Neuromuscular junction proteins

In myasthenia gravis synaptic transmission at the neuromuscular junction is impaired most cases have an autoimmune basis and some 85% of patients have a raised titre of autoantibodies to the muscle acetylcholine receptor. The condition is probably heterogeneous, however, as about 15% do not have receptor antibodies, or have antibodies to another neuromuscular junction protein (muscle specific kinase) and rarely it occurs with penicillamine used for rheumatoid arthritis. 

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. 

The amount of acetylcholine present in the synapse and the amount of time that it remains there are critical. For example, the venom of the black widow spider is highly neurotoxic. It contains a protein known as a-latrotoxin that elicits the release of massive amounts of acetylcholine at the neuromuscular junction. Too much of a good thing can be a serious problem. 

Acetylcholine (ACh) is an example of an endogenous neurotransmitter that binds to more than one receptor type, the nicotinic acetylcholine receptor (nAChR) which preferentially binds nicotine and the muscarinic receptor which binds muscarine, a mushroom alkaloid. The latter is a G protein-coupled receptor while the nACh receptor is an excitatory ligand-gated ion channel that transports Na-i- ions. Nicotinic cholinergic receptors are found in the CNS, autonomic ganglia, and at the neuromuscular junction of skeletal muscles. They are a possible target for anaesthetics. 

Acetylcholine released by an excited neuron diffuses a few micrometers across the synaptic cleft or neuromuscular junction to the postsynaptic neuron or myocyte, where it interacts with the acetylcholine receptor and triggers electrical excitation (depolarization) of the receiving cell. The acetylcholine receptor is an allosteric protein with two high-affinity binding sites for acetylcholine, about 3.0 nm from the ion gate, on the two a... 

What does a neurotransmitter do at the postsynap-tic membrane In the case of acetylcholine in neuromuscular junctions the principal action appears to be one of opening sodium channels and thereby depolarizing the postsynaptic membrane. If enough nerve impulses arrive, an action potential will be initiated in the postsynaptic neuron. In other cases, the first response may be activation of a protein kinase either directly or by opening a channel for Ca2+, which indirectly regulates protein kinases and phosphatases.592 Thus, a complex cascade may be activated. See also Fig. 30-19. 

Dystrobrevins are a small family of dystrophin-related proteins encoded by two genes, one of which—Q-dystrobrevin—encodes at least three proteins all expressed in cardiac and skeletal muscle. o-Dystrobrevin is a key component of the DPC, and its loss results in neuromuscular junction defects and muscular dystrophy. It is a cytoplasmic protein indirectly linked with the transmembrane sarcoglycan components via dystrophin and other components of the DPC (Fig. 4) (Blake and Martin-Rendon, 2002). Recently, two novel type IV IF proteins—syncoilin (Newey et al.,... 

Nguyen, T. M., Ellis, J. M., Love, D. R., Davies, K. E., Gatter, K. C., Dickson, G., and Morris, G. E. (1991). Localization of the DMDL gene-encoded dystrophin-related protein using a panel of nineteen monoclonal antibodies Presence at neuromuscular junctions, in the sarcolemma of dystrophic skeletal muscle, in vascular and other smooth muscles, and in proliferating brain cell lines. J. Cell Biol. 115, 1695-1700. 

Mechanism of action. The cellular actions of bot-ulinum toxin at the neuromuscular junction have recently been clarified.84 This toxin is attracted to glycoproteins located on the surface of the presynaptic terminal at the skeletal neuromuscular junction.33 Once attached to the membrane, the toxin enters the presynaptic terminal and inhibits proteins that are needed for acetylcholine release (Figure 13-4).84 Normally, certain proteins help fuse presynaptic vesicles with the inner surface of the presynaptic terminal, thereby allowing the vesicles to release acetylcholine via exocytosis. Botulinum toxin cleaves and destroys these fusion proteins, thus making it impossible for the neuron to release acetylcholine into the synaptic cleft.32,84 Local injection of botulinum toxin into specific muscles will therefore decrease muscle excitation by disrupting synaptic transmission at the neuromuscular junction. The affected muscle will invariably undergo some degree of paresis and subsequent... 

FIGURE 13-4 Mechanism of action of botulinum toxin at the skeletal neuromuscular junction. At a normal synapse (shown on left], fusion proteins connect acetylcholine (ACh] vesicles with the presynaptic membrane, and ACh is released via exocytosis. Botulinum toxin (represented by BTX on the right] binds to the presynaptic terminal, and enters the terminal where it destroys the fusion proteins so that ACh cannot be released. See text for details. 

Weatherbee, S. D., Anderson, K. V. and Niswander, L. A. (2006) LDL-receptor-related protein 4 is crucial for formation of the neuromuscular junction. Development 133,4993-5000. 

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... 

Rossetto O, Seveso M, Caccin P, Schiavo G, Montecucco C (2001b) Tetanus and botulinum neurotoxins turning bad guys into good by research. Toxicon 39 27—41 Rossetto O, Morbiato L, Rossetto et al. 2006 Caccin P, Rigoni M, Montecucco C (2006) Presynaptic enzymatic neurotoxins. J Neurochem 97 1534—4 5 Roux S, Colasante C, Saint Clomcnt C, Barbier J, Curie T et al. (2005) Internalization of a GFP-tetanus toxin C-terminal fragment fusion protein at mature mouse neuromuscular junctions. Mol Cell Neurosci 30 572-82... 

Frontali N, Ceccarelli B, Gorio A et al (1976) Purification from black widow spider venom of a protein factor causing the depletion of synaptic vesicles at neuromuscular junctions. J Cell Biol... 

Liu J, Wan Q, Lin X et al (2005) a-Latrotoxin modulates the secretory machinery via receptor-mediated activation of protein kinase C. Traffic 6 756-65 Long SB, Campbell EB, Mackinnon R (2005) Crystal structure of a mammalian voltage-dependent Shaker family K+ channel. Science 309 897-903 Longenecker HE, Hurlbut WP, Mauro A et al (1970) Effects of black widow spider venom on the frog neuromuscular junction. Effects on end-plate potential, miniature end-plate potential and nerve terminal spike. Nature 225 701-3... 

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