Heteromeric co‐assembly of two insect nicotinic acetylcholine receptor α subunits: influence on sensitivity to neonicotinoid insecticides

Abstract

Neonicotinoid insecticides, such as imidacloprid, are selective agonists of insect nicotinic acetylcholine receptors (nAChRs) and are used extensively in areas of crop protection and animal health to control a variety of insect pest species. Here, we describe studies performed with nAChR subunits Nlα1 and Nlα2 cloned from the brown planthopper Nilaparvata lugens, a major insect pest of rice crops in many parts of Asia. The influence of Nlα1 and Nlα2 subunits upon the functional properties of recombinant nAChRs has been examined by expression in Xenopus oocytes. In addition, the influence of a Nlα1 mutation (Y151S), which has been linked to neonicotinoid lab generated resistance in N. lugens, has been examined. As in previous studies of insect α subunits, functional expression has been achieved by co‐expression with the mammalian β2 subunit. This approach has revealed a significantly higher apparent affinity of imidacloprid for Nlα1/β2 than for Nlα2/β2 nAChRs. In addition, evidence has been obtained for the co‐assembly of Nlα1 and Nlα2 subunits into ‘triplet’ nAChRs of subunit composition Nlα1/Nlα2/β2. Evidence has also been obtained which demonstrates that the resistance‐associated Y151S mutation has a significantly reduced effect on neonicotinoid agonist activity when Nlα1 is co‐assembled with Nlα2 than when expressed as the sole α subunit in a heteromeric nAChR. These findings may be of importance in assessing the likely impact of the target‐site mutations such as Y151S upon neonicotinoid insecticide resistance in insect field populations.

Differential sensitivity of Ctenocephalides felis and Drosophila melanogaster nicotinic acetylcholine receptor α1 and α2 subunits in recombinant hybrid receptors to nicotinoids and neonicotinoid insecticides

“…Radioligand binding experiments on diverse Myzus persicae a (Mpa) subunits coexpressed with rat b2 subunit in D. melanogaster S2 cells showed high affinity of [ 3 H]imidacloprid for the rat b2 combinations with Mpa2 and Mpa3, but not Mpa1 and Mpa4 (Huang et al, 1999). Similar radioligand binding studies in S2 cell as well as Xenopus oocyte electrophysiology studies of rat b2 subunit coexpression experiments with Nilaparvata (N.) lugens a (Nla) subunits showed that Nla1/rat b2 forms a high affinity receptor, Nla2/rat b2 a low affinity receptor, while Nla3/rat b2 or Nla4/rat b2 constitute no neonicotinoid receptors (Liu et al, 2005(Liu et al, , 2006(Liu et al, , 2009a. Xenopus oocyte coexpression of Nla8 with rat b2 yielded also a low affinity neonicotinoid-gated ion channel, whose affinity could be increased 40-fold by coassembly with Nla3 (Yixi et al, 2009).…”

The cys-loop ligand-gated ion channel gene superfamily of the parasitoid wasp, Nasonia vitripennis

“…Similar studies of heterologously expressed ion channels from other insect species, including N. vitripennis, in combination with the use of three-dimensional models such as those based on the crystal structure of the molluscan acetylcholinebinding protein (Smit et al, 2003), will likely prove useful in the search for novel compounds that show selectivity for receptors of certain insect species as well as in determining the mechanisms of insecticide interaction with cysLGICs. For insect nAChRs, functional expression in heterologous systems has so far proven elusive, although a subunits of Drosophila and other insect species, such as Nilaparvata lugens, can form robust functional channels when coexpressed with a vertebrate b2 subunit and studies on such hybrid receptors have provided insights into the selectivity of neonicotinoids for insect nAChRs over those of vertebrates, regions of subunit proteins involved in imidacloprid interactions and the actions of different neonicotinoids Liu et al, 2009). Nasonia provides a powerful tool for the study of cysLGIC function.…”

Overall status of neonicotinoid insecticides in China: Production, application and innovation

“…In heterologous expression, the Nlα1 Y151S mutation showed 1.8-fold effects on imidacloprid sensitivity against Nlα1 and Nlα2 containing receptor Nlα1/Nlα2/rβ2, which was much less than the Nlα3 Y151S mutation with 9.2-fold effects on Nlα3 and Nlα8 containing receptor Nlα3/Nlα8/rβ2. 27,28) These data indicated that the Nlα3 Y151S mutation might be more important for imidacloprid resistance. All commercial neonicotinoid insecticides possess either an electron-withdrawing nitro (-NO 2 ) or cyano (-CN) group and have been postulated to contribute directly to their selectivity.…”

“…Heterologous expression and co-immunoprecipitation studies demonstrated the co-assembly of Nlα1, Nlα2 and Nlβ1 subunits into one receptor complex, Nlα1/Nlα2/Nlβ1, and of Nlα3, Nlα8 and Nlβ1 into another Nlα3/Nlα8/Nlβ1. [26][27][28] Radioligand-binding studies in N. lugens native nAChRs demonstrated that Nlα1/ Nlα2/Nlβ1 constituted the lower-affinity binding site and Nlα3/ Nlα8/Nlβ1 constituted the higher-affinity binding site of imidacloprid. In heterologous expression, the Nlα1 Y151S mutation showed 1.8-fold effects on imidacloprid sensitivity against Nlα1 and Nlα2 containing receptor Nlα1/Nlα2/rβ2, which was much less than the Nlα3 Y151S mutation with 9.2-fold effects on Nlα3 and Nlα8 containing receptor Nlα3/Nlα8/rβ2.…”