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Presynapse Formation Assay Using Presynapse Organizer Beads and “Neuron Ball” Culture
Published on: August 2, 2019
Reconstitution of the vital functions of Munc18 and Munc13 in neurotransmitter release
Cong Ma1, Lijing Su, Alpay B Seven
1Key Laboratory of Molecular Biophysics, Ministry of Education, and Institute of Biophysics and Biochemistry, Huazhong University of Science and Technology, Wuhan 430074, China. cong.ma7@gmail.com
Abstract:
Neurotransmitter release depends critically on Munc18-1, Munc13, the Ca(2+) sensor synaptotagmin-1, and the soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP) receptors (SNAREs) syntaxin-1, synaptobrevin, and SNAP-25. In vitro reconstitutions have shown that syntaxin-1-SNAP-25 liposomes fuse efficiently with synaptobrevin liposomes in the presence of synaptotagmin-1-Ca(2+), but neurotransmitter release also requires Munc18-1 and Munc13 in vivo. We found that Munc18-1 could displace SNAP-25 from syntaxin-1 and that fusion of syntaxin-1-Munc18-1 liposomes with synaptobrevin liposomes required Munc13, in addition to SNAP-25 and synaptotagmin-1-Ca(2+). Moreover, when starting with syntaxin-1-SNAP-25 liposomes, NSF-α-SNAP disassembled the syntaxin-1-SNAP-25 heterodimers and abrogated fusion, which then required Munc18-1 and Munc13. We propose that fusion does not proceed through syntaxin-1-SNAP-25 heterodimers but starts with the syntaxin-1-Munc18-1 complex; Munc18-1 and Munc13 then orchestrate membrane fusion together with the SNAREs and synaptotagmin-1-Ca(2+) in an NSF- and SNAP-resistant manner.
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