Of the seven known neurotoxins expressed by Clostridium botulinum, BoNT/A, B, and E are the most common causes of botulism in humans and the major choices for both bioterrorist and pharmaceutical uses. In previous work, researchers identified several cellular proteins that can bind to these toxins, but there were conflicting data about which, if any, of the candidate receptors actually mediate cellular entry. Once inside neurons, the toxins act as proteases to block exocytosis, ultimately leading to paralysis and death.
Using both loss-of-function and gain-of-function approaches, Dong et al. show that synaptotagmin I or synaptotagmin II can act as a receptor to internalize BoNT/B into PC-12 cells. Fusion of synaptic vesicles with the plasma membrane, and thus display of vesicle proteins on the cell surface, occurs during excitation. Indeed, BoNT internalization is dependent on electrical activity in two different cell types. Fragments of synaptotagmin II effectively block the binding of BoNT/B to cultured cells and inhibit the activity of the toxin in mice, providing strong evidence that the interaction is biologically relevant.
In the mouse experiments, an injection of synaptotagmin II fragments partially protected the animals against a subsequent challenge with BoNT/B. This is the first demonstration that a fragment from a bacterial toxin receptor can antagonize the toxin in animals. The authors are now defining the precise requirements for BoNT/B–synaptotagmin II interactions, and are also trying to identify the receptors for other Clostridium neurotoxins. ▪