Evidence of a shared binding site for Bacillus thuringiensis Cry1Ac and Cry2Aa toxins in Cnaphalocrocis medinalis cadherin

Insect midgut cadherins function as receptors and play critical roles as protein receptors of insecticidal Bacillus thuringiensis (Bt) toxins used as biopesticides and in Bt transgenic crops worldwide. Here, we cloned and characterized the full‐length midgut cadherin (CmCad) cDNA from the rice leaffolder (Cnaphalocrocis medinalis), a destructive pest of rice in many Asian countries. Expression of recombinant proteins corresponding to the extracellular domain of CmCad allowed testing binding of Cry proteins. Results from in vitro ligand blotting and enzyme‐linked immunosorbent assays supported that the extracellular domain of CmCad contains regions recognized by both Cry1Ac and Cry2Aa. Molecular modelling and docking simulations indicated that binding to both Cry1Ac and Cry2Aa is localized primarily within a CmCad motif corresponding to residues T1417–D1435. A recombinant CmCad protein produced without residues T1417–D1435 lacked binding to Cry1Ac and Cry2Aa, confirmed our modelling predictions that CmCad has a shared Cry1Ac and Cry2Aa binding site. The potential existence of a shared binding region in CmCad suggests that caution should be taken when using combinations of Cry1Ac and Cry2Aa in pyramided transgenic rice, as their combined use could speed the evolution of resistance to both toxins. The truncated CR9‐11 peptide from C. medinalis cadherin showed the binding ability to Cry1Ac and Cry2Aa. Molecular docking identified a conserved cadherin binding motif (T1417–D1435) as predicted to be involved in these binding. Lack of Cry1Ac and Cry2Aa binding to a mutant CmCad‐CR9‐11 region with a deletion of the motif provided further evidence supporting this motif as the binding site shared by Cry1Ac and Cry2Aa.