Engineered Bt Cry toxins for enhanced cotton boll weevil control

Overview

This research program focuses on engineering modified Bacillus thuringiensis (Bt) Cry toxins with enhanced binding affinity to the insect gut, aimed at achieving higher toxicity against the cotton boll weevil (Anthonomus grandis), a major pest of cotton in Argentina. While Bt proteins have been proposed for generating transgenic cotton resistant to this pest, no native Bt proteins with high toxicity to A. grandis have been identified. The project applies directed evolution strategies to modify toxin loops that interact with gut receptors, incorporating gut-binding peptides selected through phage display to increase binding specificity and toxicity.

Technical specifications

Research approach:

• Directed evolution of Bt Cry toxins (Cry1I, Cry10, Cry8) to increase gut-binding capability
• Ex vivo processing of Cry proteins with larval gut juice to assess binding ability
• Biopanning of a phage-displayed peptide library to select peptides with binding specificity for the boll weevil gut
• Incorporation of selected peptide coding sequences into toxin gene regions encoding receptor-interacting loops
• Generation of mutant cry genes and expression of modified Cry proteins
• Bioassays to assess toxicity of engineered toxins against A. grandis larvae

Scientific basis:

The strategy builds on prior work demonstrating that retargeting Bt toxins through loop replacements with gut-binding peptides can enhance toxicity, as shown with Cyt2Aa against hemipteran pests and Cry1Ab against the brown planthopper.

Technology readiness level

The research is at an early-to-mid stage of development. Initial validation steps include assessment of basal toxicity of native Bt proteins against A. grandis larvae, followed by protein engineering and bioassay validation. The approach has been validated in analogous systems for other insect pests, supporting its feasibility for application against the cotton boll weevil. The project is positioned for collaboration with partners interested in co-development of novel insect resistance traits for transgenic cotton.