Botulinum neurotoxin release

Botulinum neurotoxins (A-G), tetanus toxin Synaptic peptides a) Synapto-brevin b) Syntaxin c) SNAP25 Zinc dependent endoprotease Cleavage of synaptic peptides Inhibition of transmitter release (tetanus, botulism)... 

G. Schiavo, F. Benfenati, B. Poulain, O. Rossetto, P. Polverino de Laureto, B. R. Das-Gupta, C. Montecucco, Tetanus and Botulinum-B Neurotoxins Block Neurotransmitter Release by Proteolytic Cleavage of Synaptobrevin , Nature 1992a, 359, 832-835 G. Schiavo, O. Rossetto, A. Santucci, B. R. DasGupta, C. Montecucco, Botulinum Neurotoxins are Zinc Proteins , J. Biol. Chem. 1992b, 267, 23479-23483. 

Poulain, B., Mochida, S., Weller, U., Hogy, B., Habermann, E., Wadsworth, J. D., Shone, C.C., Dolly, J.O. and Tauc, L., Heterologous combinations of heavy and light chains from botulinum neurotoxin A and tetanus toxin inhibit neurotransmitter release in Aplysia, J. Biol. Chem., 266, 9580-9585, 1991. 

Ashton AC, Dolly JO (1988) Characterization of the inhibitory action of botulinum neurotoxin type A on the release of several transmitters from rat cerebrocortical synaptosomes. J Neurochem... 

Foran, P., Lawrence, G. W., Shone, C. C., Foster, K. A., and Dolly, J. O. (1996). Botulinum neurotoxin C1 cleaves both syntaxin and SNAR-25 in intact and permeabilized chromaffin cells Correlation with its blockade of catecholamine release. Biochemistry 25, 2630—2636. 

The first report in this area compared the actions of botulinum neurotoxin and botulinum binary toxin on transmission in the phrenic nerve-hemidiaphragm preparation (Simpson, 1982). There was the expected finding that neurotoxin blocked transmission by blocking acetylcholine release from nerve terminals, but the binary toxin had no effect. Apart from showing that C2 toxin did not block exocytosis, this study showed that the toxin did not act on the diaphragm to block muscle twitch. This observation is in keeping with the fact that the predominant form of actin in striated muscle is not that which is ADP-ribosylated by C2 toxin. 

Blasi J, Chapman ER, Yamasaki S, BinzT, Niemann H, Jahn R (1993 a) Botulinum neurotoxin C blocks neurotransmitter release by means of cleaving HPC-1/syntaxin. In EMBOJ. 12 4821-8... 

Boyd RS, Duggan MJ, Shone CC, Foster KA (1995 b) The effect of botulinum neurotoxins on the release of insulin from insulinoma cell lines HIT-15 and RINm5F. In J. Biol. Chem. 270 18216-8... 

Matsuda M, Lei DL, Sugimoto N, Ozutsumi K, OkabeT (1989) Isolation, purification and characterization of fragment B, the NH2-terminal half of the heavy chain of tetanus toxin. In Infect. Immun. 57 3588-93 Matthews BW (1988) Structural basis of the action of thermolysin and related zinc peptidases.In Acc. Chem. Res. 21 333-40 Mclnnes C, Dolly JO (1990) Ca -dependent noradrenaline release from perme-abilised PC 12 cells is blocked by botulinum neurotoxin A or its light chain. In FEBS Lett. 261 323-6... 

Schiavo G, Poulain B, Rossetto O, Benfenati F, Tauc L, Montecucco C (1992 b) Tetanus toxin is a zinc protein and its inhibition of neurotrasmitter release and protease activity depend on zinc. In EMBOJ. 11 3577-83 Schiavo G, Rossetto O, Santucci A, DasGupta BR, Montecucco C (1992 c) Botulinum neurotoxins are zinc proteins. In J. Biol. Chem. 267 23479-83 Schiavo G, Rossetto O, Catsicas S, Polverino de Laureto P, DasGupta BR, Benfenati F, Montecucco C (1993 a) Identification of the nerve-terminal targets of botulinum neurotoxins serotypes A, D and E. In J. Biol. Chem. 268 23784-7 Schiavo G, Santucci A, DasGupta BR, Metha PP, Jontes J, Benfenati F, Wilson MC, Montecucco C (1993 b) Botulinum neurotoxins serotypes A and E cleave SNAP-25 at distinct COOH-terminal peptide bonds. In FEBS Lett. 335 99-103 Schiavo G, Shone CC, Rossetto O, Alexandre FCG, Montecucco C (1993 c) Botulinum neurotoxin serotype F is a zinc endopeptidase specific for VAMP/synOp-tobrevin. In J. Biol. Chem. 268 11516-9... 

