PHIDIAS: Pathogen-Host Interaction Data Integration and Analysis System

Phinet Name: Clostridium botulinum
Phinet Information
Pathogen Name: Clostridium botulinum
Pathogen NIAID Category: NIAID Category A
Bio-objects
Bio-object 1: Absorptive cells
Type: Eukaryotic cell or cell component Location: Cytoplasm Function: Transporter Description: In the case of absorptive cells, the transport process typically does nothing more than ferry molecules across structural barriers. However, the major route utilized by toxin is transcytosis across absorptive cells. Toxin can bind to the apical surface of cells, undergo transcytosis, and be released on the basolateral side. Toxin that undergoes transport is released in a form that is structurally and functionally indistinguishable from that originally added to cells.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 2: Botulinum protoxin
Type: Protein or gene Location: Cytoplasm Function: Enzyme Description: Botulinum protoxin is synthesized as a single-chain polypeptide with a molecular mass of 150 kDa.(<a href="#reference5263">Park et al., 2003</a>)
Bio-object 3: Botulinum toxin. Food infection/Primary intoxication
Type: Protein or gene Location: Extracellular Function: Enzyme Description: The contaminated food serves as a culture medium for bacteria. The microorganisms mature and undergo autolysis and then release toxin. In primary intoxication, the patient ingests a food that is already tainted with botulinum toxin.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 4: Botulinum toxin. Primary human infection/Secondary intoxication
Type: Protein or gene Location: Extracellular Function: Enzyme Description: In primary infection, the patient ingests food that harbors botulinum spores that may germinate and colonize in the gut. In the process of growth, division, and autolysis, microorganisms release toxin in situ, and this leads to secondary intoxication. Consumption of botulinum spores is probably somewhat common, although the incidence of secondary intoxication is low. This can be attributed to several factors, with the most important being the inability of C. botulinum to compete for a niche in healthy guts that have a normal flora. This explains the fact that primary infection with secondary intoxication occurs almost exclusively in young infants (e.g., the normal flora has not developed) or in older persons who have been treated with certain antibiotics (e.g., the normal flora has been reduced or eliminated).(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 5: Clostridium botulinum
Type: Microorganism or its component Location: Extracellular Function: Toxicity Description: Botulinum toxin is one of the most potent neurotoxins known, which is synthesized by the bacteria Clostridium botulinum. The great majority of cases of botulism are contracted orally, either by ingesting food containing botulinum toxin or by ingesting food contaminated with Clostridium botulinum bacteria or spores that reproduce in the gut and release the toxin. The various etiologies of botulism can be divided into two broad categories: primary intoxication and primary infection followed by secondary intoxication.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 6: Endosome membrane. Toxin penetration by pH-induced translocation
Type: Eukaryotic cell or cell component Location: Cell membrane Function: Transporter Description: The botulinum toxin membrane translocation process includes, probably, some distinct events. These include a pH-induced change in toxin structure that results in exposure of previously occult hydrophobic domains; insertion of the toxin into the endosome membrane; translocation of the light chain from the lumenal to the cytosolic surface of the membrane; reduction of the single disulfide bond that links the heavy chain and light chain; uncoupling of the noncovalent forces that bond the heavy and light chains, with subsequent separation of chains; and restoration of light-chain structure associated with movement from an acidic environment (endosome) to a more neutral environment (cytosol).(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 7: M cells
