembly-activating site of Munc13 is autoinhibited by its neighbor. We propose that each Munc13 in the lateral hexamer ultimately assembles a single SNAREpin, explaining how only and exactly six SNARE complexes are templated. We suggest that state 1 and state 2 may represent two successive states in the synaptic vesicle supply chain leading to "primed" ready-release vesicles in which SNAREpins are clamped and ready to release (state 3).

SUBMITTER: Grushin K

PROVIDER: S-EPMC8851502 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

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Munc13 structural transitions and oligomers that may choreograph successive stages in vesicle priming for neurotransmitter release.

Grushin Kirill K Kalyana Sundaram R Venkat RV Sindelar Charles V CV Rothman James E JE

Proceedings of the National Academy of Sciences of the United States of America 20220201 7

How can exactly six SNARE complexes be assembled under each synaptic vesicle? Here we report cryo-EM crystal structures of the core domain of Munc13, the key chaperone that initiates SNAREpin assembly. The functional core of Munc13, consisting of C1-C2B-MUN-C2C (Munc13C) spontaneously crystallizes between phosphatidylserine-rich bilayers in two distinct conformations, each in a radically different oligomeric state. In the open conformation (state 1), Munc13C forms upright trimers that link the t ...[more]

PMID: 35135883

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Project description:Synaptotagmin-1 (SYT1) is a synaptic vesicle resident protein that interacts via its C2 domain with anionic lipids from the plasma membrane in a calcium-dependent manner to efficiently trigger rapid neurotransmitter (NT) release. In addition, SYT1 acts as a negative regulator of spontaneous NT release and regulates synaptic vesicle (SV) priming. How these functions relate to each other mechanistically and what role other synaptotagmin (SYT) isoforms play in supporting and complementing the role of SYT1 is still under intensive investigation. In this work, we analyzed three putative functions of SYT1 in exocytosis by systematically varying its expression in autaptic hippocampal glutamatergic neurons from mice of either sex. We find that regulation of release probability is most sensitive to variation of expression levels, whereas its impact on vesicle priming is least sensitive. Also, loss of SYT1 phenotypes on spontaneous release and vesicle priming is compensated in less mature synaptic cultures by redundant support from SYT7. Overall, our data help in resolving some controversies in SYT1 functions in exocytosis and in our understanding of how SYT1 contributes to the pathophysiology underlying SYT1-related human neurologic disorders.SIGNIFICANCE STATEMENT Our work clarifies the functions of SYT1 protein in synaptic vesicle priming and spontaneous and calcium-evoked neurotransmitter release and analyzes whether these occur at different stages of synaptic responses by examining their relative sensitivity to protein concentration at the synaptic terminal. We demonstrate that these synaptic functions are unequally sensitive to both protein levels and neuronal stage, indicating that they operate under distinct molecular mechanisms. Furthermore, we analyze how these functions are modulated by another synaptotagmin isoform expression. We show that to understand the phenotype displayed by SYT1 knock-out neurons (Syt1 -/-) is necessary to consider the interplay between SYT1 and SYT7 molecules at the presynaptic terminal.

Dataset Information

Munc13 structural transitions and oligomers that may choreograph successive stages in vesicle priming for neurotransmitter release.

Publications

Munc13 structural transitions and oligomers that may choreograph successive stages in vesicle priming for neurotransmitter release.

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