Vesicular proteins exocytosed and subsequently retrieved by compensatory endocytosis are nonidentical

Wienisch, Martin; Klingauf, Jürgen

doi:10.1038/nn1739

Cited by 221 publications

(327 citation statements)

References 49 publications

(60 reference statements)

Supporting

Mentioning

299

Contrasting

Order By: Relevance

“…1 A-C). Although a considerable fraction of the sybII constructs is localized in the plasma membrane as expected (18) and generates diffuse fluorescence, punctate fluorescence is clearly visible, indicating vesicular localization. Further evidence for vesicular localization comes from observation of vesicular movement (Movies S1-S3).…”

Section: Resultsmentioning

confidence: 53%

Role of the synaptobrevin C terminus in fusion pore formation

Ngatchou

Kisler

Fang

et al. 2010

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

106

View full text Add to dashboard Cite

Neurotransmitter release is mediated by the SNARE proteins synaptobrevin II (sybII, also known as VAMP2), syntaxin, and SNAP-25, generating a force transfer to the membranes and inducing fusion pore formation. However, the molecular mechanism by which this force leads to opening of a fusion pore remains elusive. Here we show that the ability of sybII to support exocytosis is inhibited by addition of one or two residues to the sybII C terminus depending on their energy of transfer from water to the membrane interface, following a Boltzmann distribution. These results suggest that following stimulation, the SNARE complex pulls the C terminus of sybII deeper into the vesicle membrane. We propose that this movement disrupts the vesicular membrane continuity leading to fusion pore formation. In contrast to current models, the experiments suggest that fusion pore formation begins with molecular rearrangements at the intravesicular membrane leaflet and not between the apposed cytoplasmic leaflets.chromaffin cell | patch clamp capacitance measurement | caged calcium | amperometry | electrochemical detector array T he SNARE proteins (1) mediate release of stored secretory products by exocytosis. In neurosecretion, the t-SNAREs syntaxin and SNAP-25 in the plasma membrane bind the v-SNARE synaptobrevin II (sybII, also known as VAMP2), which is anchored to the vesicle membrane by a single transmembrane (TM) domain. Upon stimulation, the SNARE complex is thought to zip up more tightly proceeding in a vectorial manner from the N to the C terminus, toward the TM domains of sybII and syntaxin (2-5), thereby transferring a force to the membranes (6). However, the molecular mechanism by which this force leads to opening of the fusion pore has not been determined (7). Several models have been proposed to explain the mechanism of fusion pore formation. In the lipid-stalk-hemifusion hypothesis, the outer and the inner leaflets of the two membranes merge via formation of a hemifusion intermediate in response to forces exerted by proteins surrounding the fusion site (8). In an alternative proteinaceous fusion pore model, the fusion pore is lined by the TM domain of syntaxin (9) and possibly synaptobrevin (10). However, it is not immediately evident how the hydrophobic transmembrane domains can line an aqueous fusion pore that allows for ion permeation by electrodiffusion (11). When the C-terminal SNARE domain interactions are reduced by mutating or deleting the C terminus of SNAP-25, or when flexible linkers are introduced between the sybII TM domain and its SNARE domain, the rate of exocytosis is reduced (12-16) and the flux of transmitter through the early fusion pore is decreased (16,17), consistent with a structural change in the fusion pore. In an attempt to interpret these findings, a proteolipidic fusion pore model has been proposed, in which the fusion pore is formed by a molecular complex of both lipids and SNARE proteins (17). However, even this model does not explain the molecular mechanism by which the N-to C-terminal zipp...

show abstract

Section: Resultsmentioning

confidence: 53%

Role of the synaptobrevin C terminus in fusion pore formation

Ngatchou

Kisler

Fang

et al. 2010

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

106

View full text Add to dashboard Cite

show abstract

“…Two recent studies found evidence for the existence of a pool of synaptic vesicle proteins on the plasma membrane that is selectively and readily retrieved after brief or mild stimuli (49,50). Partially preassembled endocytic structures waiting to be recovered after a stimulus could, in principle, account for the rapid endocytosis elicited under some conditions.…”

