Toxins

Editorial

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Open AccessEditorial

Ribosome Inactivating Proteins: From Plant Defense to Treatments against Human Misuse or Diseases

by Julien Barbier and Daniel Gillet

Toxins 2018, 10(4), 160; https://doi.org/10.3390/toxins10040160 - 18 Apr 2018

Cited by 9 | Viewed by 3627

Abstract

Ribosome inactivating proteins (RIPs) form a vast family of hundreds of toxins from plants, fungi, algae, and bacteria. RIP activities have also been detected in animal tissues. They exert an N-glycosydase catalytic activity that is targeted to a single adenine of a ribosomal [...] Read more.

Ribosome inactivating proteins (RIPs) form a vast family of hundreds of toxins from plants, fungi, algae, and bacteria. RIP activities have also been detected in animal tissues. They exert an N-glycosydase catalytic activity that is targeted to a single adenine of a ribosomal RNA, thereby blocking protein synthesis and leading intoxicated cells to apoptosis. In many cases, they have additional depurinating activities that act against other nucleic acids, such as viral RNA and DNA, or genomic DNA. Although their role remains only partially understood, their functions may be related to plant defense against predators and viruses, plant senescence, or bacterial pathogenesis. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

Research

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6199 KiB

Open AccessFeature PaperArticle

A Supercluster of Neutralizing Epitopes at the Interface of Ricin’s Enzymatic (RTA) and Binding (RTB) Subunits

by Amanda Y. Poon, David J. Vance, Yinghui Rong, Dylan Ehrbar and Nicholas J. Mantis

Toxins 2017, 9(12), 378; https://doi.org/10.3390/toxins9120378 - 23 Nov 2017

Cited by 6 | Viewed by 5086

Abstract

As part of an effort to engineer ricin antitoxins and immunotherapies, we previously produced and characterized a collection of phage-displayed, heavy chain-only antibodies (V_HHs) from alpacas that had been immunized with ricin antigens. In our initial screens, we identified nine V [...] Read more.

As part of an effort to engineer ricin antitoxins and immunotherapies, we previously produced and characterized a collection of phage-displayed, heavy chain-only antibodies (V_HHs) from alpacas that had been immunized with ricin antigens. In our initial screens, we identified nine V_HHs directed against ricin toxin’s binding subunit (RTB), but only one, JIZ-B7, had toxin-neutralizing activity. Linking JIZ-B7 to different V_HHs against ricin’s enzymatic subunit (RTA) resulted in several bispecific antibodies with potent toxin-neutralizing activity in vitro and in vivo. JIZ-B7 may therefore be an integral component of a future V_HH-based neutralizing agent (VNA) for ricin toxin. In this study, we now localize, using competitive ELISA, JIZ-B7’s epitope to a region of RTB’s domain 2 sandwiched between the high-affinity galactose/N-acetylgalactosamine (Gal/GalNAc)-binding site and the boundary of a neutralizing hotspot on RTA known as cluster II. Analysis of additional RTB (n = 8)- and holotoxin (n = 4)-specific V_HHs from a recent series of screens identified a “supercluster” of neutralizing epitopes at the RTA-RTB interface. Among the V_HHs tested, toxin-neutralizing activity was most closely associated with epitope proximity to RTA, and not interference with RTB’s ability to engage Gal/GalNAc receptors. We conclude that JIZ-B7 is representative of a larger group of potent toxin-neutralizing antibodies, possibly including many described in the literature dating back several decades, that recognize tertiary and possibly quaternary epitopes located at the RTA-RTB interface and that target a region of vulnerability on ricin toxin. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Production, Characterisation and Testing of an Ovine Antitoxin against Ricin; Efficacy, Potency and Mechanisms of Action

by Sarah J. C. Whitfield, Gareth D. Griffiths, Dominic C. Jenner, Robert J. Gwyther, Fiona M. Stahl, Lucy J. Cork, Jane L. Holley, A. Christopher Green and Graeme C. Clark

Toxins 2017, 9(10), 329; https://doi.org/10.3390/toxins9100329 - 18 Oct 2017

Cited by 13 | Viewed by 5588

Abstract

Ricin is a type II ribosome-inactivating toxin that catalytically inactivates ribosomes ultimately leading to cell death. The toxicity of ricin along with the prevalence of castor beans (its natural source) has led to its increased notoriety and incidences of nefarious use. Despite these [...] Read more.