Type: Eukaryotic cell or cell component Location: Cytoplasm Function: Transporter Description: In the case of M cells, the transport process can be more involved. The M cell Peyer's Patch complex can capture and process molecules, including partial metabolic breakdown, to present potential antigens to the mucosal immune system. M cells could outweigh the contribution of absorptive cells only if their transport rates were vastly higher. Thus, absorptive cells are the major participants in botulinum toxin transport.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 8: Nerve terminus cytosol. Toxin penetration to the nerve terminus cytosol
Type: Eukaryotic cell or cell component Location: Cytoplasm Function: Transporter Description: Since the light chains of botulinum toxins block neuroexocytosis by acting in the cytosol, at least this part of the toxin must reach the cell cytosol. All available evidence indicates that botulinum toxins do not enter the cell directly from the plasma membrane. Rather, they are endocytosed inside acidic cellular compartments.(<a href="#reference5267">Pellizzari et al., 1999</a>)
Bio-object 9: Neuromuscular junctions. Expression of toxicity-flaccid paralysis
Type: Pathway or action Location: Extracellular Description: The neurotransmitter acetylcholine is not released into neuromuscular junctions, leading to paralysis of myosin filaments. Blockade of transmitter release accounts for flaccid paralysis and autonomic dysfunction that is characteristic of the botulism disease. In untreated persons, paralysis extends beyond; death results from airway obstruction (pharyngeal and upper airway muscle paralysis) and inadequate tidal volume (diaphragmatic and accessory respiratory muscle paralysis). Although the toxin acts preferentially on cholinergic nerve endings, it does have the ability to block exocytosis from other nerve endings as well.(<a href="#reference5273">Anne et al., 2003</a>)
Bio-object 10: Plasma membrane. Toxin penetration by receptor-mediated endocytosis
Type: Eukaryotic cell or cell component Location: Cell membrane Function: Transporter Description: Botulinum toxin binds irreversibly to the presynaptic nerve ending - the heavy chain facilitates the binding and internalization of the toxin molecules into cholinergic neurons.Botulinum toxin enters the cells by receptor-mediated endocytosis and shortly thereafter escapes endosomes to act locally in the cytosol. For botulinum toxin, escape is known to be a pH-dependent phenomenon.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 11: Progenitor toxin complex. Formation and release
Type: Protein or gene complex Location: Cytoplasm Function: Enzyme Description: Neurotoxin is synthesized in seven immunologically distinct forms designated A, B, C, D, E, F, and G. Under natural conditions, each of these serotypes is released from clostridia in association with two classes of proteins: a family of hemagglutinins (HA) and a single nontoxin, nonhemagglutinin (NTNH). The complex that is composed of a toxin and noncovalently associated proteins is known as progenitor toxin. These nontoxic proteins may enhance toxicity, compared to toxin without the proteins, possibly by protecting the neurotoxin from proteolytic enzymes in the gut. Each of the serotypes responsible for human illness (mainly A, B, and E) forms a unique progenitor complex with associated proteins, but these associated proteins play no role in the signs and symptoms of clinical botulism.(<a href="#reference5263">Park et al., 2003</a>)(<a href="#reference5264">Kouguchi et al., 2002</a>)(<a href="#reference5265">Brin, 1997</a>)
Bio-object 12: Protoxin. Nicking by protease
Type: Protein or gene Location: Cytoplasm Function: Enzyme Description: This relatively inactive protoxin undergoes posttranslation modification, during which a trypsin-like enzyme cleaves (nicks) the single-chain molecule to generate a dichain molecule in which a heavy-chain polypeptide (100 kDa, HC-100) is linked by a disulfide bond to a light-chain polypeptide (50 kDa, LC-50). The dichain molecule is the agent responsible for the disease botulism.(<a href="#reference5263">Park et al., 2003</a>)
Bio-object 13: Receptor-Toxin. Toxin and target cell binding
Type: Microbe-host cell complex Location: Cell membrane Function: Transporter Description: From the site of transport cells, botulinum toxin diffuses in the body fluids, up to the presynaptic membrane of cholinergic endings where it binds.The receptor for botulinum toxin at the neuromuscular junction has not been unequivocally identified. Sialic acid-containing molecules, and perhaps gangliosides, might be implicated in toxin binding.It is clear now, that the botulinum toxin binds with high affinity to membrane; the binding domain of the toxin is localized within the heavy chain; and with the exception of serotypes C and D, which are closely related, have their own binding sites that are partially or wholly unique.(<a href="#reference5262">Simpson, 2004</a>)
