Synaptic vesicle exocytosis

Abstract

Presynaptic nerve terminals release neurotransmitters by synaptic vesicle exocytosis. Membrane fusion mediating synaptic exocytosis and other intracellular membrane traffic is affected by a universal machinery that includes SNARE (for "soluble NSF-attachment protein receptor") and SM (for "Sec1/Munc18-like") proteins. During fusion, vesicular and target SNARE proteins assemble into an α-helical trans-SNARE complex that forces the two membranes tightly together, and SM proteins likely wrap around assembling trans-SNARE complexes to catalyze membrane fusion. After fusion, SNARE complexes are dissociated by the ATPase NSF (for "N-ethylmaleimide sensitive factor"). Fusion-competent conformations of SNARE proteins are maintained by chaperone complexes composed of CSPα, Hsc70, and SGT, and by nonenzymatically acting synuclein chaperones; dysfunction of these chaperones results in neurodegeneration. The synaptic membrane-fusion machinery is controlled by synaptotagmin, and additionally regulated by a presynaptic protein matrix (the "active zone") that includes Munc13 and RIM proteins as central components.

Figure 1.

The synaptic vesicle cycle. A presynaptic nerve terminal is depicted schematically as it contacts a postsynaptic neuron. The synaptic vesicle cycle consists of exocytosis (red arrows) followed by endocytosis and recycling (yellow arrows). Synaptic vesicles (green circles) are filled with neurotransmitters (NT; red dots) by active transport (neurotransmitter uptake) fueled by an electrochemical gradient established by a proton pump that acidifies the vesicle interior (vesicle acidification; green background). In preparation to synaptic exocytosis, synaptic vesicles are docked at the active zone, and primed by an ATP-dependent process that renders the vesicles competent to respond to a Ca²⁺-signal. When an action potential depolarizes the presynaptic membrane, Ca²⁺-channels open, causing a local increase in intracellular Ca²⁺ at the active zone that triggers completion of the fusion reaction. Released neurotransmitters then bind to receptors associated with the postsynaptic density (PSD). After fusion pore opening, synaptic vesicles probably recycle via three alternative pathways: local refilling with neurotransmitters without undocking (“kiss-and-stay”), local recycling with undocking (“kiss-and-run”), and full recycling of vesicles with passage through an endosomal intermediate. (Adapted from Südhof 2004.)

Figure 2.

The SNARE/SM protein cycle. The diagram on top depicts SNARE and SM proteins prior to fusion, when they are localized to the membranes either as natively unfolded proteins (e.g., the R-SNARE synaptobrevin/VAMP) or as proteins folded in complexes distinct from canonical SNARE complexes (e.g., binding of the SM protein Munc18-1 to the closed conformation of syntaxin-1 as shown here, or as syntaxin-1/SNAP-25 heterodimeric complexes). During priming, SNARE proteins partially zipper up into trans-complexes, and the SM protein associates with the trans-complexes by binding to the syntaxin amino terminus (left diagram). Full SNARE-complex assembly then pulls the membranes apart, opening the fusion pore (bottom diagram), which expands such that the vesicle membrane collapses into the target membrane, and the trans-SNARE complexes are converted into cis-SNARE complexes (right diagram). Afterward, cis-SNARE complexes are dissociated by the ATPase NSF acting in conjunction with its adaptors α/β/γ-SNAPs (no relation to SNAP-25 and its homologs—an unfortunate coincidence of acronyms), and vesicles recycle to start another round of the cycle.

Figure 3.

Structures of synaptic SNARE and SM proteins. (A) Schematic diagram of the domain structures of syntaxin, SNAP-25, and synaptobrevin/VAMP (Habc, Habc-domain; TM, transmembrane region). (B,C) Cartoon of the two modes of interaction of the SM protein Munc18 with SNARE proteins during synaptic exocytosis: Binding of Munc18 to the closed conformation of syntaxin-1 that occludes the SNARE motif (B), and binding of Munc18 to assembling SNARE trans-complexes that depends on the syntaxin-1 amino terminus (C). Note that the precise mode of Munc18 binding to assembling SNARE complexes is unknown, apart from the fact that it is anchored by interaction of the syntaxin amino terminus (indicated by an N) with the N-lobe of Munc18; the arrow indicates the uncertain atomic nature of this binding, which may involve wrapping of Munc18 around the SNARE helical bundle analogous to the binding of Munc18 to the closed conformation of syntaxin (Südhof and Rothman 2009). (i) Atomic structures of the fully assembled SNARE complex, the synaxin-1A Habc domain, and Munc18 containing a bound syntaxin amino-terminal peptide (blue), drawn to scale. (Data for structures are from Sutton et al. 1998, Fernandez et al. 1998, and Hu et al. 2011, respectively.) Arrow indicates uncertainty in how precisely Munc18 binds to SNARE complexes apart from the interaction of the syntaxin amino terminus with the Munc18 N-lobe.

Figure 4.

Two types of chaperones support SNARE protein function. Synaptic SNARE proteins are subject to continuous folding and unfolding reactions, creating the potential for misfolding that is counteracted by at least two SNARE chaperones implicated in neurodegeneration: the classical chaperone complex containing CSPα (for cysteine-string protein α), Hsc70, and SGT (red shapes), and the nonclassical chaperones α/β/γ-synucleins (yellow shapes). CSPα and synucleins are vesicle proteins that act by distinct mechanisms. CSPα forms a complex with Hsc70 and SGT on the vesicle that binds to SNAP-25 on the target membrane and supports the functional competence of SNAP-25 to engage in SNARE complexes, whereas synucleins are bound to phospholipids and synaptobrevin (Syb)/VAMP on the vesicles, and bind to assembling SNARE complexes to support their folding.

Figure 5.

Model of active zone protein function. A complex composed of four out of the six canonical components of active zones (Munc13, α-liprins, RIMs, and RIM-BPs) is shown; the remaining two canonical active zone proteins (ELKS and Piccolo/Bassoon) bind peripherally to the complex, and are not shown. The active zone complex is formed by an interaction of the amino-terminal zinc-finger domain of RIM with the amino-terminal C2A-domain of Munc13; by an interaction of a proline-rich sequence in RIM with a RIM-BP SH3-domain, and by a binding of α-liprins to the RIM C2B-domain. The complex is anchored on synaptic vesicles via RIM binding to Rab3 and possibly also via RIM binding synaptotagmin (not shown), and on the plasma membrane via direct binding of RIM and of RIM-BP to N- and P/Q-type Ca²⁺-channels. Note that there are likely additional interactions among the various active zone proteins, and between these proteins and the presynaptic plasma membrane. Ca²⁺-ions are shown as dots; most of the active zone proteins contain C2-domains similar to synaptotagmin, but only a subset of the C2-domains bind Ca²⁺.