title: Stable expression and functional characterisation of the diamondback moth ryanodine receptor G4946E variant conferring resistance to diamide insecticides.
abstract: Diamides, such as flubendiamide and chlorantraniliprole, belong to a new chemical class of insecticides that act as conformation-sensitive activators of insect ryanodine receptors (RyRs). Both compounds are registered for use against lepidopteran species such as the diamondback moth, Plutella xylostella, a notorious global pest of cruciferous crops. Recently acquired resistance to diamide insecticides in this species is thought to be due to a target-site mutation conferring an amino acid substitution (G4946E), located within the trans-membrane domain of the RyR, though the exact role of this mutation has not yet been fully determined. To address this we have cloned a full-length cDNA encoding the P. xylostella RyR and established clonal Sf9 cell lines stably expressing either the wildtype RyR or the G4946E variant, in order to test the sensitivity to flubendiamide and chlorantraniliprole on the recombinant receptor. We report that the efficacy of both diamides was dramatically reduced in clonal Sf9 cells stably expressing the G4946E modified RyR, providing clear functional evidence that the G4946E RyR mutation impairs diamide insecticide binding.
authors.affiliation: Biological Chemistry and Crop Protection Department, Rothamsted Research, Harpenden, Hertfordshire, AL5 2JQ, UK.
authors.forename: Bartlomiej J
authors.initials: BJ
authors.lastname: Troczka
authors.affiliation: Institute of Molecular &Experimental Medicine, Cardiff University School of Medicine, Wales Heart Research Institute, Heath Park, Cardiff CF14 4XN, UK.
authors.forename: Alan J
authors.initials: AJ
authors.lastname: Williams
authors.affiliation: Biological Chemistry and Crop Protection Department, Rothamsted Research, Harpenden, Hertfordshire, AL5 2JQ, UK.
authors.forename: Martin S
authors.initials: MS
authors.lastname: Williamson
authors.affiliation: Biological Chemistry and Crop Protection Department, Rothamsted Research, Harpenden, Hertfordshire, AL5 2JQ, UK.
authors.forename: Linda M
authors.initials: LM
authors.lastname: Field
authors.affiliation: Bayer CropScience AG, 65926 Frankfurt, Germany.
authors.forename: Peter
authors.initials: P
authors.lastname: Lüemmen
authors.affiliation: Biological Chemistry and Crop Protection Department, Rothamsted Research, Harpenden, Hertfordshire, AL5 2JQ, UK.
authors.forename: T G Emyr
authors.initials: TG
authors.lastname: Davies
mesh_terms: D000818:Animals; D001549:Benzamides / Q000378:metabolism / Q000494:pharmacology; D002110:Caffeine / Q000378:metabolism / Q000494:pharmacology; D020013:Calcium Signaling; D003001:Cloning, Molecular; D004361:Drug Tolerance; D015870:Gene Expression; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism*; D007306:Insecticides / Q000378:metabolism / Q000494:pharmacology*; D020125:Mutation, Missense; D011485:Protein Binding; D012433:Ryanodine / Q000378:metabolism / Q000494:pharmacology; D019837:Ryanodine Receptor Calcium Release Channel / Q000235:genetics / Q000378:metabolism*; D061987:Sf9 Cells; D018411:Spodoptera; D013450:Sulfones / Q000378:metabolism / Q000494:pharmacology; D062367:ortho-Aminobenzoates / Q000378:metabolism / Q000494:pharmacology; D005466:Fluorocarbons; D010797:Phthalimides
chemical_list: D001549:Benzamides; D002110:Caffeine; D019476:Insect Proteins; D007306:Insecticides; D012433:Ryanodine; D019837:Ryanodine Receptor Calcium Release Channel; D013450:Sulfones; D062367:ortho-Aminobenzoates; C517733:chlorantranilipole; C508762:flubendiamide; D005466:Fluorocarbons; D010797:Phthalimides