title: Towards Coleoptera-specific high-throughput screening systems for compounds with ecdysone activity: development of EcR reporter assays using weevil (Anthonomus grandis)-derived cell lines and in silico analysis of ligand binding to A. grandis EcR ligand-binding pocket.
abstract: Molting in insects is regulated by ecdysteroids and juvenile hormones. Several synthetic non-steroidal ecdysone agonists are on the market as insecticides. These ecdysone agonists are dibenzoylhydrazine (DBH) analogue compounds that manifest their toxicity via interaction with the ecdysone receptor (EcR). Of the four commercial available ecdysone agonists, three (tebufenozide, methoxyfenozide and chromafenozide) are highly lepidopteran specific, one (halofenozide) is used to control coleopteran and lepidopteran insects in turf and ornamentals. However, compared to the very high binding affinity of these DBH analogues to lepidopteran EcRs, halofenozide has a low binding affinity for coleopteran EcRs. For the discovery of ecdysone agonists that target non-lepidopteran insect groups, efficient screening systems that are based on the activation of the EcR are needed. We report here the development and evaluation of two coleopteran-specific reporter-based screening systems to discover and evaluate ecdysone agonists. The screening systems are based on the cell lines BRL-AG-3A and BRL-AG-3C that are derived from the weevil Anthonomus grandis, which can be efficiently transduced with an EcR reporter cassette for evaluation of induction of reporter activity by ecdysone agonists. We also cloned the almost full length coding sequence of EcR expressed in the cell line BRL-AG-3C and used it to make an initial in silico 3D-model of its ligand-binding pocket docked with ponasterone A and tebufenozide.
authors: Laboratory of Agrozoology, Department of Crop Protection, Faculty of Bioscience Engineering, Ghent University, Coupure Links 653, Ghent, Belgium.
authors: Thomas
authors: T
authors: Soin
authors: Masatoshi
authors: M
authors: Iga
authors: Luc
authors: L
authors: Swevers
authors: Pierre
authors: P
authors: Rougé
authors: Colin R
authors: CR
authors: Janssen
authors: Guy
authors: G
authors: Smagghe
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D001517:Coleoptera / Q000145:classification / Q000187:drug effects* / Q000235:genetics; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004440:Ecdysone / Q000819:agonists* / Q000378:metabolism; D017930:Genes, Reporter / Q000187:drug effects; D006834:Hydrazines / Q000494:pharmacology*; D019476:Insect Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007306:Insecticides / Q000494:pharmacology; D008024:Ligands; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D010802:Phylogeny; D011485:Protein Binding / Q000187:drug effects; D011987:Receptors, Steroid / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D048889:Weevils / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism
chemical_list: D006834:Hydrazines; D019476:Insect Proteins; D007306:Insecticides; D008024:Ligands; D011987:Receptors, Steroid; C055469:ecdysone receptor; D004440:Ecdysone