Bio-object 14: Toxin (Neurotoxin). Expression of toxicity-blocking acetylcholine release
Type: Protein or gene Location: Cytoplasm Function: Enzyme Description: At normal neuromuscular transmission the SNARE (Soluble NSF-attachment Protein Receptor) proteins (SNAP-25 (Synaptosomal-Associated Protein of 25 kd), synaptobrevin (Vesicle-Associated Membrane Protein, or VAMP) and syntaxin) form a synaptic fusion complex which enables acetylcholine to be released into the neuromuscular junction. Release of acetylcholine at the neuromuscular junction is mediated by the assembly of a synaptic fusion complex that allows the membrane of the synaptic vesicle containing acetylcholine to fuse with the neuronal cell membrane. After membrane fusion, acetylcholine is released into the synaptic cleft and then bound by receptors on the muscle cell. The light chains of botulinum toxins are unique zinc proteases that have stringent substrate specificity and require exceptionally long substrates. From 15107500The light chains cleave specific sites on the SNARE proteins, preventing complete assembly of the synaptic fusion complex and thereby blocking acetylcholine release. Botulinum toxins types B, D, F, and G cleave synaptobrevin; types A, C, and E cleave SNAP-25; and type C cleaves syntaxin. Without acetylcholine release, the muscle is unable to contract and move.(<a href="#reference5269">Arnon et al., 2001</a>)(<a href="#reference5272">Segelke et al., 2004</a>)
Bio-object 15: Transport cells. Toxin binding to intestinal epithelial cell surface
Type: Eukaryotic cell or cell component Location: Extracellular Function: Enzyme Description: The two broad categories of etiology, primary or secondary intoxication, have one key feature in common: toxin in the lumen of the gut must penetrate membrane barriers to reach blood and lymph. Toxin molecules, certainly progenitor complexes of toxin or HA and NTNH, are too large for any significant rate of transcellular diffusion. It is likely that the toxin binds to a cell surface receptor that is linked to an efficient transport process.At initial stage botulinum toxin interacts with epithelial cells (namely, transport cells). The two logical choices of transport cells are absorptive epithelial cells and M cells. Both cell types, along with intercellular tight junctions, serve the purpose of creating a barrier between the lumen of the gut and the general circulation. Both cell types have the ability to capture certain molecules or macromolecular complexes and transport them.(<a href="#reference5262">Simpson, 2004</a>)
Interactions
Interaction 1: I1
Input Objects: Clostridium botulinum Output Objects: Botulinum protoxin GO Evidence Code: Inferred from Direct Assay Description: Botulinum toxin is synthesized by Clostridium botulinum as a nonactive protoxin.(<a href="#reference5263">Park et al., 2003</a>)
Interaction 2: I2
Input Objects: Botulinum protoxin Output Objects: Protoxin. Nicking by protease GO Evidence Code: Inferred from Direct Assay Description: Posttranslation modification of Botulinum protoxin. Trypsin-like protease nicks the single-chain molecule to generate a dichain molecule.(<a href="#reference5263">Park et al., 2003</a>)
Interaction 3: I3
Input Objects: Protoxin. Nicking by protease Output Objects: Progenitor toxin complex. Formation and release GO Evidence Code: Inferred from Direct Assay Description: The complex that is composed of a toxin and noncovalently associated proteins (hemagglutinins, nonhemagglutinins) is known as progenitor toxin.(<a href="#reference5264">Kouguchi et al., 2002</a>)(<a href="#reference5265">Brin, 1997</a>)
Interaction 4: I4
Input Objects: Progenitor toxin complex. Formation and release Output Objects: Botulinum toxin. Food infection/Primary intoxication, Botulinum toxin. Primary human infection/Secondary intoxication GO Evidence Code: Inferred from Direct Assay Description: Two broad categories of botulism: primary intoxication and primary infection followed by secondary intoxication.(<a href="#reference5262">Simpson, 2004</a>)
Interaction 5: I5
