Botulinum neurotoxin C1 blocks neurotransmitter release by means of cleaving HPC-1/syntaxin.
01 Dec 1993-The EMBO Journal (European Molecular Biology Organization)-Vol. 12, Iss: 12, pp 4821-4828
TL;DR: It is concluded that HPC‐1/syntaxin, a membrane protein present in axonal and synaptic membranes, is involved in exocytotic membrane fusion.
Abstract: The anaerobic bacterium Clostridium botulinum produces several related neurotoxins that block exocytosis of synaptic vesicles in nerve terminals and that are responsible for the clinical manifestations of botulism. Recently, it was reported that botulinum neurotoxin type B as well as tetanus toxin act as zinc-dependent proteases that specifically cleave synaptobrevin, a membrane protein of synaptic vesicles (Link et al., Biochem. Biophys. Res. Commun., 189, 1017-1023; Schiavo et al., Nature, 359, 832-835). Here we report that inhibition of neurotransmitter release by botulinum neurotoxin type C1 was associated with the proteolysis of HPC-1 (= syntaxin), a membrane protein present in axonal and synaptic membranes. Breakdown of HPC-1/syntaxin was selective since no other protein degradation was detectable. In vitro studies showed that the breakdown was due to a direct interaction between HPC-1/syntaxin and the toxin light chain which acts as a metallo-endoprotease. Toxin-induced cleavage resulted in the generation of a soluble fragment of HPC-1/syntaxin that is 2-4 kDa smaller than the native protein. When HPC-1/syntaxin was translated in vitro, cleavage occurred only when translation was performed in the presence of microsomes, although a full-length product was obtained in the absence of membranes. However, susceptibility to toxin cleavage was restored when the product of membrane-free translation was subsequently incorporated into artificial proteoliposomes. In addition, a translated form of HPC-1/syntaxin, which lacked the putative transmembrane domain at the C-terminus, was soluble and resistant to toxin action. We conclude that HPC-1/syntaxin is involved in exocytotic membrane fusion.(ABSTRACT TRUNCATED AT 250 WORDS)
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TL;DR: A fascinating picture of these robust nanomachines is emerging, which seems to be conserved and adaptable for fusion reactions as diverse as those involved in cell growth, membrane repair, cytokinesis and synaptic transmission.
Abstract: Since the discovery of SNARE proteins in the late 1980s, SNAREs have been recognized as key components of protein complexes that drive membrane fusion. Despite considerable sequence divergence among SNARE proteins, their mechanism seems to be conserved and is adaptable for fusion reactions as diverse as those involved in cell growth, membrane repair, cytokinesis and synaptic transmission. A fascinating picture of these robust nanomachines is emerging.
2,424 citations
TL;DR: It is reported that in the absence of SNAP and NSF, these three SNAREs form a stable complex that can also bind synaptotagmin, suggesting that synapttagmin operates as a "clamp" to prevent fusion from proceeding in the absent of a signal.
1,873 citations
TL;DR: The two universally required components of the intracellular membrane fusion machinery, SNARE and SM (Sec1/Munc18-like) proteins, play complementary roles in fusion and are spectacularly apparent in the exquisite speed and precision of synaptic exocytosis.
Abstract: The two universally required components of the intracellular membrane fusion machinery, SNARE and SM (Sec1/Munc18-like) proteins, play complementary roles in fusion. Vesicular and target membrane-localized SNARE proteins zipper up into an alpha-helical bundle that pulls the two membranes tightly together to exert the force required for fusion. SM proteins, shaped like clasps, bind to trans-SNARE complexes to direct their fusogenic action. Individual fusion reactions are executed by distinct combinations of SNARE and SM proteins to ensure specificity, and are controlled by regulators that embed the SM-SNARE fusion machinery into a physiological context. This regulation is spectacularly apparent in the exquisite speed and precision of synaptic exocytosis, where synaptotagmin (the calcium-ion sensor for fusion) cooperates with complexin (the clamp activator) to control the precisely timed release of neurotransmitters that initiates synaptic transmission and underlies brain function.
