Neuronal SNAREs and their key regulators together drive synaptic vesicle exocytosis and synaptic transmission as a single integrated membrane fusion machine. Human pathogenic mutations have now been ...reported for all eight core components, but patients are diagnosed with very different neurodevelopmental syndromes. We propose to unify these syndromes, based on etiology and mechanism, as “SNAREopathies.” Here, we review the strikingly diverse clinical phenomenology and disease severity and the also remarkably diverse genetic mechanisms. We argue that disease severity generally scales with functional redundancy and, conversely, that the large effect of mutations in some SNARE genes is the price paid for extensive integration and exceptional specialization. Finally, we discuss how subtle differences in components being rate limiting in different types of neurons helps to explain the main symptoms.
Verhage and Sørensen review the diversity in genetic mechanisms and clinical manifestations of syndromes caused by mutations in the presynaptic fusion machinery. They argue that the redundancy and phenotypic heterogeneity among neurons help to explain this diversity.
The molecular mechanisms for calcium‐triggered membrane fusion have long been sought for, and detailed models now exist that account for at least some of the functions of the many proteins involved ...in the process. Key players in the fusion reaction are a group of proteins that, upon binding to calcium, trigger the merger of cargo‐filled vesicles with the plasma membrane. Low‐affinity, fast‐kinetics calcium sensors of the synaptotagmin family – especially synaptotagmin‐1 and synaptotagmin‐2 – are the main calcium sensors for fast exocytosis triggering in many cell types. Their functions extend beyond fusion triggering itself, having been implicated in the calcium‐dependent vesicle recruitment during activity, docking of vesicles to the plasma membrane and priming, and even in post‐fusion steps, such as fusion pore expansion and endocytosis. Furthermore, synaptotagmin diversity imparts distinct properties to the release process itself. Other calcium‐sensing proteins such as Munc13s and protein kinase C play important, but more indirect roles in calcium‐triggered exocytosis. Because of their higher affinity, but intrinsic slower kinetics, they operate on longer temporal and spatial scales to organize assembly of the release machinery. Finally, the high‐affinity synaptotagmin‐7 and Doc2 (Double C2‐domain) proteins are able to trigger membrane fusion in vitro, but cellular measurements in different systems show that they may participate in either fusion or vesicle priming. Here, we summarize the properties and possible interplay of (some of) the major C2‐domain containing calcium sensors in calcium‐triggered exocytosis.
This article is part of a mini review series: “Synaptic Function and Dysfunction in Brain Diseases”.
C2‐domain containing calcium sensors are involved in releasing the vesicular content in secretory cells. Some sensors with a low calcium affinity and fast kinetics are optimally suited for triggering the release itself, whereas others work in the priming of vesicles. The interplay of several kinds of sensors give rise to the complex release kinetics found in secretory cells.
This article is part of a mini review series: “Synaptic Function and Dysfunction in Brain Diseases”.
Exocytosis requires formation of SNARE soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor complexes between vesicle and target membranes. Recent assessments in reduced model ...systems have produced divergent estimates of the number of SNARE complexes needed for fusion. Here, we used a titration approach to answer this question in intact, cultured chromaffin cells. Simultaneous expression of wild-type SNAP-25 and a mutant unable to support exocytosis progressively altered fusion kinetics and fusion-pore opening, indicating that both proteins assemble into heteromeric fusion complexes. Expressing different wild-type:mutant ratios revealed a third-power relation for fast (synchronous) fusion and a near-linear relation for overall release. Thus, fast fusion typically observed in synapses and neurosecretory cells requires at least three functional SNARE complexes, whereas slower release might occur with fewer complexes. Heterogeneity in SNARE-complex number may explain heterogeneity in vesicular release probability.
The SNAP-25 Protein Family Kádková, Anna; Radecke, Julika; Sørensen, Jakob B.
Neuroscience,
11/2019, Letnik:
420
Journal Article
Recenzirano
•We review the expression, function and regulation of the SNAP-25 protein family.•SNAP-25 and SNAP-23 are specialized for regulated exocytosis.•SNAP-25 couples to Ca2+-sensors and drives ...Ca2+-triggered exocytosis in neuronal cells.•SNAP-29 is involved in constitutive exocytosis and autophagosome fusion.•SNAP-47 drives AMPA-receptor trafficking and autophagosome fusion.
