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Loops D, E and G in the Drosophila D?1 subunit contribute to high neonicotinoid sensitivity of D?1-chicken ?2 nicotinic acetylcholine receptor.

ABSTRACT: BACKGROUND AND PURPOSE:Neonicotinoid insecticides interact with the orthosteric site formed at subunit interfaces of insect nicotinic ACh (nACh) receptors. However, their interactions with the orthosteric sites at ?-non ? and ?-? subunit interfaces remain poorly understood. The aim of this study was to elucidate the mechanism of neonicotinoid actions using the Drosophila D?1-chicken ?2 hybrid nACh receptor. EXPERIMENTAL APPROACH:Computer models of the (D?1)3 (?2)2 nACh receptor in complex with imidacloprid and thiacloprid were generated. Amino acids in the D?1 subunit were mutated to corresponding amino acids in the human ?4 subunit to examine their effects on the agonist actions of neonicotinoids on (D?1)3 (?2)2 and (D?1)2 (?2)3 nACh receptors expressed in Xenopus laevis oocytes using voltage-clamp electrophysiology. KEY RESULTS:The (D?1)3 (?2)2 nACh receptor models indicated that amino acids in loops D, E and G probably determine the effects of neonicotinoids. The amino acid mutations tested had minimal effects on the EC50 for ACh. However, the R57S mutation in loop G, although having minimal effect on imidacloprid's actions, reduced the affinity of thiacloprid for the (D?1)3 (?2)2 nACh receptor, while scarcely affecting thiacloprid's action on the (D?1)2 (?2)3 nACh receptor. Both the K140T and the combined R57S;K140T mutations reduced neonicotinoid efficacy but only for the (D?1)3 (?2)2 nACh receptor. Combining the E78K mutation with the R57S;K140T mutations resulted in a selective reduction of thiacloprid's affinity for the (D?1)3 (?2)2 nACh receptor. CONCLUSIONS AND IMPLICATIONS:These findings suggest that a triangle of residues from loops D, E and G contribute to the selective actions of neonicotinoids on insect-vertebrate hybrid nACh receptors. LINKED ARTICLES:This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc.

SUBMITTER: Ihara M

PROVIDER: S-EPMC5979822 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Loops D, E and G in the Drosophila Dα1 subunit contribute to high neonicotinoid sensitivity of Dα1-chicken β2 nicotinic acetylcholine receptor.

Ihara Makoto M Hikida Mai M Matsushita Hiroyuki H Yamanaka Kyosuke K Kishimoto Yuya Y Kubo Kazuki K Watanabe Shun S Sakamoto Mifumi M Matsui Koutaro K Yamaguchi Akihiro A Okuhara Daiki D Furutani Shogo S Sattelle David B DB Matsuda Kazuhiko K

British journal of pharmacology 20170824 11

<h4>Background and purpose</h4>Neonicotinoid insecticides interact with the orthosteric site formed at subunit interfaces of insect nicotinic ACh (nACh) receptors. However, their interactions with the orthosteric sites at α-non α and α-α subunit interfaces remain poorly understood. The aim of this study was to elucidate the mechanism of neonicotinoid actions using the Drosophila Dα1-chicken β2 hybrid nACh receptor.<h4>Experimental approach</h4>Computer models of the (Dα1)<sub>3</sub> (β2)<sub>2< ...[more]

PMID: 28616862

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Project description:Myzus persicae is a globally important aphid pest with a history of developing resistance to insecticides. Unusually, neonicotinoids have remained highly effective as control agents despite nearly two decades of steadily increasing use. In this study, a clone of M. persicae collected from southern France was found, for the first time, to exhibit sufficiently strong resistance to result in loss of the field effectiveness of neonicotinoids.Bioassays, metabolism and gene expression studies implied the presence of two resistance mechanisms in the resistant clone, one based on enhanced detoxification by cytochrome P450 monooxygenases, and another unaffected by a synergist that inhibits detoxifying enzymes. Binding of radiolabeled imidacloprid (a neonicotinoid) to whole body membrane preparations showed that the high affinity [3H]-imidacloprid binding site present in susceptible M. persicae is lost in the resistant clone and the remaining lower affinity site is altered compared to susceptible clones. This confers a significant overall reduction in binding affinity to the neonicotinoid target: the nicotinic acetylcholine receptor (nAChR). Comparison of the nucleotide sequence of six nAChR subunit (Mp?1-5 and Mp?1) genes from resistant and susceptible aphid clones revealed a single point mutation in the loop D region of the nAChR ?1 subunit of the resistant clone, causing an arginine to threonine substitution (R81T).Previous studies have shown that the amino acid at this position within loop D is a key determinant of neonicotinoid binding to nAChRs and this amino acid change confers a vertebrate-like character to the insect nAChR receptor and results in reduced sensitivity to neonicotinoids. The discovery of the mutation at this position and its association with the reduced affinity of the nAChR for imidacloprid is the first example of field-evolved target-site resistance to neonicotinoid insecticides and also provides further validation of exisiting models of neonicotinoid binding and selectivity for insect nAChRs.

Dataset Information

Loops D, E and G in the Drosophila D?1 subunit contribute to high neonicotinoid sensitivity of D?1-chicken ?2 nicotinic acetylcholine receptor.

Publications

Loops D, E and G in the Drosophila Dα1 subunit contribute to high neonicotinoid sensitivity of Dα1-chicken β2 nicotinic acetylcholine receptor.

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