1,862 citations
TL;DR: Fusion of intracellular membranes in eukaryotic cells involves several protein families including SNAREs, Rab proteins, and Sec1/Munc-18 related proteins (SM-proteins).
Abstract: ▪ Abstract Membrane fusion involves the merger of two phospholipid bilayers in an aqueous environment. In artificial lipid bilayers, fusion proceeds by means of defined transition states, including hourglass-shaped intermediates in which the proximal leaflets of the fusing membranes are merged whereas the distal leaflets are separate (fusion stalk), followed by the reversible opening of small aqueous fusion pores. Fusion of biological membranes requires the action of specific fusion proteins. Best understood are the viral fusion proteins that are responsible for merging the viral with the host cell membrane during infection. These proteins undergo spontaneous and dramatic conformational changes upon activation. In the case of the paradigmatic fusion proteins of the influenza virus and of the human immunodeficiency virus, an amphiphilic fusion peptide is inserted into the target membrane. The protein then reorients itself, thus forcing the fusing membranes together and inducing lipid mixing. Fusion of intr...
1,214 citations
TL;DR: The mechanism of action of three groups of presynaptic neurotoxins that interfere directly with the process of neurotransmitter release is reviewed, whereas presynapses acting on ion channels are not dealt with here.
Abstract: Nerve terminals are specific sites of action of a very large number of toxins produced by many different organisms. The mechanism of action of three groups of presynaptic neurotoxins that interfere directly with the process of neurotransmitter release is reviewed, whereas presynaptic neurotoxins acting on ion channels are not dealt with here. These neurotoxins can be grouped in three large families: 1) the clostridial neurotoxins that act inside nerves and block neurotransmitter release via their metalloproteolytic activity directed specifically on SNARE proteins; 2) the snake presynaptic neurotoxins with phospholipase A(2) activity, whose site of action is still undefined and which induce the release of acethylcholine followed by impairment of synaptic functions; and 3) the excitatory latrotoxin-like neurotoxins that induce a massive release of neurotransmitter at peripheral and central synapses. Their modes of binding, sites of action, and biochemical activities are discussed in relation to the symptoms of the diseases they cause. The use of these toxins in cell biology and neuroscience is considered as well as the therapeutic utilization of the botulinum neurotoxins in human diseases characterized by hyperfunction of cholinergic terminals.
1,196 citations
Cites background from "Botulinum neurotoxin C1 blocks neur..."
...TeNT and BoNT/B, /D, /F and /G cleave vesicle-associated membrane protein (VAMP)/synaptobrevin, but each at different sites (531, 535, 538, 539, 543, 666, 667); BoNT/A and /E cleave 25-kDa synaptosomal-associated protein (SNAP-25) at two different sites and BoNT/C cleaves both syntaxin and SNAP-25 (56, 67, 68, 189, 450, 539, 541, 542, 655)....
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...Several experimental data indicate that there is a direct correlation between neurotoxin-induced proteolysis of VAMP or SNAP-25 or syntaxin and inhibition of neurotransmitter release The SNARE proteins are cleaved in synaptosomes and cells intoxicated with TeNT or BoNT with a corresponding inhibition of exocytosis (3, 67, 68, 96, 189, 265, 335, 348, 375, 450, 454, 492, 500, 520, 539, 655)....
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...5) BoNT/C only cleaves membrane-bound SNAP-25 and syntaxin and is ineffective on the isolated molecules (68, 542); also, other neurotoxins are more effective on the membrane-bound substrate than on the recombinant soluble molecule (450, 467, 655)....
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...In the case of nerve cells or synaptosomes, the simple incubation of cells with CNT is sufficient to cause inhibition of neurotransmitter release and SNARE cleavage (9, 19, 67, 68, 250, 277, 339, 375, 378, 380, 382, 433, 450, 492, 525, 559, 657, 665, 666)....
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