SNARE Force Synchronizes Synaptic Vesicle Fusion and Controls the Kinetics of Quantal Synaptic Transmission

Guzman, Raul E.; Schwarz, Yvonne; Rettig, Jens; Bruns, Dieter

doi:10.1523/jneurosci.1551-10.2010

Cited by 45 publications

(55 citation statements)

References 67 publications

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“…To study the nature of release activity in the absence of syb2 we grew murine hippocampal neurons from syb2 KO (Schoch et al 2001) and WT littermates on glial microislands and performed whole cell patch-clamp experiments on autaptic neurons. Consistent with previous findings (Guzman et al 2010), excitatory evoked release was virtually absent in the KO group (Fig. 1).…”

Section: Resultssupporting

confidence: 92%

Synaptobrevin 1 mediates vesicle priming and evoked release in a subpopulation of hippocampal neurons

Zimmermann

Trimbuch

Rosenmund

2014

Journal of Neurophysiology

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Zimmermann J, Trimbuch T, Rosenmund C. Synaptobrevin 1 mediates vesicle priming and evoked release in a subpopulation of hippocampal neurons. J Neurophysiol 112: 1559 -1565, 2014. First published June 18, 2014 doi:10.1152/jn.00340.2014.-The core machinery of synaptic vesicle fusion consists of three soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins, the two t-SNAREs at the plasma membrane (SNAP-25, Syntaxin 1) and the vesicle-bound v-SNARE synaptobrevin 2 (VAMP2). Formation of the trans-oriented four-␣-helix bundle between these SNAREs brings vesicle and plasma membrane in close proximity and prepares the vesicle for fusion. The t-SNAREs are thought to be necessary for vesicle fusion. Whether the v-SNAREs are required for fusion is still unclear, as substantial vesicle priming and spontaneous release activity remain in mammalian mass-cultured synaptobrevin/cellubrevindeficient neurons. Using the autaptic culture system from synaptobrevin 2 knockout neurons of mouse hippocampus, we found that the majority of cells were devoid of any evoked or spontaneous release and had no measurable readily releasable pool. A small subpopulation of neurons, however, displayed release, and their release activity correlated with the presence and amount of v-SNARE synaptobrevin 1 expressed. Comparison of synaptobrevin 1 and 2 in rescue experiments demonstrates that synaptobrevin 1 can substitute for the other v-SNARE, but with a lower efficiency in neurotransmitter release probability. Release activity in synaptobrevin 2-deficient mass-cultured neurons was massively reduced by a knockdown of synaptobrevin 1, demonstrating that synaptobrevin 1 is responsible for the remaining release activity. These data support the hypothesis that both t-and v-SNAREs are absolutely required for vesicle priming and evoked release and that differential expression of SNARE paralogs can contribute to differential synaptic coding in the brain. SNARE; neurotransmitter release; spontaneous release; release probability; short-term plasticity RELEASE OF NEUROTRANSMITTERS in the mammalian brain is mediated by fusion of synaptic vesicles (SVs) with the neuronal plasma membrane. This process has to be both fast and reliable and is performed by evolutionarily conserved protein machinery (Rizo and Rosenmund 2008;Sudhof 2013). Three proteins from the soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) family are considered to bring the two membranes in close proximity and carry out membrane fusion (Sollner et al. 1993;Weber et al. 1998). A v-SNARE (anchored in the SV membrane) and the t-SNAREs syntaxin (Bennett et al. 1992) and synaptosomal-associated protein 25 (SNAP-25) (Oyler et al. 1989) (both anchored in the plasma membrane) zipper up from their NH 2 to COOH terminus and thus form a four-␣-helix bundle called the trans-SNARE complex (Poirier et al. 1998;Sorensen et al. 2006;Sutton et al. 1998). Whether fusion of SVs is possible without SNARE proteins is still an open question. The major v-SNARE paralog on SVs i...

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Section: Resultssupporting

confidence: 92%

Synaptobrevin 1 mediates vesicle priming and evoked release in a subpopulation of hippocampal neurons

Zimmermann

Trimbuch

Rosenmund

2014

Journal of Neurophysiology

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“…However, an earlier as well as a recent study have shown that overexpression of N-or C-terminal GFP fusion constructs of syb2 in syb2-deficient hippocampal neurons fully rescues the amplitude and kinetics of evoked excitatory postsynaptic potentials in syb2-deficient neurons (32,33). SNARE assembly is believed to generate sufficient energy to drive membrane fusion (4,6,7).…”

Section: Discussionmentioning

confidence: 98%

“…Furthermore, deletion of syb2 alone, the predominant syb isoform in SVs, was shown to result in a 10-fold reduction of spontaneous fusion and a 100-fold decrease in fast Ca 2+ -triggered exocytosis in hippocampal synapses (31), thus providing a clean loss-of-function background for rescue experiments. Indeed, expression of syb2 fused to fluorescent proteins at the luminal domain in KO neurons was previously shown to rescue evoked exocytosis (32,33). Here, we first tested the ability of spH to rescue synaptic function in these hippocampal synapses (Fig.…”

