Crystal Structures of Insect Ryanodine Receptor Domains

Abstract

Diamide insecticides are among the top-selling pesticides, generating worldwide sales over 2 billion U.S. dollars annually. They target insect ryanodine receptors (RyRs) and cause misregulation of calcium signaling in insect muscles and neurons. However, due to heavy usage, several resistance mutations identified in insect RyRs have been reported to reduce the efficacy of the diamides, but the exact binding site of diamides and the mechanism of resistance mutations remain elusive. Two options are available to overcome the resistance: modify the diamide insecticide to target the same binding pocket in resistant RyRs or develop new compounds to target a different binding site in insect RyRs. 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. Here we report four crystal structures of insect RyR domains, including N-terminal domain (NTD), SPRY2 domain, and repeat34 domain from diamondback moth (DBM), and NTD from the honeybee. Several regions in these domains showed distinguished conformations in DBM relative to honeybee and mammalian RyRs. Many of these pest-specific structural features are located in the domain-domain interfaces that would change conformation upon channel gating, making them good target for candidate insecticides. We identified several protein kinase a (PKA) phosphorylation sites clustering in a loop and a newly identified α-helix in DBM repeat34 domain, showing a pest-specific phosphorylation pattern. Interestingly, the phosphorylation of insect RyR is temperature-dependent, facilitated by the low thermal stability and dynamic structure of insect Repeat34 domain. The accurate structural information of insect RyRs would provide valuable templates for the design of insecticidal molecules.