CONICET | Buscador de Institutos y Recursos Humanos

Poisonous eggs from Pomacea snails have evolved a defensive protein that damage cell membranes with a pore-forming toxin delivered by a lectin We found that the eggs from the invasive freshwater snail Pomacea maculata have evolved a defensive strategy against predation involving a neurotoxin, PmPV2. PmPV2 is a dimeric pore-forming toxin which combines two immune proteins: a lectin and a membrane attack complex/perforin (MACPF) module. The aim of this study was to investigate PmPV2 interaction with membranes using enterocytes as a model of the first cells that the toxin would encounter when ingested by a predator. The toxin withstands digestive proteases in vitro and can attach to mice enterocyteglycocalyx in vivo, suggesting it reaches the intestinal tract in an active form. At the cellular level, PmPV2 specifically interacts with membrane cholesterol (MACPF) and its lectin module interacts with glycoproteins. It has affinity to negatively charged glycans as revealed by a glycan array and hemagglutination assays. Though Atomic Force Microscopy we observed that PmPV2inserts into phosphoplipid/cholesterol lipid bilayers and oligomerizes into pore-like structures. Additionally, PmPV2 binds to the plasma membrane of intestinal Caco-2 cells recognizing specific glycoproteins, alters their surface morphology and eventually leads to cell death. Intestinal cells exposed to the toxin changed their membrane permeability in patch-clamp experiments in a fashion compatible with the formation of ~14nm pores. This effect is inhibitedby aminated monosaccharides, highlighting the importance of the lectin module. The study provides the first evidence to our knowledge of the molecular mechanism of pore formation for an animal defensive toxin containing a MACPF/CDC domain.