Carbohydrate moieties play an important role in binding and toxicity of Bt toxin Cry1Ac in the midgut of cabbage loopers

Cry proteins from the soil bacterium Bacillus thuringiensis (Bt) are major insecticidal toxins in formulated Bt-sprays and in transgenic Bt-crops widely used in agriculture. In order to understand the modes of action of Cry toxins and mechanisms of insect resistance to Bt toxins, it is important to understand the specific interaction of Cry toxins with the insect midgut brush border membranes. We have previously found that the fluorescent brightener Calcofluor white (CFW) could inhibit the formation of the peritrophic membrane (PM) in larval midgut and significantly inhibit the toxicity of Cry1Ac in the cabbage looper, Trichoplusia ni. In this study, we used CFW as a unique reagent to study the specific interaction of Cry1Ac with the larval midgut in the mode of action of the toxin. T. ni larvae and midgut samples were treated with CFW in vivo or in vitro. Cry1Ac toxicity in larvae was determined, larval midgut structure was examined and proteins were analyzed by MS/MS proteomic analysis, and binding of Cry1Ac to the midgut brush border membranes was analyzed. Our results indicated that carbohydrate moieties in the midgut play an important role in binding and toxicity of Cry1Ac in T. ni larvae.