SNARE-complexes drive the fusion of membrane-bound vesicles with target membranes or with each other (homotypic fusion). The SNARE-proteins are subdivided into Qa, Qb, Qc and R-SNAREs depending on their position in the four-helical SNARE-bundle. Here, we review the SNAP-25 protein sub-family, which includes both the Qb and Qc SNARE-domains within a single protein. In vertebrates, this sub-family consists of SNAP-25, SNAP-23, SNAP-29 and SNAP-47, named for their apparent molecular weights. SNAP-25 and SNAP-23 are specialized for driving regulated exocytosis. SNAP-25 performs this function in the nervous system, and in neuroendocrine cells, where fast Ca2+-dependent triggering is required in order to synchronize release with an electrical signal, whereas SNAP-23 drives regulated exocytosis in most other cases that have been studied, e.g. platelet exocytosis or glucose transporter trafficking. SNAP-25 is regulated by alternative splicing, phosphorylation and by G-protein binding, and it regulates Ca2+-channels, neuronal survival and postsynaptic spine development. SNAP-23 is primarily regulated by phosphorylation within the linker connecting Qb to Qc. Cross-rescue experiments show that SNAP-25 and SNAP-23 can (at least partly) substitute for each other, whereas SNAP-29 and SNAP-47 cannot. SNAP-29 is present on intracellular membranes and performs functions in autophagosome-to-lysosome fusion, among others. An overlapping function for SNAP-47 was described; in addition, SNAP-47 mediates postsynaptic AMPA-receptor insertion. Overall, the presence of two SNARE-domains confers members of this family the ability to associate to different Qa and R-SNAREs and drive diverse membrane fusion reactions; one member of the family, SNAP-25, has been devoted entirely to Ca2+-triggered fusion and has taken on a number of additional, regulatory roles.
Vesicle Docking in Regulated Exocytosis Verhage, Matthijs; Sørensen, Jakob B
Traffic (Copenhagen, Denmark),
September 2008, Letnik:
9, Številka:
9
Journal Article
Recenzirano
Odprti dostop
In electron micrographs, many secretory and synaptic vesicles are found 'docked' at the target membrane, but it is unclear why and how. It is generally assumed that docking is a necessary first step ...in the secretory pathway before vesicles can acquire fusion competence (through 'priming'), but recent studies challenge this. New biophysical methods have become available to detect how vesicles are tethered at the target membrane, and genetic manipulations have implicated many genes in tethering, docking and priming. However, these studies have not yet led to consistent working models for these steps. In this study, we review recent attempts to characterize these early steps and the cellular factors to orchestrate them. We discuss whether assays for docking, tethering and priming report on the same phenomena and whether all vesicles necessarily follow the same linear docking-priming-fusion pathway. We conclude that most evidence to date is consistent with such a linear pathway assuming several refinements that imply that some vesicles can be nonfunctionally docked ('dead-end' docking) or, conversely, that the linear pathway can be greatly accelerated (crash fusion).
Synchronization of neurotransmitter release with the presynaptic action potential is essential for maintaining fidelity of information transfer in the central nervous system. However, synchronous ...release is frequently accompanied by an asynchronous release component that builds up during repetitive stimulation, and can even play a dominant role in some synapses. Here, we show that substitution of SNAP-23 for SNAP-25 in mouse autaptic glutamatergic hippocampal neurons results in asynchronous release and a higher frequency of spontaneous release events (mEPSCs). Use of neurons from double-knock-out (SNAP-25, synaptotagmin-7) mice in combination with viral transduction showed that SNAP-23-driven release is triggered by endogenous synaptotagmin-7. In the absence of synaptotagmin-7 release became even more asynchronous, and the spontaneous release rate increased even more, indicating that synaptotagmin-7 acts to synchronize release and suppress spontaneous release. However, compared to synaptotagmin-1, synaptotagmin-7 is a both leaky and asynchronous calcium sensor. In the presence of SNAP-25, consequences of the elimination of synaptotagmin-7 were small or absent, indicating that the protein pairs SNAP-25/synaptotagmin-1 and SNAP-23/synaptotagmin-7 might act as mutually exclusive calcium sensors. Expression of fusion proteins between pHluorin (pH-sensitive GFP) and synaptotagmin-1 or -7 showed that vesicles that fuse using the SNAP-23/synaptotagmin-7 combination contained synaptotagmin-1, while synaptotagmin-7 barely displayed activity-dependent trafficking between vesicle and plasma membrane, implying that it acts as a plasma membrane calcium sensor. Overall, these findings support the idea of alternative syt∶SNARE combinations driving release with different kinetics and fidelity.
