Alternative titles; symbols
HGNC Approved Gene Symbol: STX1A
Cytogenetic location: 7q11.23 Genomic coordinates (GRCh38): 7:73,699,210-73,719,669 (from NCBI)
Intracellular vesicles travel among cellular compartments and deliver their specific cargo to target membranes by membrane fusion. The specificity of cargo delivery and membrane fusion is controlled, in part, by the pairing of vesicle v-SNAREs (soluble N-ethylmaleimide-sensitive factor attachment protein receptors) with target membrane t-SNAREs, such as STX1A (summary by McNew et al., 2000).
Bennett et al. (1992) identified two 35-kD proteins (p35 or syntaxins) that interact with synaptic vesicle protein p65 (synaptotagmin; 185605). The p35 proteins are expressed only in the nervous system. The 2 proteins are 84% identical, include C-terminal membrane anchors, and are concentrated on the plasma membrane at synaptic sites. The authors speculated that the p35 proteins may function in docking synaptic vesicles near calcium channels at presynaptic active zones.
Zhang et al. (1995) screened a human brain cDNA library with a partial rat STX1A cDNA. The isolated 2.1-kb clone predicted a 288-amino acid protein that was 98% identical to the rat protein. The authors referred to this gene as human STX1A.
Bennett et al. (1993) further characterized the syntaxin family of vesicular transport receptors in rats. Microinjection studies suggested that the rat nervous system-specific syntaxin-1A is important for calcium-regulated secretion from neuroendocrine PC12 cells.
By PCR analysis of human/rodent somatic cell hybrid panels and fluorescence in situ hybridization, Nakayama et al. (1997) mapped the STX1A gene to chromosome 7q11.2.
Zhang et al. (1995) noted that syntaxins, such as syntaxin-1A, function in the vesicle fusion process. Syntaxins also serve as substrates for botulinum neurotoxin type C, a metalloprotease that blocks exocytosis, and have high affinity for a molecular complex that includes the receptor for alpha-latrotoxin (600565), a substance that produces explosive exocytosis.
To elucidate the mechanisms that regulate neutrophil exocytosis, Martin-Martin et al. (1999) studied the expression of syntaxins in neutrophils. See syntaxin-6 (603944). The authors found that syntaxin-1A is expressed in neutrophils and peripheral blood lymphocytes. Immunoblotting of subcellular fractions and immunofluorescence analysis using an antibody against syntaxin-1A/1B (601485) revealed that syntaxin-1 is primarily located in the membranes of cytoplasmic granules in human resting neutrophils.
Naren et al. (2000) demonstrated that syntaxin-1A is expressed in airway epithelial cells, and is not a neural-specific protein. They also showed that syntaxin-1A regulates CFTR (602421) activity in airway epithelial cells.
McNew et al. (2000) tested all of the potential v-SNAREs encoded in the yeast genome for their capacity to trigger fusion by partnering with t-SNAREs that mark the Golgi, the vacuole, and the plasma membrane. McNew et al. (2000) found that, to a marked degree, the pattern of membrane flow in the cell is encoded and recapitulated by its isolated SNARE proteins, as predicted by the SNARE hypothesis. The heterodimer of syntaxin Sso1, which is homologous to syntaxin 1A, and Sec9, which is homologous to SNAP25 (600322), is a t-SNARE of the yeast plasma membrane, with Snc2, which is homologous to VAMP2 (185881), as its cognate v-SNARE. Thus, the yeast plasma membrane t-SNARE complex closely resembles its neuronal counterpart (Weber et al., 1998).
In adrenal chromaffin cells, Fisher et al. (2001) expressed a Munc18 (602926) mutant with reduced affinity for syntaxin, which specifically modified the kinetics of single-granule exocytotic release events, consistent with an acceleration of fusion pore expansion. This observation demonstrated that Munc18 functions in a late stage in the intracellular membrane fusion process, where its dissociation from syntaxin determines the kinetics of postfusion events.
The priming step of synaptic vesicle exocytosis is thought to require the formation of the SNARE complex, which comprises the proteins synaptobrevin (185880), SNAP25, and syntaxin. In solution, syntaxin adopts a default, closed configuration that is incompatible with formation of the SNARE complex. Specifically, the amino terminus of syntaxin binds the SNARE motif and occludes interactions with the other SNARE proteins. The N terminus of syntaxin also binds the presynaptic protein UNC13 (UNC13B; 605836). Studies in mouse, Drosophila, and Caenorhabditis elegans suggest that UNC13 functions at a post-docking step of exocytosis, most likely during synaptic vesicle priming. Therefore, UNC13 binding to the N terminus of syntaxin may promote the open configuration of syntaxin. To test this model, Richmond et al. (2001) engineered mutations into C. elegans syntaxin that caused the protein to adopt the open configuration constitutively. Richmond et al. (2001) demonstrated that the open form of syntaxin can bypass the requirement for UNC13 in synaptic vesicle priming. Thus, Richmond et al. (2001) concluded that it is likely that UNC13 primes synaptic vesicles for fusion by promoting the open configuration of syntaxin.