Ricin is a type II ribosome-inactivating toxin that catalytically inactivates ribosomes ultimately leading to cell death. The toxicity of ricin along with the prevalence of castor beans (its natural source) has led to its increased notoriety and incidences of nefarious use. Despite these concerns, there are no licensed therapies available for treating ricin intoxication. Here, we describe the development of a F(ab’)₂ polyclonal ovine antitoxin against ricin and demonstrate the efficacy of a single, post-exposure, administration in an in vivo murine model of intoxication against aerosolised ricin. We found that a single dose of antitoxin afforded a wide window of opportunity for effective treatment with 100% protection observed in mice challenged with aerosolised ricin when given 24 h after exposure to the toxin and 75% protection when given at 30 h. Treated mice had reduced weight loss and clinical signs of intoxication compared to the untreated control group. Finally, using imaging flow cytometry, it was found that both cellular uptake and intracellular trafficking of ricin toxin to the Golgi apparatus was reduced in the presence of the antitoxin suggesting both actions can contribute to the therapeutic mechanism of a polyclonal antitoxin. Collectively, the research highlights the significant potential of the ovine F(ab’)₂ antitoxin as a treatment for ricin intoxication. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

A Monoclonal–Monoclonal Antibody Based Capture ELISA for Abrin

by Christina C. Tam, Luisa W. Cheng, Xiaohua He, Paul Merrill, David Hodge and Larry H. Stanker

Toxins 2017, 9(10), 328; https://doi.org/10.3390/toxins9100328 - 18 Oct 2017

Cited by 6 | Viewed by 5836

Abstract

Abrin, one of the most highly potent toxins in the world, is derived from the plant, Abrus precatorius. Because of its high toxicity, it poses potential bioterror risks. Therefore, a need exists for new reagents and technologies that would be able to [...] Read more.

Abrin, one of the most highly potent toxins in the world, is derived from the plant, Abrus precatorius. Because of its high toxicity, it poses potential bioterror risks. Therefore, a need exists for new reagents and technologies that would be able to rapidly detect abrin contamination as well as lead to new therapeutics. We report here a group of abrin-specific monoclonal antibodies (mAbs) that recognize abrin A-chain, intact A–B chain toxin, and agglutinin by Western blot. Additionally, these mAbs were evaluated for their ability to serve as capture antibodies for a sandwich (capture) ELISA. All possible capture–detector pairs were evaluated and the best antibody pair identified and optimized for a capture ELISA. The capture ELISA based on this capture–detector mAb pair had a limit of detection (L.O.D) of ≈1 ng/mL measured using three independent experiments. The assay did not reveal any false positives with extracts containing other potential ribosome-inactivating proteins (RIPs). Thus, this new capture ELISA uses mAbs for both capture and detection; has no cross-reactivity against other plant RIPs; and has a sensitivity comparable to other reported capture ELISAs using polyclonal antibodies as either capture or detector. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Abrin Toxicity and Bioavailability after Temperature and pH Treatment

by Christina C. Tam, Thomas D. Henderson, Larry H. Stanker, Xiaohua He and Luisa W. Cheng

Toxins 2017, 9(10), 320; https://doi.org/10.3390/toxins9100320 - 13 Oct 2017

Cited by 10 | Viewed by 8683

Abstract

Abrin, one of most potent toxins known to man, is derived from the rosary pea (jequirity pea), Abrus precatorius and is a potential bioterror weapon. The temperature and pH stability of abrin was evaluated with an in vitro cell free translation (CFT) assay, [...] Read more.

Abrin, one of most potent toxins known to man, is derived from the rosary pea (jequirity pea), Abrus precatorius and is a potential bioterror weapon. The temperature and pH stability of abrin was evaluated with an in vitro cell free translation (CFT) assay, a Vero cell culture cytotoxicity assay, and an in vivo mouse bioassay. pH treatment of abrin had no detrimental effect on its stability and toxicity as seen either in vitro or in vivo. Abrin exposure to increasing temperatures did not completely abrogate protein translation. In both the cell culture cytotoxicity model and the mouse bioassay, abrin’s toxic effects were completely abrogated if the toxin was exposed to temperatures of 74 °C or higher. In the cell culture model, 63 °C-treated abrin had a 30% reduction in cytotoxicity which was validated in the in vivo mouse bioassay with all mice dying but with a slight time-to-death delay as compared to the non-treated abrin control. Since temperature inactivation did not affect abrin’s ability to inhibit protein synthesis (A-chain), we hypothesize that high temperature treatment affected abrin’s ability to bind to cellular receptors (affecting B-chain). Our results confirm the absolute need to validate in vitro cytotoxicity assays with in vivo mouse bioassays. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Shiga Toxins Induce Apoptosis and ER Stress in Human Retinal Pigment Epithelial Cells