Tetanus toxin poisoning produces tetanus, i.e. muscle contractions resulting in spastic paralysis. In contrast, Botulinum neurotoxins cause botulism, which is characterized by flaccid paralysis. This difference reflects differences in the anatomical level of action of these toxins. TeTx acts primarily on the CNS where it blocks exocytosis from inhibitory glycinergic synapses in the spinal cord. Loss of inhibitory control results in motoneuron firing. BoNTs act primarily in the periphery where they inhibit acetylcholine release at the neuromuscular junctions. 

Mochida S, Saisu H, Kobayashi H etal. (1995) Impairment of synatxin by botulinum neurotoxin Ci or antibodies inhibits acetylcholine release but not Ca channel activity. Neuroscience. 65 905—15. 

Poulain B, Mochida S, Wadsworth JD et al. (1990) Inhibition of neurotransmitter release by botulinum neurotoxins and tetanus toxin at Aplysia synapses role of the constituent chains. J. Physiol. (Paris) 84 247-61. 

The botulinum neurotoxins (BoNTs) comprise a family of seven distinct neurotoxic proteins (A-G) produced by immunologically discrete strains of the anaerobic bacterium Clostridium botulinum and in rare cases by Clostridium baratii and Clostridium butyricum (Habermann and Dreyer, 1986 Harvey et ah, 2002 Simpson, 2004). These toxins act on peripheral cholinergic synapses to inhibit spontaneous and impulse-dependent release of acetylcholine (ACh) (Brooks, 1956 Kao et al., 1976). Intoxication by BoNT results in muscle weakness, which can be fatal when the diaphragm and intercostal muscles become sufficiently compromised to impair ventilation (Dickson and Shevky, 1923). The BoNTs are the most potent substances in nature, and exposure to as httle as 1-3 ng/kg may be sufficient to cause human lethahty (GUI, 1982 Middlebrook and Franz, 1997 Amon et al., 2001). 

Blasi, J., Chapman, E.R., Yamasaki, S., Binz, T., Niemann, H., and Jahn, R. 1993b. Botulinum neurotoxin Cl blocks neurotransmitter release by means of cleaving HPC-l/syntaxin. EMBO J. 12 4821 4X28. 

Ferrer-Montiel, A.V., Gutierrez, L.M., Apland, J.P., Canaves, J.M., Gd, A., Viniegra, S., Biser, J.A., Adler, M., and Montal, M., The 26-mer peptide released from SNAP-25 cleavage by botulinum neurotoxin E inhibits vesicle docking, FEES Lett., 435, 84, 1998. 

Type Cl and D botulinum neurotoxins as ADP-ribosyl transferases. As shown by Knight et al. (3), type D botulinum neurotoxin was able to inhibit exocytosis in cultured chromaffin cells. Fig. 1 represents the results of our experiments showing the time course of this inhibition. When cultured bovine adrenal chromaffin cells were incubated with type D botulinum neurotoxin, inhibition of acetylcholine-evoked catecholamine release appeared. This inhibition, however, did not occur instantaneously but appeared and increased with days of incubation, suggesting involvement... 

Fig. 1. Geft) Inhibition of acetylcholine-induced catecholamine release by botulinum toxin in cultured adtenal chromaffin cells. Bovine adrenal chromaffin cells were isolated and cultured at a density of 5 x lOVml. After two days of culture, the cells were washed and suspended in a fresh medium containing 20 pg/ml of type D botulinum neurotoxin. After incubation for indicated days, the cells were washed and stimulated 500 with pM acetylcholine. Catecholamines in the media and cells were extracted separately and quantified electrochemically. Catecholamine release is expressed as % of the total amount , acetylcholine-evoked release O, basal release.

So far eight different botulinum toxins (A, B, Cl, C2, D, E, F, G) have been described which are produced by various strains of Clostridium botulinum (1). Whereas seven of the botulinum toxins are neurotoxins and block the release at the cholinergic synapses, botulinum C2 toxin is not neurotoxic and acts on various non-neuronal tissues (1-3). It has been shown that component I of the binary botulinum C2 toxin possesses ADP-ribosyltransferase activity (4) on the eukaryotic substrate non-muscle actin (5). Here we describe another ADP-ribosyltransferase which is produced by certain strains of Clostridium botulinum type C. In order to distinguish the novel ADP-ribosyltransferase from botulinum neurotoxin Cl and botulinum C2 toxin we termed this enzyme C3. 

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