Pore-forming protein complexes from Pleurotus mushrooms kill western corn rootworm and Colorado potato beetle through targeting membrane ceramide phosphoethanolamine

Aegerolysins ostreolysin A (O1yA) and pleurotolysin A (PIyA), and pleurotolysin B (PIyB) with the membrane-attack-complex/perforin domain are proteins from the mushroom genus Pleurotus. Upon binding to sphingomyelin/cholesterol-enriched membranes, OlyA and PIyA can recruit PIyB to form multimeric bi-component transmembrane pores. Recently, Pleurotus aegerolysins OlyA, PIyA2 and erylysin A (EryA) were demonstrated to preferentially bind to artificial lipid membranes containing 50 mol% ceramide phosphoethanolamine (CPE), the main sphingolipid in invertebrate cell membranes. In this study, we demonstrate that OlyA6, PIyA2 and EryA bind to insect cells and to artificial lipid membranes with physiologically relevant CPE concentrations. Moreover, these aegerolysins permeabilize these membranes when combined with PIyB. These aegerolysin/PIyB complexes show selective toxicity toward western corn rootworm larvae and adults and Colorado potato beetle larvae. These data strongly suggest that these aegerolysin/PIyB complexes recognize CPE as their receptor molecule in the insect midgut. This mode of binding is different from those described for similar aegerolysin-based bacterial complexes, or other Bacillus thuringiensis Cry toxins, which have protein receptors. Targeting of Pleurotus aegerolysins to CPE and formation of transmembrane pores in concert with PIyB suggest the use of aegerolysin/PIyB complexes as novel biopesticides for the control of western corn rootworm and Colorado potato beetle.