by Jun-Young Park, Yu-Jin Jeong, Sung-Kyun Park, Sung-Jin Yoon, Song Choi, Dae Gwin Jeong, Su Wol Chung, Byung Joo Lee, Jeong Hun Kim, Vernon L. Tesh, Moo-Seung Lee and Young-Jun Park

Toxins 2017, 9(10), 319; https://doi.org/10.3390/toxins9100319 - 13 Oct 2017

Cited by 25 | Viewed by 7278

Abstract

Shiga toxins (Stxs) produced by Shiga toxin-producing bacteria Shigella dysenteriae serotype 1 and select serotypes of Escherichia coli are the most potent known virulence factors in the pathogenesis of hemorrhagic colitis progressing to potentially fatal systemic complications such as acute renal failure, blindness [...] Read more.

Shiga toxins (Stxs) produced by Shiga toxin-producing bacteria Shigella dysenteriae serotype 1 and select serotypes of Escherichia coli are the most potent known virulence factors in the pathogenesis of hemorrhagic colitis progressing to potentially fatal systemic complications such as acute renal failure, blindness and neurological abnormalities. Although numerous studies have defined apoptotic responses to Shiga toxin type 1 (Stx1) or Shiga toxin type 2 (Stx2) in a variety of cell types, the potential significance of Stx-induced apoptosis of photoreceptor and pigmented cells of the eye following intoxication is unknown. We explored the use of immortalized human retinal pigment epithelial (RPE) cells as an in vitro model of Stx-induced retinal damage. To the best of our knowledge, this study is the first report that intoxication of RPE cells with Stxs activates both apoptotic cell death signaling and the endoplasmic reticulum (ER) stress response. Using live-cell imaging analysis, fluorescently labeled Stx1 or Stx2 were internalized and routed to the RPE cell endoplasmic reticulum. RPE cells were significantly sensitive to wild type Stxs by 72 h, while the cells survived challenge with enzymatically deficient mutant toxins (Stx1A⁻ or Stx2A⁻). Upon exposure to purified Stxs, RPE cells showed activation of a caspase-dependent apoptotic program involving a reduction of mitochondrial transmembrane potential (Δψ_m), increased activation of ER stress sensors IRE1, PERK and ATF6, and overexpression CHOP and DR5. Finally, we demonstrated that treatment of RPE cells with Stxs resulted in the activation of c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38MAPK), suggesting that the ribotoxic stress response may be triggered. Collectively, these data support the involvement of Stx-induced apoptosis in ocular complications of intoxication. The evaluation of apoptotic responses to Stxs by cells isolated from multiple organs may reveal unique functional patterns of the cytotoxic actions of these toxins in the systemic complications that follow ingestion of toxin-producing bacteria. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Total Body Irradiation Mitigates Inflammation and Extends the Therapeutic Time Window for Anti-Ricin Antibody Treatment against Pulmonary Ricinosis in Mice

by Yoav Gal, Anita Sapoznikov, Reut Falach, Sharon Ehrlich, Moshe Aftalion, Chanoch Kronman and Tamar Sabo

Toxins 2017, 9(9), 278; https://doi.org/10.3390/toxins9090278 - 11 Sep 2017

Cited by 9 | Viewed by 3968

Abstract

Ricin, a highly toxic plant-derived toxin, is considered a potential weapon in biowarfare and bioterrorism due to its pronounced toxicity, high availability, and ease of preparation. Pulmonary exposure to ricin results in the generation of an acute edematous inflammation followed by respiratory insufficiency [...] Read more.

