ADP‐ribosylation of skeletal muscle and non‐muscle actin by Clostridium perfringens iota toxin

Abstract

The enzymatically active component i, of Clostridium perfringens iota toxin ADP-ribosylated actin in human platelet cytosol and purified platelet Ply-actin, in a similar way to that been reported for component I of botulinum C2 toxin. ADP-ribosylation of cytosolic and purified actin by either toxin was inhibited by 0.1 mM phalloidin indicating that monomeric G-actin but not polymerized F-actin was the toxin substrate. Perfringens iota toxin and botulinum C2 toxin were not additive in ADP-ribosylation of platelet actin. Treatment of intact chicken embryo cells with botulinum C2 toxin decreased subsequent ADP-ribosylation of actin in cell lysates by perfringens iota or botulinum C2 toxin. In contrast to botulinum C2 toxin, perfringens iota toxin ADP-ribosylated skeletal muscle or-actin with a potency and efficiency similar to non-muscle actin. ADP-ribosylation of purified skeletal muscle and non-muscle actin by perfringens iota toxin led to a dose-dependent impairment of the ability of actin to polymerize.Several bacterial toxins modify functions in eukaryotic cells by an ADP-ribosylation of intracellular target proteins. Diphtheria toxin and Pseudomonas exotoxin A ADPribosylate elongation factor I1 and thereby inhibit protein synthesis [l]. Cholera toxin and pertussis toxin modify guanylnucleotide-binding regulatory (G) proteins involved in trans-Botulinum C2 toxin is another member of the family of microbial ADP-ribosyltransferases [5]. It has been shown that botulinum C2 toxin modifies actin in intact cells and isolated cytoplasmic /?/y actin in its monomeric G form, but not F-actin [6 -91. The covalent modification of purified actin by botulinum C2 toxin severely reduces its ability to polymerize Recently it has been demonstrated that perfringens iota toxin also possesses ADP-ribosyltransferase activity [lo]. Furthermore, it has been reported in a preliminary note that it also accepts actin as substrate [lo]. We were prompted to compare the molecular mode of action of these two toxins because of their striking similarities. Both toxins are clostridial in origin and are produced during opportunistic intestinal infections. They both cause increased fluid transport across membranes [ll, 121. They both are binary in structure and consist of a high and low-molecular-mass component, which are components I and I1 of botulinum C2 toxin and components i, and ib of perfringens iota toxin. The respective components are linked neither by covalent nor non-covalent bonds [13-151. In each case the smaller components of botulinum C2 toxin (C21, molecular mass 50 kDa) and of perfringens iota toxin (i,, molecular mass 48 kDa) are enzymatically active as ADP-ribosyltransferases [5,6, lo]. The larger components have a molecular mass of 72 kDa in iota toxin [14] and 88 kDa in trypsin-activated C2 toxin [16] and are respectively responsible for the binding of the toxin to the eukaryotic cell surface. Thus, to elicit toxic effects, both components of either toxin have to act together [13, 151. Here we report on studies on the ADP-r...

ADP‐ribosylation of actin causes increase in the rate of ATP exchange and inhibition of ATP hydrolysis

“…ADP-ribosylation of actin catalyzed by perfringens iota and botulinum C2 toxin has been shown to inhibit its ability to polymerize [l, 5,7,111. Here we report that the ADPribosylation increases the rate of exchange of ATP bound to G-actin about twofold without changes in the rank order of affinity of actin for nucleotides (ATP > ATP[yS] > ADP > AMP = NAD).…”

“…On the other hand, ADP-ribosylation of actin increases the rate of ATP exchange, inhibits ATPase activity and blocks polymerization. Finally, polymerization shields actin against ADP-ribosylation by the toxins [7].…”

“…Both toxins differ in their substrate specificity. Whereas botulinum C2 toxin modifies non-muscle actin and (much less) skeletal muscle actin, perfringens iota toxin ADP-ribosylates both actin species readily [7]. Recently it has been shown that both toxins ADP-ribosylate actin at Arg-177 [9, 101, a modification which largely reduces the ability of actin to polymerize [5, 7, 1 I].…”

Nonmuscle actin ADP‐ribosylated by botulinum C2 toxin caps actin filaments

“…In addition to botulinum C2 toxin, other microbial toxins such as Perfringens iota toxin [16], C. spiroforme toxin [21] and an ADPribosyltransferase produced by C. difficile [22] belong to the novel class of actin-modifying ADP-ribosyltransferases. Recent reports indicate that these agents most likely modify actin at an identical amino acid [21][22][23][24].…”

Analysis of the catalytic site of the actin ADP‐ribosylating Clostridium perfringens iota toxin

“…Clostridium perfringens iota toxin belongs to a family of actin-ADP-ribosylating toxins [1]. Other members of this family are C. botulinum C2 toxin [2] and C. spiroforme toxin [3,4] (for review see [5,6]).…”