Vesicle-associated membrane protein VAMP

Sander LE. Frank SP, Bolat S, Blank U, Galli T. Bigalke H. Bischoff SC, Lorentz A Vesicle-associated membrane protein (VAMP)-7 and VAMP-8, but not VAMP-2 or VAMP-3, are required for activation-induced degranulation of mature human mast cells. Eur J Immunol 2008 38 855-863. 

All botulin neurotoxins act in a similar way. They only differ in the amino-acid sequence of some protein parts (Prabakaran et al., 2001). Botulism symptoms are provoked both by oral ingestion and parenteral injection. Botulin toxin is not inactivated by enzymes present in the gastrointestinal tracts. Foodborne BoNT penetrates the intestinal barrier, presumably due to transcytosis. It is then transported to neuromuscular junctions within the bloodstream and blocks the secretion of the neurotransmitter acetylcholine. This results in muscle limpness and palsy caused by selective hydrolysis of soluble A-ethylmalemide-sensitive factor activating (SNARE) proteins which participate in fusion of synaptic vesicles with presynaptic plasma membrane. SNARE proteins include vesicle-associated membrane protein (VAMP), synaptobrevin, syntaxin, and synaptosomal associated protein of 25 kDa (SNAP-25). Their degradation is responsible for neuromuscular palsy due to blocks in acetylcholine transmission from synaptic terminals. In humans, palsy caused by BoNT/A lasts four to six months. 

Stimulus-evoked, calcium-dependent release of acetylcholine (ACh) from the cholinergic synapse normally occurs through the formation of a fusion complex between ACh-containing vesicles and the intracellular leaflet of the nerve terminal membrane (Amon et al., 2001). This synaptic vesicle fusion complex consists of several proteins of the SNARE family, including a 25 kDa synaptosomal associated protein (SNAP-25), vesicle-associated membrane protein (VAMP, or synaptobrevin), and the synaptic membrane protein syntaxin. Other SNARE proteins have been identified as components of membrane transport systems in yeast and mammals but have not been implicated as targets for BoNTs. Meanwhile, type A and E neurotoxins cleave SNAP-25 while types B, D, F, and G act on VAMP and type C1 toxin cleaves both syntaxin and SNAP-25. Neurotoxin-mediated cleavage of any of these substrates disrupts the processes involved in the exocytotic release of ACh and leads to flaccid paralysis of the affected skeletal muscles. 

It is now well established that in vivo efficient membrane fusion requires the interaction of small cytoplasmically exposed membrane proteins called soluble N-ethylmaleimide sensitive factor (NSF) attachment receptors (SNAREs) (Sollner et al., 1993). For synaptic vesicle exocytosis, the relevant SNAREs are synaptobrevin/ vesicle-associated membrane protein (VAMP) 1 and 2, syntaxin 1, and synaptosome-associ-ated protein of 25,000 daltons (SNAP-25). Synaptobrevins/ VAMPs are localized primarily on synaptic vesicles, while syntaxin and SNAP-25 are localized primarily on the plasma membrane. Fusion is driven by the progressive zippering of vesicle and plasma membrane SNAREs forming a four-helix bundle (Sutton et al., 1998). Although many other proteins appear to play critical roles in synaptic vesicle exocytosis, it seems likely that SNAREs are the minimal machinery required for fusion (Weber et al., 1998). Once assembled, SNARE complexes are disassembled by NSF, which functions in conjunction with SNAP proteins. 

Braun JEA, Fritz BA, Wong SME, Lowe AW (1994) Identification of a vesicle-associated membrane protein (VAMP-like) membrane protein in zymogen granules of the rat exocryne pancreas. In J. Biol. Chem. 269 5328-35 Brennwald P, Kearns B, Champion K, Keranen S, Bankaitis V, Novick P (1994) Sec9 is a SNAP-25-like component of a yeast SNARE complex that may be the effector of Sec4 function in excytosis. In Cell 79 245—58 Broadie K, Prokop A, Bellen HJ, O Kane C, Schulze KL, Sweeney ST (1995) Syntaxin and synaptobrevin function downstream of vesicle docking in Drosophila. In Neuron 15 663-73... 

Gaisano HY, Sheu L, Foskett JK et al. (1994) Tetanus toxin light chain cleaves a vesicle-associated membrane protein (VAMP) isiform 2 in rat pancreatic zymogen granules and inhibits enzyme secretion. J. Biol. Chem. 269 17062-6. 

The vesicle-associated membrane proteins (VAMP or synaptobrevin) occur in three isoforms and are proteins that are anchored to the cytoplasmic portion of synaptic membrane vesicles and secretory granules. VAMP2 and 3 are present in pancreatic beta cells, but the roles of this family of proteins have not been widely studied as markers of NE tumors. In contrast to synaptophysin and other synaptic vesicle proteins, SNAP-25 (synaptosomal protein of 25 KD) and syntaxin are present in the plasma membranes. At present, there are few studies on the application of these markers in diagnostic pathology... 

Inside the cytosol, the L chain of the toxin acts as a proteolytic enzyme whose activity is dependent on the Zn ions present in the molecules. It hydrolyses protein components of the exocytosis apparatus to block the release of the transmitter. It is known that botulinum toxins B, D and G can cleave vesicle associated membrane protein (VAMP), a protein present in the membranes of acetylcholine vesicles. Botulinum toxins A, C and E, on the other hand, act on proteins of the pres maptic plasma membrane. A and E cleave synaptosomal associated protein 25 (SNAP 2s) and C cleaves... 