Ricin, a highly toxic plant-derived toxin, is considered a potential weapon in biowarfare and bioterrorism due to its pronounced toxicity, high availability, and ease of preparation. Pulmonary exposure to ricin results in the generation of an acute edematous inflammation followed by respiratory insufficiency and death. Massive neutrophil recruitment to the lungs may contribute significantly to ricin-mediated morbidity. In this study, total body irradiation (TBI) served as a non-pharmacological tool to decrease the potential neutrophil-induced lung injury. TBI significantly postponed the time to death of intranasally ricin-intoxicated mice, given that leukopenia remained stable following intoxication. This increase in time to death coincided with a significant reduction in pro-inflammatory marker levels, and led to marked extension of the therapeutic time window for anti-ricin antibody treatment. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Two Saporin-Containing Immunotoxins Specific for CD20 and CD22 Show Different Behavior in Killing Lymphoma Cells

by Letizia Polito, Daniele Mercatelli, Massimo Bortolotti, Stefania Maiello, Alice Djemil, Maria Giulia Battelli and Andrea Bolognesi

Toxins 2017, 9(6), 182; https://doi.org/10.3390/toxins9060182 - 30 May 2017

Cited by 20 | Viewed by 5085

Abstract

Immunotoxins (ITs) are hybrid proteins combining the binding specificity of antibodies with the cytocidal properties of toxins. They represent a promising approach to lymphoma therapy. The cytotoxicity of two immunotoxins obtained by chemical conjugation of the plant toxin saporin-S6 with the anti-CD20 chimeric [...] Read more.

Immunotoxins (ITs) are hybrid proteins combining the binding specificity of antibodies with the cytocidal properties of toxins. They represent a promising approach to lymphoma therapy. The cytotoxicity of two immunotoxins obtained by chemical conjugation of the plant toxin saporin-S6 with the anti-CD20 chimeric antibody rituximab and the anti-CD22 murine antibody OM124 were evaluated on the CD20-/CD22-positive cell line Raji. Both ITs showed strong cytotoxicity for Raji cells, but the anti-CD22 IT was two logs more efficient in killing, probably because of its faster internalization. The anti-CD22 IT gave slower but greater caspase activation than the anti-CD20 IT. The cytotoxic effect of both immunotoxins can be partially prevented by either the pan-caspase inhibitor Z-VAD or the necroptosis inhibitor necrostatin-1. Oxidative stress seems to be involved in the cell killing activity of anti-CD20 IT, as demonstrated by the protective role of the H₂O₂ scavenger catalase, but not in that of anti-CD22 IT. Moreover, the IT toxicity can be augmented by the contemporary administration of other chemotherapeutic drugs, such as PS-341, MG-132, and fludarabine. These results contribute to the understanding of the immunotoxin mechanism of action that is required for their clinical use, either alone or in combination with other drugs. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Differences in Ribosome Binding and Sarcin/Ricin Loop Depurination by Shiga and Ricin Holotoxins

by Xiao-Ping Li and Nilgun E. Tumer

Toxins 2017, 9(4), 133; https://doi.org/10.3390/toxins9040133 - 11 Apr 2017

Cited by 20 | Viewed by 5102

Abstract

Both ricin and Shiga holotoxins display no ribosomal activity in their native forms and need to be activated to inhibit translation in a cell-free translation inhibition assay. This is because the ribosome binding site of the ricin A chain (RTA) is blocked by [...] Read more.

Both ricin and Shiga holotoxins display no ribosomal activity in their native forms and need to be activated to inhibit translation in a cell-free translation inhibition assay. This is because the ribosome binding site of the ricin A chain (RTA) is blocked by the B subunit in ricin holotoxin. However, it is not clear why Shiga toxin 1 (Stx1) or Shiga toxin 2 (Stx2) holotoxin is not active in a cell-free system. Here, we compare the ribosome binding and depurination activity of Stx1 and Stx2 holotoxins with the A1 subunits of Stx1 and Stx2 using either the ribosome or a 10-mer RNA mimic of the sarcin/ricin loop as substrates. Our results demonstrate that the active sites of Stx1 and Stx2 holotoxins are blocked by the A2 chain and the B subunit, while the ribosome binding sites are exposed to the solvent. Unlike ricin, which is enzymatically active, but cannot interact with the ribosome, Stx1 and Stx2 holotoxins are enzymatically inactive but can interact with the ribosome. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessCommunication

Yeast Reporter Assay to Identify Cellular Components of Ricin Toxin A Chain Trafficking

by Björn Becker, Tina Schnöder and Manfred J. Schmitt

Toxins 2016, 8(12), 366; https://doi.org/10.3390/toxins8120366 - 6 Dec 2016

Cited by 6 | Viewed by 4878

Abstract

RTA, the catalytic A-subunit of the ribosome inactivating A/B toxin ricin, inhibits eukaryotic protein biosynthesis by depurination of 28S rRNA. Although cell surface binding of ricin holotoxin is mainly mediated through its B-subunit (RTB), sole application of RTA is also toxic, albeit to [...] Read more.