Section: +mentioning

confidence: 99%

Two synaptobrevin molecules are sufficient for vesicle fusion in central nervous system synapses

Sinha¹,

Ahmed

Jahn

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

171

138

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Exocytosis of synaptic vesicles (SVs) during fast synaptic transmission is mediated by soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex assembly formed by the coil-coiling of three members of this protein family: vesicle SNARE protein, synaptobrevin 2 (syb2), and the presynaptic membrane SNAREs syntaxin-1A and SNAP-25. However, it is controversially debated how many SNARE complexes are minimally needed for SV priming and fusion. To quantify this effective number, we measured the fluorescence responses from single fusing vesicles expressing pHluorin (pHl), a pH-sensitive variant of GFP, fused to the luminal domain of the vesicular SNARE syb2 (spH) in cultured hippocampal neurons lacking endogenous syb2. Fluorescence responses were quantal, with the unitary signals precisely corresponding to single pHluorin molecules. Using this approach we found that two copies of spH per SV fully rescued evoked fusion whereas SVs expressing only one spH were unable to rapidly fuse upon stimulation. Thus, two syb2 molecules and likely two SNARE complexes are necessary and sufficient for SV fusion during fast synaptic transmission.synaptopHluorin | membrane fusion | single molecule bleaching | SNARE density I n conventional neuronal synapses, fast synaptic transmission is mediated by release of neurotransmitter upon Ca 2+ -triggered synaptic vesicle (SV) exocytosis. This process is exquisitely regulated both spatially and temporally. The core of the SV fusion machinery is formed by three members of the soluble N-ethylmaleimidesensitive factor attachment protein receptor (SNARE) protein family, which is characterized by conserved sequences of 60-70 amino acids called SNARE motifs: vesicle SNARE protein, synaptobrevin-2 (syb2), and the presynaptic membrane SNAREs syntaxin-1A and SNAP-25 (1-3).Zipper-like assembly of the SNARE motifs from their N-terminal ends toward their membrane-proximal C termini results in the formation of a highly stable heterotrimeric "trans-SNARE complex" (also called "SNAREpin"), consisting of four parallel α-helices, which brings the two membranes into close apposition for fusion (4-7). Previous studies have suggested that several of these SNARE complexes might assemble in rosette-shaped multivalent supercomplexes, forming a ring, around the fusion pore; however, there is no direct evidence in support of this model (5,8,9). Therefore, the precise number of SNARE complexes minimally required to drive membrane fusion is highly debated and current estimates range between 1 and 15 (10-18). Some of these results are based on single-molecule fluorescence measurements in artificially reconstituted liposomes, whereas others are based on theoretical models, kinetic analysis, and extrapolations from dose-response relationships. Therefore, it is essential to apply a more direct method capable of visualizing single SNARE complexes in real time in a physiological setting.In the present study we have used a unique direct approach to count the number of syb2 molecules required for fast...

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“…Hippocampal neurons were isolated as described previously 38 . Chromaffin cells were isolated as described previously 39,40 .…”

Section: Methodsmentioning

confidence: 99%

“…The methods used for electrophysiological recordings of hippocampal neurons 38,41 and chromaffin cells 40,42 were previously described.…”

Section: Methodsmentioning

confidence: 99%

Synaptobrevin2 is the v-SNARE required for cytotoxic T-lymphocyte lytic granule fusion

et al. 2013

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Cytotoxic T lymphocytes kill virus-infected and tumorigenic target cells through the release of perforin and granzymes via fusion of lytic granules at the contact site, the immunological synapse. It has been postulated that this fusion process is mediated by non-neuronal members of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor complex protein family. Here, using a synaptobrevin2-monomeric red fluorescence protein knock-in mouse we demonstrate that, surprisingly, the major neuronal v-SNARE synaptobrevin2 is expressed in cytotoxic T lymphocytes and exclusively localized on granzyme B-containing lytic granules. Cleavage of synaptobrevin2 by tetanus toxin or ablation of the synaptobrevin2 gene leads to a complete block of lytic granule exocytosis while leaving upstream events unaffected, identifying synaptobrevin2 as the v-SNARE responsible for the fusion of lytic granules at the immunological synapse.

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SNARE Force Synchronizes Synaptic Vesicle Fusion and Controls the Kinetics of Quantal Synaptic Transmission

Cited by 45 publications

References 67 publications

Synaptobrevin 1 mediates vesicle priming and evoked release in a subpopulation of hippocampal neurons

Synaptobrevin 1 mediates vesicle priming and evoked release in a subpopulation of hippocampal neurons

Two synaptobrevin molecules are sufficient for vesicle fusion in central nervous system synapses

Synaptobrevin2 is the v-SNARE required for cytotoxic T-lymphocyte lytic granule fusion

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