Synaptosomal-associated protein SNAP

At the level of SNARE complex formation and dissolution, synaptosome-associated protein (SNAP-25) (Hepp et al. 2002 Risinger and Bennett 1999), a-SNAP (Hirling and Scheller 1996) and syntaphilin (Hirling and Scheller 1996) are phosphorylated by PKA. These are therefore candidate regulators of exocytosis by GPCR/AC signaling. 

Fatemi SH, Earle JA, Stary JM, Lee S, Sedgewick J. 2001. Altered levels of the synaptosomal associated protein SNAP-25 in hippocampus of subjects with mood disorders and schizophrenia. Neuroreport 12 3257-3262. 

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. 

Some intracellular signal transduction molecules are reduced in schizophrenia. The release of neurotransmitters is regulated by a family of proteins that coordinate vesicular trafficking (see Ch. 9). Of these, the expression of complexin I and II appears to be decreased in prefrontal cortex and subfields of the hippocampal formation, and the ratio of complexin I to complexin II is elevated in the hippocampus [35], SNAP-25 (Synaptosomal Associated Protein, kDa 25) has inconsistently been found to be down-regulated in both these regions. Synapsin expression is also reduced, but more robust decrements have been observed in bipolar disorder (Ch. 55). 

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. 

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.)... 

Criado M, Gil A, Viniegra S, Gutierrez LM (1999) A single amino acid near the C terminus of the synaptosome associated protein of 25 kKa (SNAP-25) is essential for exocytosis in chromaffin cells. Proc Natl Acad Sci USA 96 7256-61... 

Raciborska DA, Charlton MP (1999) Retention of cleaved synaptosome-associated protein of 25 kDa (SNAP-25) in neuromuscular junctions a new hypothesis to explain persistence of botulinum A poisoning. Can J Physiol Pharmacol 77 679-88 Raciborska DA, Trimble WS, Charlton MP (1998) Presynaptic protein interactions in vivo evidence from botulinum A, C, D and E action at frog neuromuscular junction. Eur J Neurosci 10 2617-28... 

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. 

Synaptosomal proteins are involved in synaptic transmission, which is deranged in AD. The machinery consists of synaptosomal proteins, including soluble N-ethylmaleimide-sensitive factor attachment proteins (SNAPs) present as three isoforms, a, (3 and y, synaptosomal-associated protein 25 (SNAP 25), synapto-tagmin, and vesicular proteins. Proteomics studies showed reduced expression... 

SNAP-25 (synaptosomal-associated protein of 25kDa) is a 206 residue protein that lacks a classical transmembrane segment, but is bound to the cytosolic surface of the neuronal plasmalemma via pal-mitoylation of four cysteine residues located at the center of the molecule (Fig. 3) (Bennett and Scheller, 1994 Sudhof, 1995). SNAP-25 is required for axonal growth during development, and for nerve terminal plasticity in the mature nervous system (Osen-Sand et al., 1993). The tissue distribution of SNAP-25 is less well characterized than that of VAMP however, its presence in pancreatic cells (Jacobson et al., 1994 Sadoul et al., 1995) may indicate that it is also expressed outside the nervous system. A SNAP-25 related protein required for post-Golgi transport has been cloned from yeast (Brenwald efal., 1994). 

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... 

The LC then targets and cleaves one or more of three soluble NSF attachment protein receptor (SNARE) proteins with exquisite specificity synaptosomal-associated protein 25 (SNAP-25 BoNT/A, /C, /E) vesicle associated membrane protein 1-3 (VAMPl-3 BoNT-/B, /D, /F, /G) or syntaxin (BoNT/C). The SNARE proteins are essential components of the synaptic exocytosis mechanism, and their cleavage prevents functional assembly of the ternary complex, thereby blocking neurotransmitter release. The combination of efficient neuronal targeting and presynaptic activation renders BoNTs the most potent substances known, with estimated human lethal doses as low as 0.1-1 ng/kg. 

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. 

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... 

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