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Justin Hessinger Tess Demeter. A toxin is a poisonous substance that is produced by one organism that has negative effects on another organism A toxin.

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Presentation on theme: "Justin Hessinger Tess Demeter. A toxin is a poisonous substance that is produced by one organism that has negative effects on another organism A toxin."— Presentation transcript:

1 Justin Hessinger Tess Demeter

2 A toxin is a poisonous substance that is produced by one organism that has negative effects on another organism A toxin is a poisonous substance that is produced by one organism that has negative effects on another organism They include small molecules, peptides, or proteins They include small molecules, peptides, or proteins

3 No, but really, we felt that amatoxins (alpha-amitin) were interesting because they are the cause of over 100 human deaths annually worldwide. The organisms that are affected by these seemingly harmless shrooms suffer mild symptoms hours after ingestion that completely disappear only to kill their unexpecting victims days later. No, but really, we felt that amatoxins (alpha-amitin) were interesting because they are the cause of over 100 human deaths annually worldwide. The organisms that are affected by these seemingly harmless shrooms suffer mild symptoms hours after ingestion that completely disappear only to kill their unexpecting victims days later. Cause they’re freakin’ cool!!!

4 Class of toxins produced by certain types of Amanita species of mushrooms Class of toxins produced by certain types of Amanita species of mushrooms Bicyclic octapeptide derivatives Bicyclic octapeptide derivatives zymogenic until they reach the host There are at least 10 known variants of amanitin toxin There are at least 10 known variants of amanitin toxin Amatoxins account for 95% of fatal mushroom poisoning Amatoxins account for 95% of fatal mushroom poisoning

5 Alpha-Amanitin is produced by some of the mushrooms from the genus Amanita. Alpha-Amanitin is produced by some of the mushrooms from the genus Amanita. The most common/deadly species: The most common/deadly species: Death cap mushroom (Amanita phalloides) Destroying Angel (Amanita virosa)

6 There are around 600 known species, some of which are actually edible There are around 600 known species, some of which are actually edible They are distributed throughout the world and found in regions such as; They are distributed throughout the world and found in regions such as; Mexico Mexico Central Africa Central Africa Europe Europe Canada Canada South America South America United States United States

7 Alpha-amanitin causes inhibition of RNA Polymerase II by: Alpha-amanitin causes inhibition of RNA Polymerase II by: strong hydrogen bonding to the bridge helix on RNA Polymerase referred to as pore 1. strong hydrogen bonding to the bridge helix on RNA Polymerase referred to as pore 1. penetrates at pore1 and regulates translocation and stops elongation at or before translocation occurs. penetrates at pore1 and regulates translocation and stops elongation at or before translocation occurs. it does not affect NTP loading, rather it prevents the bridge helix from flexing thus preventing DNA translocation. it does not affect NTP loading, rather it prevents the bridge helix from flexing thus preventing DNA translocation.

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9 Inhibition of RNA Polymerase II induces stress activated kinases. Inhibition of RNA Polymerase II induces stress activated kinases. Phosphorylation of p53 protein fraction on serine residues Phosphorylation of p53 protein fraction on serine residues This causes suppression of p53 degredation This causes suppression of p53 degredation The accumulated p53 are translocated to mitochondria and this increases permeability of the outer membrane which causes the release of cytochrome C to the cytosol. The accumulated p53 are translocated to mitochondria and this increases permeability of the outer membrane which causes the release of cytochrome C to the cytosol. This causes programmed cell death. This causes programmed cell death.

10 Amanitin toxicity is also associated with an increase in the production of reactive oxygen species through lipid peroxidation Amanitin toxicity is also associated with an increase in the production of reactive oxygen species through lipid peroxidation Increases hepatotoxicity Increases hepatotoxicity Although many studies note this reaction during amanitin toxicity, they are unclear in stating how amanintin itself induces this response Although many studies note this reaction during amanitin toxicity, they are unclear in stating how amanintin itself induces this response

11 These are all very similar in structure, often only differing by 1 R-group. These are all very similar in structure, often only differing by 1 R-group. α-amanitin β-amanitin γ-amanitin ε-amanitin Amanullin Amanullinic Acid Amaninamide Amanin Proamanullin

12 Toxicity depends on many factors: Toxicity depends on many factors: Amount of toxin ingested Amount of toxin ingested Age; multi-organ failure between 6-8 days ranges Age; multi-organ failure between 6-8 days ranges 10-20% adults 10-20% adults 22-50% children 22-50% children Immune Function Immune Function Exposure Pathway; majority of exposure is through oral administration, but there are cases of intravenous injection. Exposure Pathway; majority of exposure is through oral administration, but there are cases of intravenous injection.