Fast exocytosis of synaptic vesicles differs from other membrane fusion reactions by being under tight temporal control by the intracellular calcium concentration. This is achieved by subjecting the ...SNARE-dependent fusion pathway to additional layers of control, both upstream and downstream of the assembly of the fusogenic SNARE-complex. Here, I review conflicting views on the function of the core fusion machinery consisting of the SNAREs, Munc18, complexin, and synaptotagmin. Munc18 controls docking of vesicles to the plasma membrane and initial SNARE-complex assembly, whereas complexin and synaptotagmin cooperate in holding the SNARE complex in an intermediate release-ready or cocked state. Different effects of complexin and synaptotagmin shape the energy landscape for fusion and make final fusion calcium triggered. The final steps are fusion pore formation and expansion, which allow release of the water-soluble vesicle content. The fusion pore remains the most elusive part of the exocytosis pathway, owing to its short lifetime.
The SNAP receptor (SNARE) complex, consisting of synaptosome-associated protein of 25 kDa (SNAP-25), synaptobrevin-2, and syntaxin-1, is involved in synaptic vesicles exocytosis. In addition, SNAP-25 ...has been implicated in constitutive exocytosis processes required for neurite outgrowth. However, at least three isoforms of SNAP-25 have been reported from neurons: SNAP-23, which is also present in non-neuronal cells, and the two alternative splice variants SNAP-25a and SNAP-25b. Here, we studied the differential ability of these isoforms to support the functions previously broadly ascribed to "SNAP-25." We studied the rescue of snap-25 null neurons in culture with different SNAP-25 homologs. We find that deletion of SNAP-25 leads to strongly reduced neuron survival, and, in the few surviving cells, impaired arborization, reduced spontaneous release, and complete arrest of evoked release. Lentiviral expression of SNAP-25a, SNAP-25b, or SNAP-23 rescued neuronal survival, arborization, amplitude, and frequency of spontaneous events. Also evoked release was rescued by all isoforms, but synchronous release required SNAP-25a/b in both glutamatergic and GABAergic neurons. SNAP-23 supported asynchronous release only, reminiscent of synaptotagmin-1 null neurons. SNAP-25b was superior to SNAP-25a in vesicle priming, resembling the shift to larger releasable vesicle pools that accompanies synaptic maturation. These data demonstrate a differential ability of SNAP-25b, SNAP-25a, and SNAP-23 to support neuronal function.
Presynaptic activation of the diacylglycerol (DAG)/protein kinase C (PKC) pathway is a central event in short-term synaptic plasticity. Two substrates, Munc13-1 and Munc18-1, are essential for ...DAG-induced potentiation of vesicle priming, but the role of most presynaptic PKC substrates is not understood. Here, we show that a mutation in synaptotagmin-1 (Syt1T112A), which prevents its PKC-dependent phosphorylation, abolishes DAG-induced potentiation of synaptic transmission in hippocampal neurons. This mutant also reduces potentiation of spontaneous release, but only if alternative Ca2+ sensors, Doc2A/B proteins, are absent. However, unlike mutations in Munc13-1 or Munc18-1 that prevent DAG-induced potentiation, the synaptotagmin-1 mutation does not affect paired-pulse facilitation. Furthermore, experiments to probe vesicle priming (recovery after train stimulation and dual application of hypertonic solutions) also reveal no abnormalities. Expression of synaptotagmin-2, which lacks a seven amino acid sequence that contains the phosphorylation site in synaptotagmin-1, or a synaptotagmin-1 variant with these seven residues removed (Syt1Δ109–116), supports normal DAG-induced potentiation. These data suggest that this seven residue sequence in synaptotagmin-1 situated in the linker between the transmembrane and C2A domains is inhibitory in the unphosphorylated state and becomes permissive of potentiation upon phosphorylation. We conclude that synaptotagmin-1 phosphorylation is an essential step in PKC-dependent potentiation of synaptic transmission, acting downstream of the two other essential DAG/PKC substrates, Munc13-1 and Munc18-1.
The functions of four of the five proteins in the mammalian uncoordinated-13 (Munc13) family have been identified as priming factors in SNARE-dependent exocytosis. In this issue, Zhang et al. (2017. ...J. Cell Biol. https://doi.org/10.1083/jcb.201702099) show that the fifth member, BAIAP3 (brain-specific angiogenesis inhibitor I–associated protein 3), acts in retrograde trafficking by returning secretory vesicle material to the trans-Golgi network. In its absence, secretory vesicle formation is impaired, leading to accumulation of immature vesicles, or lysosomal vesicle degradation.
PDF