RTA, the catalytic A-subunit of the ribosome inactivating A/B toxin ricin, inhibits eukaryotic protein biosynthesis by depurination of 28S rRNA. Although cell surface binding of ricin holotoxin is mainly mediated through its B-subunit (RTB), sole application of RTA is also toxic, albeit to a significantly lower extent, suggesting alternative pathways for toxin uptake and transport. Since ricin toxin trafficking in mammalian cells is still not fully understood, we developed a GFP-based reporter assay in yeast that allows rapid identification of cellular components required for RTA uptake and subsequent transport through a target cell. We hereby show that Ypt6p, Sft2p and GARP-complex components play an important role in RTA transport, while neither the retromer complex nor COPIB vesicles are part of the transport machinery. Analyses of yeast knock-out mutants with chromosomal deletion in genes whose products regulate ADP-ribosylation factor GTPases (Arf-GTPases) and/or retrograde Golgi-to-ER (endoplasmic reticulum) transport identified Sso1p, Snc1p, Rer1p, Sec22p, Erv46p, Gea1p and Glo3p as novel components in RTA transport, suggesting the developed reporter assay as a powerful tool to dissect the multistep processes of host cell intoxication in yeast. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Improvement of the Pharmacological Properties of Maize RIP by Cysteine-Specific PEGylation

by Ka-Yee Au, Wei-Wei Shi, Shuai Qian, Zhong Zuo and Pang-Chui Shaw

Toxins 2016, 8(10), 298; https://doi.org/10.3390/toxins8100298 - 17 Oct 2016

Cited by 3 | Viewed by 4246

Abstract

To improve the pharmacological properties of maize ribosome-inactivating protein (maize RIP) for targeting HIV-infected cells, the previously engineered TAT-fused active form of maize RIP (MOD) was further engineered for cysteine-directed PEGylation. In this work, two potential antigenic sites, namely Lys-78 and Lys-264, were [...] Read more.

To improve the pharmacological properties of maize ribosome-inactivating protein (maize RIP) for targeting HIV-infected cells, the previously engineered TAT-fused active form of maize RIP (MOD) was further engineered for cysteine-directed PEGylation. In this work, two potential antigenic sites, namely Lys-78 and Lys-264, were identified. They were mutated to cysteine residue and conjugated with PEG_5k or PEG_20k. The resultant PEG derivatives of MOD variants were examined for ribosome-inactivating activity, circulating half-life and immunogenicity. Our results showed that MOD-PEG conjugates had two- to five-fold lower biological activity compared to the wild-type. Mutation of the two sites respectively did not decrease the anti-MOD IgG and IgE level in mice, but the conjugation of PEG did dramatically reduce the antigenicity. Furthermore, pharmacokinetics studies demonstrated that attachment of PEG_20k prolonged the plasma half-life by five-fold for MOD-K78C and 17-fold for MOD-K264C, respectively. The site-specific mutation together with PEGylation therefore generated MOD derivatives with improved pharmacological properties. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessArticle

Crystal Structure of Ribosome-Inactivating Protein Ricin A Chain in Complex with the C-Terminal Peptide of the Ribosomal Stalk Protein P2

by Wei-Wei Shi, Yun-Sang Tang, See-Yuen Sze, Zhen-Ning Zhu, Kam-Bo Wong and Pang-Chui Shaw

Toxins 2016, 8(10), 296; https://doi.org/10.3390/toxins8100296 - 13 Oct 2016

Cited by 31 | Viewed by 7397

Abstract

Ricin is a type 2 ribosome-inactivating protein (RIP), containing a catalytic A chain and a lectin-like B chain. It inhibits protein synthesis by depurinating the N-glycosidic bond at α-sarcin/ricin loop (SRL) of the 28S rRNA, which thereby prevents the binding of elongation factors [...] Read more.

