Clostridium perfringens Iota-Toxin b Induces Rapid Cell Necrosis

Nagahama, Masahiro; Umezaki, Mariko; Oda, Masataka; Kobayashi, Kazuo; Toné, Shigenobu; Suda, Taiji; Ishidoh, Kazumi; Sakùrai, Jun

doi:10.1128/iai.05677-11

Cited by 45 publications

(71 citation statements)

References 30 publications

(44 reference statements)

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“…Absorbance was read at 490 nm using an enzyme-linked immunosorbent assay plate reader. Cell viability was calculated as the mean absorbance of the toxin group divided by that of the control (17,27). In some experiments, heatinactivated beta-toxin was prepared by heating at 95°C for 10 min.…”

Section: Methodsmentioning

confidence: 99%

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The p38 MAPK and JNK Pathways Protect Host Cells against Clostridium perfringens Beta-Toxin

et al. 2013

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Clostridium perfringens beta-toxin is an important agent of necrotic enteritis and enterotoxemia. Beta-toxin is a pore-forming toxin (PFT) that causes cytotoxicity. Two mitogen-activated protein kinase ( Incubation in K؉ -free medium intensified p38 MAPK activation and cell death induced by the toxin, while incubation in K ؉ -high medium prevented those effects. While streptolysin O (SLO) reportedly activates p38 MAPK via reactive oxygen species (ROS), we showed that this pathway did not play a major role in p38 phosphorylation in beta-toxin-treated cells. Therefore, we propose that beta-toxin induces activation of the MAPK pathway to promote host cell survival.

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Section: Methodsmentioning

confidence: 99%

“…The separation of lipid rafts was carried out by flotation-centrifugation on a sucrose gradient (27,29). THP-1 cells were incubated in fresh medium containing beta-toxin at 37°C for 30 min.…”

Section: Methodsmentioning

confidence: 99%

The p38 MAPK and JNK Pathways Protect Host Cells against Clostridium perfringens Beta-Toxin

et al. 2013

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“…These cytotoxic effects are responsible for the pore formation of Ib in the plasma membrane. Our previous study also reported that Ib causes an elevation of intracellular Ca 2ϩ levels from the extracellular space during endocytosis (11). On the other hand, bacterial pore-forming protein-induced Ca 2ϩ influx and the binding of pathogenic microorganisms lead to the lysosomal exocytosis and release of the lysosomal acid sphingomyelinase (ASMase) extracellularly (12,13).…”

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confidence: 99%

“…We previously reported that the binding component (Ib) of iota-toxin causes cell necrosis in A431 and A549 cells (11). These cytotoxic effects are responsible for the pore formation of Ib in the plasma membrane.…”

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Cellular Uptake of Clostridium botulinum C2 Toxin Requires Acid Sphingomyelinase Activity

et al. 2017

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Clostridium botulinum C2 toxin consists of an enzyme component (C2I) and a binding component (C2II). Activated C2II (C2IIa) binds to a cell receptor, giving rise to lipid raft-dependent oligomerization, and it then assembles with C2I. The whole toxin complex is then endocytosed into the cytosol, resulting in the destruction of the actin cytoskeleton and cell rounding. Here, we showed that C2 toxin requires acid sphingomyelinase (ASMase) activity during internalization. In this study, inhibitors of ASMase and lysosomal exocytosis blocked C2 toxin-induced cell rounding. C2IIa induced Ca 2ϩ influx from the extracellular medium to cells. C2 toxininduced cell rounding was enhanced in the presence of Ca 2ϩ . ASMase was released extracellularly when cells were incubated with C2IIa in the presence of Ca 2ϩ . Small interfering RNA (siRNA) knockdown of ASMase reduced C2 toxin-induced cell rounding. ASMase hydrolyzes sphingomyelin to ceramide on the outer leaflet of the membrane at acidic pH. Ceramide was detected in cytoplasmic vesicles containing C2IIa. These results indicated that ASMase activity is necessary for the efficient internalization of C2 toxin into cells. Inhibitors of ASMase may confer protection against infection.