13 Although amatoxins are extremely toxic to humans and difficult to treat, there are species that are immune to these toxins. The mechanisms of their immunity are highly misunderstood Although amatoxins are extremely toxic to humans and difficult to treat, there are species that are immune to these toxins. The mechanisms of their immunity are highly misunderstood Caenorhabditis elegans Drosophila melanogaster

14 Structural mutations of RNAPII Structural mutations of RNAPII Alpha-amanitin is unable to bind in the proper location Alpha-amanitin is unable to bind in the proper location Most plausible Most plausible Metabolic degradation Metabolic degradation There are 3 possible modes of degredation although the most plausible is by cytochrome P450’s There are 3 possible modes of degredation although the most plausible is by cytochrome P450’s Behavioral Avoidance Behavioral Avoidance

15 Why are the mushrooms that develop these amatoxins not affected by them? Why are the mushrooms that develop these amatoxins not affected by them? Protein precursers are usually used when the protein is potentially harmful to the host, but needs to be available on short notice and in large quantities. Protein precursers are usually used when the protein is potentially harmful to the host, but needs to be available on short notice and in large quantities. The active form of the toxin is not present in the host organisms. The active form of the toxin is not present in the host organisms. When entering a target species activation occurs by cleavage of the 8 terminal amino acids present on the toxin When entering a target species activation occurs by cleavage of the 8 terminal amino acids present on the toxin Enzymatic reaction catalyzed by prolyloligopeptidase Enzymatic reaction catalyzed by prolyloligopeptidase

16 The most common pathway is through oral ingestion The most common pathway is through oral ingestion The toxin may be isolated and injected peritoneally or intravenously, although this method is really only reserved for lab research and due to ethicalities, humans are not generally the test subjects The toxin may be isolated and injected peritoneally or intravenously, although this method is really only reserved for lab research and due to ethicalities, humans are not generally the test subjects

17 For humans: For humans: 6-12 hours: incubation (no observable adverse effects) 6-12 hours: incubation (no observable adverse effects) Starts with gastrointestinal symptoms Starts with gastrointestinal symptoms Vomiting, diarrhea, abdominal pain, hypoglycemia, and dehydration Vomiting, diarrhea, abdominal pain, hypoglycemia, and dehydration hours: evident hepatocellular damage hours: evident hepatocellular damage Coagulopathy and acute liver failure Coagulopathy and acute liver failure 6-8 days: acute liver and kidney failure 6-8 days: acute liver and kidney failure Haemorrhaging, encephalopathy, coma, and death Haemorrhaging, encephalopathy, coma, and death

18 About 10% of individuals exposed to amanitin in will die in the U.S. About 10% of individuals exposed to amanitin in will die in the U.S. World wide, about 50% of those exposed to aminitin toxin will die due to inadequate and/or less accessible medical care World wide, about 50% of those exposed to aminitin toxin will die due to inadequate and/or less accessible medical care Of those that survive, severe liver and kidney damage is extremely common and some patients may die due to complications pertaining to organ failure Of those that survive, severe liver and kidney damage is extremely common and some patients may die due to complications pertaining to organ failure

19 Silybin Silybin Comes from the Milk Thistle (Silybum marianum) Comes from the Milk Thistle (Silybum marianum) Competes with amanitin for transport systems in the plasma membranes in cells Competes with amanitin for transport systems in the plasma membranes in cells Causes a decrease in penetration of amanitin into hepatocytes Causes a decrease in penetration of amanitin into hepatocytes Thioctic Acid Thioctic Acid Powerful antioxidant Powerful antioxidant Inhibits free radical formation through lipid peroxidation Inhibits free radical formation through lipid peroxidation Prevents further oxidative tissue damage Prevents further oxidative tissue damage

20 Benzylpenicillin Benzylpenicillin Inhibits trasporting polypeptide 1B3 in the plasma membrane which impedes amanitin penetration of hepatocytes Inhibits trasporting polypeptide 1B3 in the plasma membrane which impedes amanitin penetration of hepatocytes N-acetylcysteine N-acetylcysteine Unclear mechanism Unclear mechanism May inhibit hepatic uptake of amanitin May inhibit hepatic uptake of amanitin May act as strong antioxidant May act as strong antioxidant Cimetidine Cimetidine Listed as a treatment, but there was no known mechanism found Listed as a treatment, but there was no known mechanism found

21 To date it seems that the most effective treatment involves pumping the stomach right after eating the toxin. To date it seems that the most effective treatment involves pumping the stomach right after eating the toxin. Because this treatment is extremely time dependant and symptoms may take hours to show up, it’s not largely reliable. Because this treatment is extremely time dependant and symptoms may take hours to show up, it’s not largely reliable. Although there are other proposed treatments, the ones that we have included are the most common across notable sources. Although there are other proposed treatments, the ones that we have included are the most common across notable sources. The only treatment other than emptying the stomach content, that has shown any potential benefits through decrease in hepatic necrosis is Silybin. The only treatment other than emptying the stomach content, that has shown any potential benefits through decrease in hepatic necrosis is Silybin. These results, as noted in various studies, can be limited or nullified by the experimental conditions. These results, as noted in various studies, can be limited or nullified by the experimental conditions. There are other studies that have resulted positively with different toxins, but themselves list a variety of limitations and problems with their research There are other studies that have resulted positively with different toxins, but themselves list a variety of limitations and problems with their research

