Bacterial toxins modifying the actin cytoskeleton

Numerous bacterial toxins recognize the actin cytoskeleton as a target. The clostridial binary toxins (Iota and C2 families) ADP-ribosylate the actin monomers causing the dissociation of the actin filaments. The large clostridial toxins from Clostridium difficile, Clostridium sordellii and Clostridium novyi inactivate, by glucosylation, proteins from the Rho family that regulate actin polymerization. In contrast, the cytotoxic necrotic factor from Escherichia coli activates Rho by deamidation and increases the formation of actin filaments. The enterotoxin of Bacteroides fragilis is a protease specific for E-cadherin and it promotes the reorganization of the actin cytoskeleton. The bacterial toxins that modify the actin cytoskeleton induce various cell disfunctions including changes in cell barrier permeability and disruption of intercellular junctions

Comparative genome and phenotypic analysis of Clostridium difficile 027 strains provides insight into the evolution of a hypervirulent bacterium

A genome comparison of non-epidemic and epidemic strains of Clostridium difficile reveals gene gains that could explain how a hypervirulent strain has emerge

CD44 Promotes Intoxication by the Clostridial Iota-Family Toxins

International audienceVarious pathogenic clostridia produce binary protein toxins associated with enteric diseases of humans and animals. Separate binding/translocation (B) components bind to a protein receptor on the cell surface, assemble with enzymatic (A) component(s), and mediate endocytosis of the toxin complex. Ultimately there is translocation of A component(s) from acidified endosomes into the cytosol, leading to destruction of the actin cytoskeleton. Our results revealed that CD44, a multifunctional surface protein of mammalian cells, facilitates intoxication by the iota family of clostridial binary toxins. Specific antibody against CD44 inhibited cytotoxicity of the prototypical Clostridium perfringens iota toxin. Versus CD44(+) melanoma cells, those lacking CD44 bound less toxin and were dose-dependently resistant to C. perfringens iota, as well as Clostridium difficile and Clostridium spiroforme iota-like, toxins. Purified CD44 specifically interacted in vitro with iota and iota-like, but not related Clostridium botulinum C2, toxins. Furthermore, CD44 knockout mice were resistant to iota toxin lethality. Collective data reveal an important role for CD44 during intoxication by a family of clostridial binary toxins

Structural insights into Clostridium perfringens delta toxin pore formation

Clostridium perfringens Delta toxin is one of the three hemolysin-like proteins produced by C. perfringens type C and possibly type B strains. One of the others, NetB, has been shown to be the major cause of Avian Nectrotic Enteritis, which following the reduction in use of antibiotics as growth promoters, has become an emerging disease of industrial poultry. Delta toxin itself is cytotoxic to the wide range of human and animal macrophages and platelets that present GM2 ganglioside on their membranes. It has sequence similarity with Staphylococcus aureus β-pore forming toxins and is expected to heptamerize and form pores in the lipid bilayer of host cell membranes. Nevertheless, its exact mode of action remains undetermined. Here we report the 2.4 Å crystal structure of monomeric Delta toxin. The superposition of this structure with the structure of the phospholipid-bound F component of S. aureus leucocidin (LukF) revealed that the glycerol molecules bound to Delta toxin and the phospholipids in LukF are accommodated in the same hydrophobic clefts, corresponding to where the toxin is expected to latch onto the membrane, though the binding sites show significant differences. From structure-based sequence alignment with the known structure of staphylococcal α-hemolysin, a model of the Delta toxin pore form has been built. Using electron microscopy, we have validated our model and characterized the Delta toxin pore on liposomes. These results highlight both similarities and differences in the mechanism of Delta toxin (and by extension NetB) cytotoxicity from that of the staphylococcal pore-forming toxins

Clostridium perfringens Epsilon Toxin Induces a Rapid Change of Cell Membrane Permeability to Ions and Forms Channels in Artificial Lipid Bilayers

International audienceEpsilon toxin is a potent toxin produced by Clostri-dium perfringens types B and D, which are responsible for a rapidly fatal enterotoxemia in animals. One of the main properties of epsilon toxin is the production of edema. We have previously found that epsilon toxin causes a rapid swelling of Madin-Darby canine kidney cells and that the toxin does not enter the cytosol and remains associated with the cell membrane by forming a large complex (Petit, L., Gibert, M., Gillet, D., Laurent-Winter, C., Boquet, P., and Popoff, M. R. (1997) J. Bacte-riol. 179, 6480 – 6487). Here, we report that epsilon toxin induced in Madin-Darby canine kidney cells a rapid decrease of intracellular K , and an increase of Cl and Na , whereas the increase of Ca 2 occurred later. The entry of propidium iodide that was correlated with the loss of cell viability monitored by the 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test indicates that epsilon toxin formed large pores. In artificial lipid bilayers, epsilon toxin caused current steps with a single-channel conductance of 60 pS in 100 mM KCl, which represented general diffusion pores. The channels were slightly selective for anions, but cations could also penetrate. Epsilon toxin formed wide and water-filled channels permeable to hydrophilic solutes up to a molecular mass of at least 1 kDa, which probably represents the basic mechanism of toxin action on target cells

Endocytosis and toxicity of clostridial binary toxins depend on a clathrin-independent pathway regulated by Rho-GDI

International audienceClostridial binary toxins, such as Clostridium perfringens Iota and Clostridium botulinum C2, are composed of a binding protein (Ib and C2II respectively) that recognizes distinct membrane receptors and mediates internalization of a catalytic protein (Ia and C2-I respectively) with ADP-ribosyltransferase activity that disrupts the actin cytoskeleton. We show here that the endocytic pathway followed by these toxins is independent of clathrin but requires the activity of dynamin and is regulated by Rho-GDI. This endocytic pathway is similar to a recently characterized clathrin-independent pathway followed by the interleukin-2 (IL2) receptor. We found indeed that Ib and C2II colocalized intracellularly with the IL2 receptor but not the transferrin receptor after different times of endocytosis. Accordingly, the intracellular effects of Iota and C2 on the cytoskeleton were inhibited by inactivation of dynamin or by Rho-GDI whereas inhibitors of clathrin-dependent endocytosis had no protective effect