Ricin is a type 2 ribosome-inactivating protein (RIP), containing a catalytic A chain and a lectin-like B chain. It inhibits protein synthesis by depurinating the N-glycosidic bond at α-sarcin/ricin loop (SRL) of the 28S rRNA, which thereby prevents the binding of elongation factors to the GTPase activation center of the ribosome. Here, we present the 1.6 Å crystal structure of Ricin A chain (RTA) complexed to the C-terminal peptide of the ribosomal stalk protein P2, which plays a crucial role in specific recognition of elongation factors and recruitment of eukaryote-specific RIPs to the ribosomes. Our structure reveals that the C-terminal GFGLFD motif of P2 peptide is inserted into a hydrophobic pocket of RTA, while the interaction assays demonstrate the structurally untraced SDDDM motif of P2 peptide contributes to the interaction with RTA. This interaction mode of RTA and P protein is in contrast to that with trichosanthin (TCS), Shiga-toxin (Stx) and the active form of maize RIP (MOD), implying the flexibility of the P2 peptide-RIP interaction, for the latter to gain access to ribosome. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Review

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Open AccessFeature PaperReview

Microvesicle Involvement in Shiga Toxin-Associated Infection

by Annie Villysson, Ashmita Tontanahal and Diana Karpman

Toxins 2017, 9(11), 376; https://doi.org/10.3390/toxins9110376 - 19 Nov 2017

Cited by 30 | Viewed by 10859

Abstract

Shiga toxin is the main virulence factor of enterohemorrhagic Escherichia coli, a non-invasive pathogen that releases virulence factors in the intestine, causing hemorrhagic colitis and, in severe cases, hemolytic uremic syndrome (HUS). HUS manifests with acute renal failure, hemolytic anemia and thrombocytopenia. [...] Read more.

Shiga toxin is the main virulence factor of enterohemorrhagic Escherichia coli, a non-invasive pathogen that releases virulence factors in the intestine, causing hemorrhagic colitis and, in severe cases, hemolytic uremic syndrome (HUS). HUS manifests with acute renal failure, hemolytic anemia and thrombocytopenia. Shiga toxin induces endothelial cell damage leading to platelet deposition in thrombi within the microvasculature and the development of thrombotic microangiopathy, mostly affecting the kidney. Red blood cells are destroyed in the occlusive capillary lesions. This review focuses on the importance of microvesicles shed from blood cells and their participation in the prothrombotic lesion, in hemolysis and in the transfer of toxin from the circulation into the kidney. Shiga toxin binds to blood cells and may undergo endocytosis and be released within microvesicles. Microvesicles normally contribute to intracellular communication and remove unwanted components from cells. Many microvesicles are prothrombotic as they are tissue factor- and phosphatidylserine-positive. Shiga toxin induces complement-mediated hemolysis and the release of complement-coated red blood cell-derived microvesicles. Toxin was demonstrated within blood cell-derived microvesicles that transported it to renal cells, where microvesicles were taken up and released their contents. Microvesicles are thereby involved in all cardinal aspects of Shiga toxin-associated HUS, thrombosis, hemolysis and renal failure. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessReview

The Use of Plant-Derived Ribosome Inactivating Proteins in Immunotoxin Development: Past, Present and Future Generations

by Aleksander Rust, Lynda J. Partridge, Bazbek Davletov and Guillaume M. Hautbergue

Toxins 2017, 9(11), 344; https://doi.org/10.3390/toxins9110344 - 27 Oct 2017

Cited by 24 | Viewed by 6330

Abstract

Ribosome inactivating proteins (RIPs) form a class of toxins that was identified over a century ago. They continue to fascinate scientists and the public due to their very high activity and long-term stability which might find useful applications in the therapeutic killing of [...] Read more.

Ribosome inactivating proteins (RIPs) form a class of toxins that was identified over a century ago. They continue to fascinate scientists and the public due to their very high activity and long-term stability which might find useful applications in the therapeutic killing of unwanted cells but can also be used in acts of terror. We will focus our review on the canonical plant-derived RIPs which display ribosomal RNA N-glycosidase activity and irreversibly inhibit protein synthesis by cleaving the 28S ribosomal RNA of the large 60S subunit of eukaryotic ribosomes. We will place particular emphasis on therapeutic applications and the generation of immunotoxins by coupling antibodies to RIPs in an attempt to target specific cells. Several generations of immunotoxins have been developed and we will review their optimisation as well as their use and limitations in pre-clinical and clinical trials. Finally, we endeavour to provide a perspective on potential future developments for the therapeutic use of immunotoxins. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessFeature PaperReview

Shiga Toxin—A Model for Glycolipid-Dependent and Lectin-Driven Endocytosis

by Ludger Johannes

Toxins 2017, 9(11), 340; https://doi.org/10.3390/toxins9110340 - 25 Oct 2017

Cited by 60 | Viewed by 9610

Abstract

The cellular entry of the bacterial Shiga toxin and the related verotoxins has been scrutinized in quite some detail. This is due to their importance as a threat to human health. At the same time, the study of Shiga toxin has allowed the [...] Read more.