Section: Discussionmentioning

confidence: 99%

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Newton

Kirchhausen

Murthy

2006

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

220

190

View full text Add to dashboard Cite

The ability of synapses to sustain signal propagation relies on rapid recycling of transmitter-containing presynaptic vesicles. Clathrinand dynamin-mediated retrieval of vesicular membrane has an undisputed role in synaptic vesicle recycling. There is also evidence for other modes of vesicle retrieval, including bulk retrieval and the so-called kiss-and-run recycling. Whether dynamin in required for these other modes of synaptic vesicle endocytosis remains unclear. Here, we have tested the role of dynamin in synaptic vesicle endocytosis by using a small molecule called dynasore, which rapidly inhibits the GTPase activity of dynamin with high specificity. Endocytosis after sustained or brief stimuli was completely and reversibly blocked by dynasore in cultured hippocampal neurons expressing the fluorescent tracer synaptopHluorin. By contrast, dynasore had no effect on exocytosis. In the presence of dynasore, low-frequency stimulation led to sustained accumulation of synaptopHluorin and other vesicular proteins on the surface membrane at a rate predicted from net exocytosis. These vesicular components remained on surface membranes even after the stimulus was terminated, suggesting that all endocytic events rely on dynamin during low-frequency activity as well as in the period after it. Ultrastructural analysis revealed a reduction in the density of synaptic vesicles and the presence of endocytic structures only at synapses that were stimulated in the presence of dynasore. In sum, our data indicate that dynamin is essential for all forms of compensatory synaptic vesicle endocytosis including any kiss-andrun events.clathrin ͉ hippocampal ͉ vesicle recycling ͉ kiss-and-run D espite substantial progress in understanding the mechanistic basis of the synaptic vesicle cycle, several fundamental questions remain. Evidence for a ''classical'' pathway of vesicle recycling through clathrin-mediated endocytosis comes from a variety of studies and techniques, and many molecular players in this process have been identified (1-4). There is also evidence for other modes of vesicle retrieval, including bulk retrieval (5, 6) and the so-called kiss-and-run recycling (7-12). Mechanistic studies of kiss-and-run recycling have been difficult, in part, because of a lack of agreement on the exact definition of this process.The final mechanical step of the separation of a budding clathrincoated vesicle from the plasma membrane relies on the GTPase dynamin (13,14). Studies using a temperature-sensitive dynamin ortholog shibire in Drosophila have indicated that dynamin function is critical for synaptic vesicle endocytosis (15, 16). If and how dynamin is involved in alternate modes of vesicle recycling such as kiss-and-run is unclear (17-22), especially at small central synapses where these modes have been proposed (8,12,23,24). At several giant synapses, the role of dynamin in synaptic vesicle endocytosis has been assayed by using nonhydrolyzable analogs of GTP such as guanosine 5Ј-[␥-thio]triphosphate (GTP␥S) or peptides that interfere wit...

show abstract

“…These proteins are mobile and intermix with newly exocytosed synaptic vesicle proteins 44,45 . By contrast, super-resolution stimulated emission depletion (STED) microscopy (BOX 4) has suggested that newly exocytosed synaptic vesicle proteins are clustered at the plasma membrane 46 .…”

Section: Box 1 | Presynaptic Organization -Exocytic and Endocytic Zonesmentioning

confidence: 99%

“…This apparent contradiction could be explained by differences between pHluorin-tagged synaptic vesicle membrane proteins and their native endogenous counterparts 47 . Alternatively, synaptic vesicle proteins may undergo postexocytic reclustering within the periactive zone 44,45 (FIG. 4), a process that could become rate-limiting under conditions of sustained activity (see below).…”

Section: Box 1 | Presynaptic Organization -Exocytic and Endocytic Zonesmentioning

confidence: 99%

See 1 more Smart Citation

Protein scaffolds in the coupling of synaptic exocytosis and endocytosis

2011

View full text Add to dashboard Cite

Communication between nerve cells largely occurs at chemical synapses -specialized sites of cell-cell contact where electrical signals trigger the exocytic release of neurotransmitter, which in turn activates postsynaptic receptor channels. The efficacy with which such chemical signals are transmitted is crucial for the functioning of the nervous system. Modulation of neurotransmission enables nerve cells to respond to a vast range of stimulus patterns. Synaptic activity can also be tremendously diverse; from the fast synapses of the hippocampus, to slow neuropeptide-containing synapses. Indeed, some signalling pathways are active in the absence of stimulation, such as those involved in the processing of sensory information, which transmit tonically at high rates of up to 100 Hz or more 1,2 .Intriguing questions arise from these facts, including how high rates of neurotransmission can be maintained over long periods of time, and what limits the ability of a given synapse to release neurotransmitter during sustained periods of activity. Recent data indicate that in many synapses, exocytosis of neurotransmitter is coupled to endocytosis, and that synapses have evolved a specialized apparatus of scaffolding proteins to comply with these demands. Such scaffolds aid the temporal and spatial coupling of exocytosis and endocytosis, and are crucial for maintaining rapid neurotransmission during sustained activity 3 .Exocytosis of neurotransmitter is triggered by stimulusinduced calcium influx into the nerve terminal 4,5 and the subsequent fusion of synaptic vesicles with the presynaptic plasma membrane at specialized regions called active zones (BOX 1). Exocytosed synaptic vesicle membrane proteins and lipids in turn are recycled at the endocytic or periactive zone (BOX 1) that surrounds the release site, to restore functional synaptic vesicle pools for reuse and to ensure long-term functionality of the synapse 6-8 (FIG. 1). Depending on the type of synapse and the stimulation frequency, several modes of endocytosis with different time constants -for example, fast and slow endocytosisseem to operate 7,9,10 . Clathrin-mediated endocytosis arguably represents the main pathway of synaptic vesicle endocytosis 6,8,11 , although other modes -for example, fast kiss-and-run exocytosis and endocytosis -could operate in parallel.Although the machineries for exocytic membrane fusion 12,13 and for endocytic retrieval of synaptic vesicle membranes have been studied separately in some detail 6,14 , comparatively little is known about the coupling between exocytosis and endocytosis. Here we synthesize recent data from physiological, morphological and biochemical studies in several model systems into hypothetical models for scaffold-based mechanisms underlying exocytic-endocytic coupling.The synaptic vesicle cycle Synaptic vesicle pools. Some defining features of chemical synapses are the presence of one or several clusters of synaptic vesicles in the presynaptic nerve terminal, and ultrastructural specializations at the pre-and postsy...

show abstract

Vesicular proteins exocytosed and subsequently retrieved by compensatory endocytosis are nonidentical

Cited by 221 publications

References 49 publications

Role of the synaptobrevin C terminus in fusion pore formation

Role of the synaptobrevin C terminus in fusion pore formation

Inhibition of dynamin completely blocks compensatory synaptic vesicle endocytosis

Protein scaffolds in the coupling of synaptic exocytosis and endocytosis

Contact Info

Product

Resources

About