Input Objects: Progenitor toxin complex. Formation and release, Botulinum toxin. Food infection/Primary intoxication, Botulinum toxin. Primary human infection/Secondary intoxication Output Objects: Transport cells. Toxin binding to intestinal epithelial cell surface GO Evidence Code: Inferred from Direct Assay Description: At initial stage of action botulinum toxin interacts with gut epithelial cells (transport cells). It is more likely that the toxin binds exploitatively to a receptor that is linked to an efficient transport process.(<a href="#reference5270">Maksymowych et al., 1998</a>)(<a href="#reference5271">Maksymowych et al., 1999</a>)
Interaction 6: I6
Input Objects: Transport cells. Toxin binding to intestinal epithelial cell surface Output Objects: Absorptive cells, M cells GO Evidence Code: Inferred from Direct Assay Description: Regardless of whether poisoning is due to primary or secondary intoxication, these two broad categories of etiology have one key feature in common: toxin in the lumen of the gut must penetrate membrane barriers to reach blood and lymph. Absorptive epithelial cells and M cells have the ability to transport certain molecules or macromolecular complexes. The major route utilized by toxin is transcytosis across absorptive cells.(<a href="#reference5262">Simpson, 2004</a>)
Interaction 7: I7
Input Objects: Absorptive cells, M cells Output Objects: Receptor-Toxin. Toxin and target cell binding GO Evidence Code: Inferred from Direct Assay Description: At transport cells, receptor binding leads to receptor-mediated endocytosis that is followed by a cascade of receptor-dependent events. The toxin is not modified during transport.(<a href="#reference5266">Kalandakanond et al., 2001</a>)
Interaction 8: I8
Input Objects: Receptor-Toxin. Toxin and target cell binding Output Objects: Nerve terminus cytosol. Toxin penetration to the nerve terminus cytosol GO Evidence Code: Inferred from Direct Assay Description: Botulinum toxin is endocytosed inside acidic cellular compartment.(<a href="#reference5267">Pellizzari et al., 1999</a>)
Interaction 9: I9
Input Objects: Nerve terminus cytosol. Toxin penetration to the nerve terminus cytosol Output Objects: Plasma membrane. Toxin penetration by receptor-mediated endocytosis GO Evidence Code: Inferred from Direct Assay Description: Botulinum toxin binds irreversibly to the presynaptic nerve cell membrane ending and enters the cells by receptor-mediated endocytosis and then shortly escapes endosomes to act locally in the cytosol.(<a href="#reference5262">Simpson, 2004</a>)
Interaction 10: I10
Input Objects: Plasma membrane. Toxin penetration by receptor-mediated endocytosis Output Objects: Endosome membrane. Toxin penetration by pH-induced translocation GO Evidence Code: Inferred from Direct Assay Description: Toxin that enters nerve endings is largely confined to early endosomes that possess a membrane proton pump. Acidification of the lumen causes marked structural changes in the toxin molecule. Although many details of the process are still unknown, one can deduce that the toxin inserts into the endosome membrane, and this is accompanied by reduction of the disulfide bond, separation of the two chains, and eventual escape of the light chain to the cytosol. Productively internalized toxin that reaches the cytosol blocks exocytosis.(<a href="#reference5268">Maksymowych et al., 2004</a>)
Interaction 11: I11
Input Objects: Endosome membrane. Toxin penetration by pH-induced translocation Output Objects: Toxin (Neurotoxin). Expression of toxicity-blocking acetylcholine release GO Evidence Code: Inferred from Direct Assay Description: The light chain (zinc protease) of botulinum toxin cleaves specific sites on the SNARE (Soluble NSF-attachment Protein Receptor) proteins, preventing complete assembly of the synaptic fusion complex and thereby blocking acetylcholine release.(<a href="#reference5269">Arnon et al., 2001</a>)
Interaction 12: I12
Input Objects: Toxin (Neurotoxin). Expression of toxicity-blocking acetylcholine release Output Objects: Neuromuscular junctions. Expression of toxicity-flaccid paralysis GO Evidence Code: Inferred from Direct Assay Description: All Clostridium botulinum serotypes have similar neurotoxic properties resulting in flaccid muscle paralysis.(<a href="#reference5273">Anne et al., 2003</a>)(<a href="#reference5274">Matarasso, 2003</a>)
Pathways

Phinet Name: Clostridium botulinum