Neuronal exocytosis is triggered by calcium and requires 3 SNARE proteins: synaptobrevin on the synaptic vesicle, and syntaxin and SNAP25 on the plasma membrane. Neuronal SNARE proteins form a parallel 4-helix bundle that is thought to drive the fusion of opposing membranes. Hu et al. (2002) demonstrated that whereas syntaxin and SNAP25 in target membranes are freely available for SNARE complex formation, availability of synaptobrevin on synaptic vesicles is very limited. Calcium at micromolar concentrations triggers SNARE complex formation and fusion between synaptic vesicles and reconstituted target membranes. Although calcium does promote interaction of SNARE proteins between opposing membranes, it does not act by releasing synaptobrevin from synaptic vesicle restriction. Hu et al. (2002) concluded that their data suggests a mechanism in which calcium-triggered membrane apposition enables syntaxin and SNAP25 to engage synaptobrevin, leading to membrane fusion.
SNARE proteins normally face the cytoplasm, within which their helical domains can pair to link membranes for fusion. To ascertain whether SNAREs can fuse cells, Hu et al. (2003) flipped their orientation and engineered cognate cells to express either the v- or t-SNAREs. Hu et al. (2003) found that cells expressing the interacting domains of v- (VAMP2) and t-SNAREs (syntaxin 1A and SNAP25) on the cell surface fused spontaneously, demonstrating that SNAREs are sufficient to fuse biological membranes.
Han et al. (2004) found that mutations of some residues within the transmembrane segment of syntaxin, a plasma membrane protein essential for exocytosis, altered neurotransmitter flux through fusion pores and altered pore conductance. The residues that influenced fusion-pore flux lay along 1 face of an alpha-helical model. Thus, the fusion pore is formed at least in part by a circular arrangement of 5 to 8 syntaxin transmembrane segments in the plasma membrane. The relevant amino acids were I269, V273, G276, I278, and I283. Szule and Coorssen (2004) suggested that the interpretations of Han et al. (2004) did not address sequence conservation or other data in the field, and that the results were equally consistent with the well-established stalk-pore model for membrane fusion. Szule and Coorssen (2004) suggested that unambiguous interpretation of the data presented by Han et al. (2004) is difficult, as the mutations produced only inhibitory effects; enhancement of flux would be more indicative of a direct role for the syntaxin transmembrane domain in pore formation. They cautioned that this may represent a shortcoming of overexpression approaches. Han and Jackson (2004) responded to the comments of Szule and Coorssen (2004), suggesting that overexpression studies may have shortcomings but their experiments in which the wildtype and 11 mutant proteins were overexpressed without altering fusion-pore flux provided an adequate control for an explanation based on expression levels. Han and Jackson (2004) emphasized that despite their results suggesting that the initial fusion pore is composed of protein, a lipidic fusion pore is likely to come into play in the ensuing steps of calcium-triggered exocytosis.
Tucker et al. (2004) investigated the effect of synaptotagmin I (SYT1; 185605) on membrane fusion mediated by neuronal SNARE proteins SNAP25 (600322), syntaxin, and synaptobrevin (see 185880), which were reconstituted into vesicles. In the presence of calcium, the cytoplasmic domain of SYT1 strongly stimulated membrane fusion when synaptobrevin densities were similar to those found in native synaptic vesicles. The calcium dependence of SYT1-stimulated fusion was modulated by changes in lipid composition of the vesicles and by a truncation that mimics cleavage of SNAP25 by botulinum neurotoxin A. Stimulation of fusion was abolished by disrupting the calcium-binding activity, or by severing the tandem C2 domains, of SYT1. Thus, SYT1 and SNAREs are likely to represent the minimal protein complement for calcium-triggered exocytosis.
By use of the large calyx of Held presynaptic terminal from rats, Sakaba et al. (2005) demonstrated that cleavage of different SNARE proteins by clostridial neurotoxins caused distinct kinetic changes in neurotransmitter release. When elevating calcium ion concentration directly at the presynaptic terminal with the use of caged calcium, cleavage of SNAP25 by botulinum toxin A produced a strong reduction in the calcium sensitivity for release, whereas cleavage of syntaxin using botulinum toxin C1 and synaptobrevin using tetanus toxin produced an all or nothing block without changing the kinetics of remaining vesicles. When stimulating release by calcium influx through channels, a difference between botulinum toxin C1 and tetanus toxin emerged, which suggests that cleavage of synaptobrevin modifies the coupling between channels and release-competent vesicles.
