Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2008 Sep 12;321(5895):1507-10.
doi: 10.1126/science.1163174. Epub 2008 Aug 14.

Conformational switch of syntaxin-1 controls synaptic vesicle fusion

Stefan H Gerber  1 , Jong-Cheol Rah, Sang-Won Min, Xinran Liu, Heidi de Wit, Irina Dulubova, Alexander C Meyer, Josep Rizo, Marife Arancillo, Robert E Hammer, Matthijs Verhage, Christian Rosenmund, Thomas C Südhof
Affiliations

Conformational switch of syntaxin-1 controls synaptic vesicle fusion

Stefan H Gerber et al. Science. .

Abstract

During synaptic vesicle fusion, the soluble N-ethylmaleimide-sensitive factor-attachment protein receptor (SNARE) protein syntaxin-1 exhibits two conformations that both bind to Munc18-1: a "closed" conformation outside the SNARE complex and an "open" conformation in the SNARE complex. Although SNARE complexes containing open syntaxin-1 and Munc18-1 are essential for exocytosis, the function of closed syntaxin-1 is unknown. We generated knockin/knockout mice that expressed only open syntaxin-1B. Syntaxin-1B(Open) mice were viable but succumbed to generalized seizures at 2 to 3 months of age. Binding of Munc18-1 to syntaxin-1 was impaired in syntaxin-1B(Open) synapses, and the size of the readily releasable vesicle pool was decreased; however, the rate of synaptic vesicle fusion was dramatically enhanced. Thus, the closed conformation of syntaxin-1 gates the initiation of the synaptic vesicle fusion reaction, which is then mediated by SNARE-complex/Munc18-1 assemblies.

PubMed Disclaimer

Figures

Figure 1
Figure 1. Syntaxin-1AKO/syntaxin-1BOpen double mutant mice perish postnatally
A. Survival of syntaxin-1 mutant mice B. and C. Representative immunoblots (B) and levels of synaptic proteins (C) from syntaxin-1AWT/1BWT, -1AWT/1BOpen, -1AKO/1BWT, and -1AKO/1BOpen mutant mice determined by quantitative immunoblotting using 125I-labeled secondary antibodies (see also Table S1). D. Representative immunoblots (left) and quantitations (right) analyzing co-immunoprecipitation of Munc18-1, SNAP-25, synaptobrevin-2, and synaptotagmin-1 with syntaxin-1BWT and -1BOpen in Triton X-100 solubilized brain proteins. The amounts of co-immunoprecipitated Munc18-1, SNAP-25, synaptobrevin-2, and synaptotagmin-1 were determined by quantitative immunoblotting and normalized for the immunoprecipitated syntaxin-1. Data in C and D are means ± SEMs; *=p<0.05; **=p<0.01; ***=p<0.001 by Student's t-test compared to wild-type. Abbreviations: Synt., syntaxin; Syb-2, synaptobrevin-2; Synph-1, synaptophysin-1; Syt-1, synaptotagmin-1; VCP, p97/vasolin-containing protein; GDI, GDP dissociation inhibitor.
Figure 2
Figure 2. Syntaxin-1BOpen impairs chromaffin but not synaptic vesicle docking
A. and B. Representative electron micrographs of neurons cultured from syntaxin-1AKO mice containing wild-type syntaxin-1BWT (A) or ‘open’ syntaxin-1BOpen (B; scale bar, 250 nm). C. Number of docked vesicles per active zone (n = 3 experiments with 1BWT=49, 55, and 32 synapses; 1BOpen=21, 38, and 49 synapses; normalized for wild-type values). D. Plot of the cumulative distribution of docked vesicles per active zone (statistical significance with Kolmogorov-Simirnov test: p<0.01; 1BWT=136, 1BOpen=108 synapses). E. and F. Size of the postsynaptic density (E) and number of docked vesicles/length of postsynaptic density (F; both normalized for wild-type values). G. - J. Representative electron micrographs of chromaffin cells from control (G,H) and syntaxin-1BOpen (I,J) littermate mice at embryonic day E18 at two magnifications (G,I: bar, 1 μm; H,J: bar, 200 nm). K. Distribution of the distance of secretory granules from the plasma membrane in chromaffin cells from syntaxin-1BWT, -1BOpen, Munc18-1 KO, and synaptobrevin-2 KO mice (analyzed separately with wildtype controls and binned as indicated). L. Total number of secretory granules per chromaffin cell in syntaxin-1BWT or -1BOpen mice, and in Munc18-1 and synaptobrevin-2 KO mice (K and L: N=3 animals, n = 60 chromaffin cells). Data are means ± SEMs; **=p<0.01 by Student's t-test compared to wild-type.
Figure 3
Figure 3. Neurotransmitter release in syntaxin-1BOpen synapses
A.- C. Summary graphs of the frequency (A), amplitude (B), and charge (C) of spontaneous synaptic events (mEPSCs) D. - F. Representative traces (D) and mean EPSC amplitudes (E) and charges (F) in synaptic responses induced by isolated action potentials G. and H. EPSC amplitudes of evoked synaptic responses elicited by 10 Hz (G) and 20 Hz stimulus trains (H). Data are means ± SEMs; *=p<0.05 by Student's t-test compared to wild-type; numbers in bars show numbers of neurons analyzed.
Figure 4
Figure 4. Increased fusogenicity of synaptic vesicles in syntaxin-1BOpen synapses
A. Average postsynaptic currents elicited by application of 0.5 M sucrose in syntaxin-1BWT and -1BOpen synapses B. Mean RRP size determined as the transient charge integral induced by application of 0.5 M hypertonic solution C. Summary graph of the spontaneous vesicular release rate (mini frequency divided by the number of vesicles in the RRP) D. Mean vesicular release probability (Pvr = evoked EPSC charge divided by the RRP charge). E. Time-course of the average cumulative synaptic charge transfer during sucrose-induced release. For syntaxin-1BOpen synapses, both absolute (blue; left y-axis) and normalized responses (red; right y-axis) are depicted. In B and E, the steady-state component of release during the responses was subtracted. F. Plot of the half-width vs. time-to-onset of sucrose-induced synaptic responses. G. Representative traces of synaptic responses induced by 0.25 M, 0.35 M and 0.50 M sucrose. H. and I. Plot of the released fraction of the RRP (H; defined as the response to 0.5 M sucrose) and of the vesicular release rate (I) as a function of the sucrose concentration. In G, the 0 mM sucrose value represents the spontaneous vesicular release rate (Fig. 4D). Data are means ± SEMs; **=p<0.01, ***=p<0.001 by Student's t-test compared to wild-type; numbers in bars show numbers of neurons analyzed.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Brunger AT. Q. Rev. Biophys. 2005;9:1. - PubMed
    1. Jahn R, Scheller RH. Nat Rev Mol Cell Biol. 2006;7:631. - PubMed
    1. Khvotchev M, et al. J. Neurosci. 2007;27:12147. - PMC - PubMed
    1. Dulubova I, et al. EMBO J. 1999;18:4372. - PMC - PubMed
    1. Misura KM, Scheller RH, Weis WI. Nature. 2000;404:355. - PubMed

Publication types