Diamide insecticides target insect ryanodine receptors (RyRs) and cause misregulation of calcium signaling in insect muscles and neurons, generating worldwide sales over 2 billion U.S. dollars annually. Several resistance mutations have been reported to reduce the efficacy of the diamides, but the exact binding sites and mechanism of resistance mutations are not clear. The recent breakthrough in the structural studies of mammalian RyRs has deepened our understanding of the channel, but the structural information about insect RyRs is still scarce. Recently we solved the cryo-electron microscopy (cryo-EM) structure of RyR in complex with the anthranilic diamide chlorantraniliprole (CHL). CHL binds to the pseudo-voltage-sensor domain (pVSD) of RyR, a site in proximity to the previously identified resistance mutations. Mutagenesis studies in silico, in mutant cell lines, and in transgenic drosophila strains reveal the key residues involved in diamide coordination and the molecular mechanism under species-selectivity and resistance mutations. We have also solved the crystal structures of several RyR domains from the diamondback moth and the bee, revealing insect-specific structural features which could be potentially targeted by novel insecticides. Interestingly, we found that the phosphorylation of insect RyR is temperature-dependent, facilitated by the low thermal stability and dynamic structure of the insect RyR. Our structures provide a foundation for developing novel pesticides to overcome the resistance crisis.


 

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