The cellular entry of the bacterial Shiga toxin and the related verotoxins has been scrutinized in quite some detail. This is due to their importance as a threat to human health. At the same time, the study of Shiga toxin has allowed the discovery of novel molecular mechanisms that also apply to the intracellular trafficking of endogenous proteins at the plasma membrane and in the endosomal system. In this review, the individual steps that lead to Shiga toxin uptake into cells will first be presented from a purely mechanistic perspective. Membrane-biological concepts will be highlighted that are often still poorly explored, such as fluctuation force-driven clustering, clathrin-independent membrane curvature generation, friction-driven scission, and retrograde sorting on early endosomes. It will then be explored whether and how these also apply to other pathogens, pathogenic factors, and cellular proteins. The molecular nature of Shiga toxin as a carbohydrate-binding protein and that of its cellular receptor as a glycosylated raft lipid will be an underlying theme in this discussion. It will thereby be illustrated how the study of Shiga toxin has led to the proposal of the GlycoLipid-Lectin (GL-Lect) hypothesis on the generation of endocytic pits in processes of clathrin-independent endocytosis. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessReview

Plant Ribosome-Inactivating Proteins: Progesses, Challenges and Biotechnological Applications (and a Few Digressions)

by Maria Serena Fabbrini, Miku Katayama, Ikuhiko Nakase and Riccardo Vago

Toxins 2017, 9(10), 314; https://doi.org/10.3390/toxins9100314 - 12 Oct 2017

Cited by 57 | Viewed by 9731

Abstract

Plant ribosome-inactivating protein (RIP) toxins are EC3.2.2.22 N-glycosidases, found among most plant species encoded as small gene families, distributed in several tissues being endowed with defensive functions against fungal or viral infections. The two main plant RIP classes include type I (monomeric) and [...] Read more.

Plant ribosome-inactivating protein (RIP) toxins are EC3.2.2.22 N-glycosidases, found among most plant species encoded as small gene families, distributed in several tissues being endowed with defensive functions against fungal or viral infections. The two main plant RIP classes include type I (monomeric) and type II (dimeric) as the prototype ricin holotoxin from Ricinus communis that is composed of a catalytic active A chain linked via a disulphide bridge to a B-lectin domain that mediates efficient endocytosis in eukaryotic cells. Plant RIPs can recognize a universally conserved stem-loop, known as the α-sarcin/ ricin loop or SRL structure in 23S/25S/28S rRNA. By depurinating a single adenine (A4324 in 28S rat rRNA), they can irreversibly arrest protein translation and trigger cell death in the intoxicated mammalian cell. Besides their useful application as potential weapons against infected/tumor cells, ricin was also used in bio-terroristic attacks and, as such, constitutes a major concern. In this review, we aim to summarize past studies and more recent progresses made studying plant RIPs and discuss successful approaches that might help overcoming some of the bottlenecks encountered during the development of their biomedical applications. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessReview

Treatments for Pulmonary Ricin Intoxication: Current Aspects and Future Prospects

by Yoav Gal, Ohad Mazor, Reut Falach, Anita Sapoznikov, Chanoch Kronman and Tamar Sabo

Toxins 2017, 9(10), 311; https://doi.org/10.3390/toxins9100311 - 3 Oct 2017

Cited by 43 | Viewed by 7381

Abstract

Ricin, a plant-derived toxin originating from the seeds of Ricinus communis (castor beans), is one of the most lethal toxins known, particularly if inhaled. Ricin is considered a potential biological threat agent due to its high availability and ease of production. The clinical [...] Read more.