22 Before eating a random mushroom, there are a few things you can do: Before eating a random mushroom, there are a few things you can do: Meixner Test: tests for presence of amanitin in the mushroom Meixner Test: tests for presence of amanitin in the mushroom Limitations: cannot differentiate between toxic and hallucinogenic species because psilocin gives a false positive Limitations: cannot differentiate between toxic and hallucinogenic species because psilocin gives a false positive Mezler’s Reagent: tests for spores of amyloid, non-amyloid, and dextranoid Mezler’s Reagent: tests for spores of amyloid, non-amyloid, and dextranoid Used to identify different types of fungi Used to identify different types of fungi

23 There is potential for therapeutic use against pancreatic carcinoma There is potential for therapeutic use against pancreatic carcinoma Through conjugation of Amanitin to Anti- epithelial cell adhesion molecules (EpCAM), antitumor properties have been shown Through conjugation of Amanitin to Anti- epithelial cell adhesion molecules (EpCAM), antitumor properties have been shown EpCAM’s are over expressed in many cancers. EpCAM’s are over expressed in many cancers. The conjugated molecules are able to bind these markers and initiate apoptosis The conjugated molecules are able to bind these markers and initiate apoptosis

24 Although amatoxins are widely distributed and extremely toxic, they are still very misunderstood. Although amatoxins are widely distributed and extremely toxic, they are still very misunderstood. The toxic effects suffered from exposure to amatoxins are difficult to treat and the mechanisms behind their action is still unclear. Despite these difficulties, continued research and understanding is very important because they may be the key to future treatments for other illnesses and survivorship. The toxic effects suffered from exposure to amatoxins are difficult to treat and the mechanisms behind their action is still unclear. Despite these difficulties, continued research and understanding is very important because they may be the key to future treatments for other illnesses and survivorship.

25 So remember kids, be careful when picking your shrooms!!! So remember kids, be careful when picking your shrooms!!! As we understand that peer-reviewing is a time-consuming process and you may be tired, here is some legal fun you can have with some awesome shrooms. ENJOY!!!

26 Brueckner, F. and Cramer, P Structural basis of transcription inhibition by α-amanitin and implications for RNA polymerase II translocation. Nature Structural & Molecular Biology. Vol. 15, No. 8: Madalan, J., Piotrowska, A., Gomulkiewicz, A., Sozanski, T., Podhorska-Okolow, M., Szelag, A., and Dziegiel, P Benzylpenicyllin and acetylcysteine protection from α-amanitin-induced apoptosis in human hepatocyte cultures. Experimental and Toxicologic Pathology. Vol 63: Moldenhauer, G., Salnikiv, A., Luttgau, S., & Harr, I. (2012). Therapeutic potential of amanitin-conjugated anti-epithelial cell adhesion molecule monoclonal antibody against pancreatic carcinoma. JNCI, Nikolova, G., Karamalakova, Y., Hadjibojeva, P., Georgiev, T., Tolekova, A., Gadjeva, V., and Zheleva, A Severe mushroom toxin alpha amanitin causes generation of reactive oxygen species in liver tissues of mice – A comparative study by two different instrumental methods. Trakia Journal of Sciences. Vol. 8, No. 2: Springer, M Effect of Alpha-Amanitin. Retrieved April 16, 2012 from Springer, M Effect of Alpha-Amanitin. Retrieved April 16, 2012 from Stump, A., Jabonski, S., Bouton, L., and Wilder, J Distribution and mechanism of α-amanitin tolerance in mycophagous Drosophila. Environmental Entomology. Vol. 40, No. 6: Thiel, C., Thiel, K., Klingert, W., Diewold, A., Scheuermann, K., Hawerkamp, E., Lauber, J., Scheppach, J., Morgalla, M., Konigsrainer, A., and Schenk, M The enterohepatic circulation of amanitin: Kinetics and therapeutical implications. Toxicology Letters. Vol. 203: Tsao, D., Park, N., Nag, A., and Martinson, H Prolonged α-amanitin treatment of cells for studying mutated polymerases causes degradation of DSIF160 and other proteins. RNA Society. Vol. 2: Tong, Tri C. "Comparative Treatment of."Annals of Emergency Medicine. 50. (2007): Print. Zheleva, A., Gadjeva, V., and Zhelev. M Free radical formation might contribute to the severe amatoxin hepatotoxicity. Trakia Journal of Sciences. Vol. 1, No. 3: Zheleva, A., Gadjeva, V., and Zhelev. M Free radical formation might contribute to the severe amatoxin hepatotoxicity. Trakia Journal of Sciences. Vol. 1, No. 3: mushroom poisoning syndromes. (2012). Retrieved from


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