Abstract
Syntaxin-1 (STX1) and Munc18-1 are two requisite components of synaptic vesicular release machinery, so much so synaptic transmission cannot proceed in their absence. They form a tight complex through two major binding modes: through STX1's N-peptide and through STX1's closed conformation driven by its Habc- domain. However, physiological roles of these two reportedly different binding modes in synapses are still controversial. Here we characterized the roles of STX1's N-peptide, Habc-domain, and open conformation with and without N-peptide deletion using our STX1-null mouse model system and exogenous reintroduction of STX1A mutants. We show, on the contrary to the general view, that the Habc-domain is absolutely required and N-peptide is dispensable for synaptic transmission. However, STX1A's N-peptide plays a regulatory role, particularly in the Ca2+-sensitivity and the short-term plasticity of vesicular release, whereas STX1's open conformation governs the vesicle fusogenicity. Strikingly, we also show neurotransmitter release still proceeds when the two interaction modes between STX1A and Munc18-1 are presumably intervened, necessitating a refinement of the conceptualization of STX1A-Munc18-1 interaction.
Highlights
The synaptic vesicle (SV) fusion is the fundamental process in synaptic transmission, and it is catalyzed by the merger of plasma and vesicular membranes by the neuronal SNAREs syntaxin-1 (STX1 collectively refers to STX1A and STX1B throughout this study), synaptobrevin-2 (Syb-2 ), and SNAP25 (Rizo and Sudhof, 2012; Rizo and Xu, 2015; Baker and Hughson, 2016)
We utilized immunocytochemistry in high-d ensity hippocampal neuronal culture to quantify the exogenous expression of STX1A∆N2-9, STX1A∆Habc, and STX1ALEOpen at presynaptic compartments as defined by Bassoon-positive puncta and normalized fluorescence signals to the signals caused by expression of STX1AWT, all in STX1-n ull neurons
Loss of STX1 leads to a severe reduction in Munc18-1 expression, which can be rescued by the expression of either STX1A or STX1B (Zhou et al, 2013; Vardar et al, 2016; Vardar et al, 2020)
Summary
The synaptic vesicle (SV) fusion is the fundamental process in synaptic transmission, and it is catalyzed by the merger of plasma and vesicular membranes by the neuronal SNAREs syntaxin-1 (STX1 collectively refers to STX1A and STX1B throughout this study), synaptobrevin-2 (Syb-2 ), and SNAP25 (Rizo and Sudhof, 2012; Rizo and Xu, 2015; Baker and Hughson, 2016). STX1 is the most important neuronal SNARE because synaptic transmission grinds to a halt in its absence, and neurons cannot survive (Vardar et al, 2016). Besides its interaction with the other SNAREs, STX1 binds to its cognate SM protein Munc18-1 forming a tight binary complex with an affinity in the nanomolar range (Pevsner et al, 1994; Burkhardt et al, 2008). Munc18-1 , which is an assistor of SNARE-m ediated vesicular release, is an important protein as its absence leads to inhibition of synaptic transmission (Verhage et al, 2000). Two major modes for STX1 binding to Munc18-1 have been defined: one through its N-p eptide, the other through its closed conformation driven by the intramolecular interaction between its Habc- and SNARE domains (Dulubova et al, 1999; Misura et al, 2000). Several issues regarding these reportedly different binding modes of STX1 to Munc18-1 are still subjects of dispute
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