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“…Recently, we reported that Ib alone caused the death of A431 and A549 cells (21). Ib bound and formed oligomers in the membranes of A431 cells.…”

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Intracellular Trafficking of Clostridium perfringens Iota-Toxin b

et al. 2012

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c Clostridium perfringens iota-toxin is composed of an enzymatic component (Ia) and a binding component (Ib). Ib binds to a cell surface receptor, undergoes oligomerization in lipid rafts, and binds Ia. The resulting complex is then endocytosed. Here, we show the intracellular trafficking of iota-toxin. After the binding of the Ib monomer with cells at 4°C, oligomers of Ib formed at 37°C and later disappeared. Immunofluorescence staining of Ib revealed that the internalized Ib was transported to early endosomes. Some Ib was returned to the plasma membrane through recycling endosomes, whereas the rest was transported to late endosomes and lysosomes for degradation. Degraded Ib was delivered to the plasma membrane by an increase in the intracellular Ca 2؉ concentration caused by Ib. Bafilomycin A1, an endosomal acidification inhibitor, caused the accumulation of Ib in endosomes, and both nocodazole and colchicine, microtubule-disrupting agents, restricted Ib's movement in the cytosol. These results indicated that an internalized Ia and Ib complex was delivered to early endosomes and that subsequent delivery of Ia to the cytoplasm occurs mainly in early endosomes. Ib was either sent back to the plasma membranes through recycling endosomes or transported to late endosomes and lysosomes for degradation. Degraded Ib was transported to plasma membranes. C lostridium perfringens type E, which produces an iota-toxin consisting of an enzyme component (Ia) and a binding component (Ib), causes enterotoxemia in calves, lambs, and piglets (24,26,28). Ia ADP-ribosylates skeletal muscle ␣-actin and nonmuscle ␤/␥-actin (1, 3), and Ib binds to the cell, forming oligomers (15,18,30). The components lack toxic activity when each is injected alone, but together they have cytotoxic, lethal, and dermonecrotic effects (1, 24, 26). Iota-toxin belongs to a family of binary actin-ADP-ribosylating toxins that includes Clostridium botulinum C2 toxin, Clostridium spiroforme iota-like toxin, Clostridium difficile ADP-ribosyltransferase, and vegetative insecticidal protein from Bacillus cereus (1, 26).Crystallography of Ia revealed that it is divided into two domains, the N domain (residues 1 to 210), which is responsible for interaction with Ib, and the C domain (residues 211 to 413), which is involved in the catalytic activity of ADP-ribosyltransferase (25, 32). We described the structure of a Michaelis complex with iotatoxin, actin, and a nonhydrolyzable NAD ϩ analogue (33). Based on this structure, we provided some insight into substrate recognition. In particular, we were able to show that Tyr 62 on loop I and Arg 248 on loop II in Ia play an essential role at the actin-toxin interface. We also proposed a common reaction mechanism for the actin-targeting mono-ADP-ribosylating toxins whereby an oxocarbenium intermediate is formed following the cleavage of the nicotinamide moiety from NAD ϩ . Rotation then allows for the release of the conformational strain and the formation of a second cationic intermediate. Finally, the nucleophilic attac...

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Clostridium perfringens Iota-Toxin b Induces Rapid Cell Necrosis

Cited by 45 publications

References 30 publications

The p38 MAPK and JNK Pathways Protect Host Cells against Clostridium perfringens Beta-Toxin

The p38 MAPK and JNK Pathways Protect Host Cells against Clostridium perfringens Beta-Toxin

Cellular Uptake of Clostridium botulinum C2 Toxin Requires Acid Sphingomyelinase Activity

Intracellular Trafficking of Clostridium perfringens Iota-Toxin b

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