Pobbati et al. (2006) found that liposome fusion was dramatically accelerated when a stabilized syntaxin/SNAP25 acceptor complex was used. Thus, SNAREs do have the capacity to execute fusion at a speed required for neuronal secretion, demonstrating that the maintenance of acceptor complexes is a critical step in biologic fusion reactions.
During synaptic vesicle fusion, the SNARE protein syntaxin-1 exhibits 2 conformations that both bind to Munc18-1 (602926): a 'closed' conformation outside the SNARE complex and an 'open' conformation in the SNARE complex. Gerber et al. (2008) 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-1B 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, Gerber et al. (2008) concluded that 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.
Shi et al. (2012) used in vitro membrane fusion and exocytosis assays that paired liposomes containing a t-SNARE complex of rat syntaxin-1A and mouse Snap25 with flat nanodisc proteolipid particles containing the mouse v-SNARE Vamp2. They found that a single Vamp2 protein could mediate efficient SNARE complex formation, vesicle fusion, and lipid mixing between the liposome and nanodisc, but not pore formation or release of liposome cargo. Cargo release was highly sensitive to the number of SNARE complexes formed between the liposome and nanodisc, and maximum efflux required 3 or 4 Vamp2 proteins per nanodisc. Use of chimeric proteins revealed that the membrane-spanning transmembrane domain of VAMP2 mediated efficient release of vesicle contents by stabilizing the nascent fusion pore formed between VAMP2 and the t-SNAREs. Shi et al. (2012) concluded that membrane fusion requires only a single SNARE complex between membranes, but pore formation, widening, and stabilization, as well as efficient cargo efflux, requires several SNARE complexes.
Ma et al. (2013) found that Munc18-1 (602926) could displace SNAP25 (600322) from syntaxin-1 and that fusion of syntaxin-1-Munc18-1 liposomes with synaptobrevin (see 185880) liposomes required Munc13 (UNC13B; 605836), in addition to SNAP25 and synaptotagmin-1-Ca(2+). Moreover, when starting with syntaxin-1-SNAP25 liposomes, NSF (N-ethylmaleimide-sensitive factor)-alpha-SNAP disassembled the syntaxin-1-SNAP25 heterodimers and abrogated fusion, which then required Munc18-1 and Munc13. Ma et al. (2013) proposed that fusion does not proceed through syntaxin-1-SNAP25 heterodimers but starts with the syntaxin-1-Munc18-1 complex; Munc18-1 and Munc13 then orchestrate membrane fusion together with the SNAREs and synaptotagmin-1-Ca(2+) in an NSF- and SNAP-resistant manner.
3-Dimensional Structure
Syntaxin-1A plays a central role in neurotransmitter release through multiple protein-protein interactions. Fernandez et al. (1998) used nuclear magnetic resonance spectroscopy to identify an autonomously folded N-terminal domain in syntaxin-1A and to elucidate its 3-dimensional structure. This 120-residue N-terminal domain is conserved in plasma membrane syntaxins but not in other syntaxins, indicating a specific role in exocytosis. The domain contains 3 long alpha helices that form an up-and-down bundle with a left-handed twist. A striking residue conservation is observed throughout a long groove that is likely to provide a specific surface for protein-protein interactions. A highly acidic region binds to the C(2)A domain of synaptotagmin-1 in a Ca(2+)-dependent interaction that may serve as an electrostatic switch in neurotransmitter release.
Crystal Structure
Stein et al. (2009) reported the x-ray structure of the neuronal SNARE complex, consisting of the SNARE motifs of rat syntaxin-1A, Snap25, and synaptobrevin-2 (VAMP2), with the C-terminal linkers and transmembrane regions of both syntaxin-1A and synaptobrevin-2 at 3.4-angstrom resolution. The structure showed that assembly proceeds beyond the known core SNARE complex, resulting in a continuous helical bundle that is further stabilized by side-chain interactions in the linker region. The results suggested that the final phase of SNARE assembly is directly coupled to membrane merger.
Physical Chemistry
Gao et al. (2012) used optical tweezers to observe in a cell-free reconstitution experiment in real time a long-sought SNARE assembly intermediate in which only the membrane-distal amino-terminal half of the bundle is assembled. Their findings supported the zippering hypothesis, but suggested that zippering proceeds through 3 sequential binary switches, not continuously, in the amino- and carboxyl-terminal halves of the bundle and the linker domain. The half-zippered intermediate was stabilized by externally applied force that mimicked the repulsion between apposed membranes being forced to fuse. This intermediate then rapidly and forcefully zippered, delivering free energy of 36 k(B)T (where k(B) is the Boltzmann constant and T is temperature) to mediate fusion.
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