In the sea urchin embryo, cells moving from the mesoderm to the ectoderm secrete SNARE proteins (soluble NSF attachment protein receptor, N-ethylmaleimide-sensitive factor), syntaxin, vesicle-associated membrane protein (VAMP) and Rab3 in order to break ceU adhesions (intercellular matrix) for their locomotion [522]. SNARE, syntaxins, VAMP and SNAP (soluble NSF-attachment protein), physiological directors of cellular traffic, are predecessors of the metalloproteinases (MMPs), that cancer cells secrete for the clearance of intercellular passageways for their locomotion (invasiveness). 

Amperometry at single PC 12 cells has also been used in conjunction with a genetic cell transfection protocol to examine the effects of toxin expression on basal and evoked exocytosis. PC 12 cells have been transfected with the specific endoprotease Botulinum neurotoxin Cl light chain (BoNT/Cl), which cleaves the proteins syntaxin and SNAP-25 [5], The molecular dissection of the mechanisms underlying exocytosis has been motivated by the SNARE hypothesis, which postulates that exocytosis requires the assembly of the plasma membrane proteins syntaxin 1, SNAP-25, and the vesicle associated membrane protein (VAMP) into a complex [5], This SNARE complex then acts as a receptor for cytosolic components of the proposed fusion machinery. Direct evidence for the role of the SNARE proteins in neurotransmission comes from molecular genetic studies in which syntaxin and VAMP have been shown to be required for neurotransmission in Drosophila [47 9] and Caenorhabditis elegans [50,51]. To assess the effects of the disruption of SNARE proteins on exocytosis in PC 12 cells, amperometry has been used in conjunction with a genetic cell transfection assay to establish a... 

Fig. 6.4 Model for protein-mediated membrane fusion and exocytosis. a The release of acetylcholine from the vesicles is mediated by a series of proteins collectively called SNARE proteins. Synaptotagmin is the neuronal Ca " receptor detecting C entry. Synaptobrevin (i.e. vesicle-associated membrane protein, VAMP) is a filament-like protein on the vesicle, b During depolarisation and calcium entry, synaptobrevin on the vesicle unfolds and forms a ternary complex with syntaxin/SNAP-25. This process is facilitated by phosphorylation of synapsin, also present on the vesicle membrane, c Assembly of the ternary complex forces the vesicle in close apposition to the nerve membrane at the active zone with release of its contents, acetylcholine. The fusion is disassembled, and the vesicle is recycled. (From Martyn 2005, p 864 copyright Elsevier)...

Schematic illustration of a generalized cholinergic junction (not to scale). Choline is transported into the presynaptic nerve terminal by a sodium-dependent choline transporter (CHT). This transporter can be inhibited by hemicholinium drugs. In the cytoplasm, acetylcholine is synthesized from choline and acetyl -A (AcCoA) by the enzyme choline acetyltransferase (ChAT). Acetylcholine is then transported into the storage vesicle by a second carrier, the vesicle-associated transporter (VAT), which can be inhibited by vesamicol. Peptides (P), adenosine triphosphate (ATP), and proteoglycan are also stored in the vesicle. Release of transmitter occurs when voltage-sensitive calcium channels in the terminal membrane are opened, allowing an influx of calcium. The resulting increase in intracellular calcium causes fusion of vesicles with the surface membrane and exocytotic expulsion of acetylcholine and cotransmitters into the junctional cleft (see text). This step can he blocked by botulinum toxin. Acetylcholine s action is terminated by metabolism by the enzyme acetylcholinesterase. Receptors on the presynaptic nerve ending modulate transmitter release. SNAPs, synaptosome-associated proteins VAMPs, vesicle-associated membrane proteins.

Release can be blocked by drugs such as guanethidine and bretylium. After release, norepinephrine diffuses out of the cleft or is transported into the cytoplasm of the terminal (uptake 1 [1], blocked by cocaine, tricyclic antidepressants) or into the postjunctional cell (uptake 2 [2]). Regulatory receptors are present on the presynaptic terminal. (SNAPs, synaptosome-associated proteins VAMPs, vesicle-associated membrane proteins.)... 

In neurons, the SNARE complex consists of three main proteins the v-SNARE synaptobrevin or VAMP (vesicle-associated membrane protein), and two t-SNAREs, syntaxin and SNAP-25 (synaptosomal associated protein of 25 kD). Synaptobrevins traverse the synaptic vesicle membrane in an asymmetric manner a few amino acids are found inside the vesicle, but most of the molecule lies outside the vesicle, within the cytoplasm. Synaptobrevin makes contact with another protein anchored to the plasma membrane of the presynaptic neuron, syntaxin, which is associated with SNAP-25. Via these interactions, the SNARE proteins play a role in the docking and fusion of synaptic vesicles to the active zone. 

TDO TE TEC Teff TE TG TGE Thl Th2 TIA d yptophan 2,3-dioxygenase echo time thymic epithelial cell T effector cells d anscripdon factor digeminal ganglia d ansforming growth factor T helper type 1 cell T helper type 2 cell d ansient ischemic attack VI VAMP VCAM primary visual cortex vesicle-associated membrane protein vascular cell adhesion molecule... 

VAMP vesicle-associated membrane protein (= synaptobre-vin)... 

Protease-free preparations of TeTx and BoNT/B, D, F and G cleave a membrane protein of small synaptic vesicles (SSV) called VAMP or synaptobrevin (Schiavo ef ai, 1992 a, 1993 a,c, Yamasaki et ai, 1994 a, b). Conversely, BoNT/A, C and E act on proteins associated with the presynaptic membrane BoNT/A and E cut SNAP-25, while serotype C cleaves syntaxin, in addition to SNAP-25 (Blasi efai, 1993 a, b ... 

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