Ricin, a plant-derived toxin originating from the seeds of Ricinus communis (castor beans), is one of the most lethal toxins known, particularly if inhaled. Ricin is considered a potential biological threat agent due to its high availability and ease of production. The clinical manifestation of pulmonary ricin intoxication in animal models is closely related to acute respiratory distress syndrome (ARDS), which involves pulmonary proinflammatory cytokine upregulation, massive neutrophil infiltration and severe edema. Currently, the only post-exposure measure that is effective against pulmonary ricinosis at clinically relevant time-points following intoxication in pre-clinical studies is passive immunization with anti-ricin neutralizing antibodies. The efficacy of this antitoxin treatment depends on antibody affinity and the time of treatment initiation within a limited therapeutic time window. Small-molecule compounds that interfere directly with the toxin or inhibit its intracellular trafficking may also be beneficial against ricinosis. Another approach relies on the co-administration of antitoxin antibodies with immunomodulatory drugs, thereby neutralizing the toxin while attenuating lung injury. Immunomodulators and other pharmacological-based treatment options should be tailored according to the particular pathogenesis pathways of pulmonary ricinosis. This review focuses on the current treatment options for pulmonary ricin intoxication using anti-ricin antibodies, disease-modifying countermeasures, anti-ricin small molecules and their various combinations. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessReview

Shiga Toxin Therapeutics: Beyond Neutralization

by Gregory Hall, Shinichiro Kurosawa and Deborah J. Stearns-Kurosawa

Toxins 2017, 9(9), 291; https://doi.org/10.3390/toxins9090291 - 19 Sep 2017

Cited by 26 | Viewed by 18237

Abstract

Ribotoxic Shiga toxins are the primary cause of hemolytic uremic syndrome (HUS) in patients infected with Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC), a pathogen class responsible for epidemic outbreaks of gastrointestinal disease around the globe. HUS is a leading cause of pediatric renal [...] Read more.

Ribotoxic Shiga toxins are the primary cause of hemolytic uremic syndrome (HUS) in patients infected with Shiga toxin-producing enterohemorrhagic Escherichia coli (STEC), a pathogen class responsible for epidemic outbreaks of gastrointestinal disease around the globe. HUS is a leading cause of pediatric renal failure in otherwise healthy children, resulting in a mortality rate of 10% and a chronic morbidity rate near 25%. There are currently no available therapeutics to prevent or treat HUS in STEC patients despite decades of work elucidating the mechanisms of Shiga toxicity in sensitive cells. The preclinical development of toxin-targeted HUS therapies has been hindered by the sporadic, geographically dispersed nature of STEC outbreaks with HUS cases and the limited financial incentive for the commercial development of therapies for an acute disease with an inconsistent patient population. The following review considers potential therapeutic targeting of the downstream cellular impacts of Shiga toxicity, which include the unfolded protein response (UPR) and the ribotoxic stress response (RSR). Outcomes of the UPR and RSR are relevant to other diseases with large global incidence and prevalence rates, thus reducing barriers to the development of commercial drugs that could improve STEC and HUS patient outcomes. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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Open AccessReview

Extensive Evolution of Cereal Ribosome-Inactivating Proteins Translates into Unique Structural Features, Activation Mechanisms, and Physiological Roles

by Jeroen De Zaeytijd and Els J. M. Van Damme

Toxins 2017, 9(4), 123; https://doi.org/10.3390/toxins9040123 - 29 Mar 2017

Cited by 20 | Viewed by 6320

Abstract

Ribosome-inactivating proteins (RIPs) are a class of cytotoxic enzymes that can depurinate rRNAs thereby inhibiting protein translation. Although these proteins have also been detected in bacteria, fungi, and even some insects, they are especially prevalent in the plant kingdom. This review focuses on [...] Read more.

Ribosome-inactivating proteins (RIPs) are a class of cytotoxic enzymes that can depurinate rRNAs thereby inhibiting protein translation. Although these proteins have also been detected in bacteria, fungi, and even some insects, they are especially prevalent in the plant kingdom. This review focuses on the RIPs from cereals. Studies on the taxonomical distribution and evolution of plant RIPs suggest that cereal RIPs have evolved at an enhanced rate giving rise to a large and heterogeneous RIP gene family. Furthermore, several cereal RIP genes are characterized by a unique domain architecture and the lack of a signal peptide. This advanced evolution of cereal RIPs translates into distinct structures, activation mechanisms, and physiological roles. Several cereal RIPs are characterized by activation mechanisms that include the proteolytic removal of internal peptides from the N-glycosidase domain, a feature not documented for non-cereal RIPs. Besides their role in defense against pathogenic fungi or herbivorous insects, cereal RIPs are also involved in endogenous functions such as adaptation to abiotic stress, storage, induction of senescence, and reprogramming of the translational machinery. The unique properties of cereal RIPs are discussed in this review paper. Full article

(This article belongs to the Special Issue Ribosome Inactivating Toxins)

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