The long term use of many insecticides is continually threatened by the ability of insects to evolve resistance mechanisms that render the chemicals ineffective. Such resistance poses a serious threat to insect pest control both in the UK and worldwide. Resistance may result from either an increase in the ability of the insect to detoxify the insecticide or by changes in the target protein with which the insecticide interacts. DDT, the pyrethrins and the synthetic pyrethroids (the latter currently accounting for around 17% of the world insecticide market), act on the voltage‐gated sodium channel proteins found in insect nerve cell membranes. The correct functioning of these channels is essential for normal transmission of nerve impulses and this process is disrupted by binding of the insecticides, leading to paralysis and eventual death. Some insect pest populations have evolved modifications of the sodium channel protein which prevent the binding of the insecticide and result in the insect developing resistance. Here we review some of the work (done at Rothamsted Research and elsewhere) that has led to the identification of specific residues on the sodium channel that may constitute the DDT and pyrethroid binding sites. IUBMB Life, 59: 151‐162, 2007

We describe the first complete inventory of ATP-binding cassette (ABC) proteins from a multicellular organism, the model plant Arabidopsis thaliana. By the application of several search criteria, Arabidopsis was found to contain a total of 129 open reading frames (ORFs) capable of encoding ABC proteins, of which 103 possessed contiguous transmembrane spans and were identified as putative intrinsic membrane proteins. Fifty-two of the putative intrinsic membrane proteins contained at least two transmembrane domains (TMDs) and two nucleotide-binding folds (NBFs) and could be classified as belonging to one of five subfamilies of full-molecule transporters. The other 51 putative membrane proteins, all of which were half-molecule transporters, fell into five subfamilies. Of the remaining ORFs identified, all of which encoded proteins lacking TMDs, 11 could be classified into three subfamilies. There were no obvious homologs in other organisms for 15 of the ORFs which encoded a heterogeneous group of non-intrinsic ABC proteins (NAPs). Unrooted phylogenetic analyses substantiated the subfamily designations. Notable features of the Arabidopsis ABC superfamily was the presence of a large yeast-like PDR subfamily, and the absence of genes encoding bona fide cystic fibrosis transmembrane conductance regulator (CFTR), sulfonylurea receptor (SUR), and heavy metal tolerance factor 1 (HMT1) homologs. Arabidopsis was unusual in its large allocation of ORFs (a minimum of 0.5%) to members of the ABC protein superfamily.

The impact of neonicotinoid insecticides on the health of bee pollinators is a topic of intensive research and considerable current debate [1]. As insecticides, certain neonicotinoids, i.e., N-nitroguanidine compounds such as imidacloprid and thiamethoxam, are as intrinsically toxic to bees as to the insect pests they target. However, this is not the case for all neonicotinoids, with honeybees orders of magnitude less sensitive to N-cyanoamidine compounds such as thiacloprid [2]. Although previous work has suggested that this is due to rapid metabolism of these compounds [2-5], the specific gene(s) or enzyme(s) involved remain unknown. Here, we show that the sensitivity of the two most economically important bee species to neonicotinoids is determined by cytochrome P450s of the CYP9Q subfamily. Radioligand binding and inhibitor assays showed that variation in honeybee sensitivity to N-nitroguanidine and N-cyanoamidine neonicotinoids does not reside in differences in their affinity for the receptor but rather in divergent metabolism by P450s. Functional expression of the entire CYP3 clade of P450s from honeybees identified a single P450, CYP9Q3, that metabolizes thiacloprid with high efficiency but has little activity against imidacloprid. We demonstrate that bumble bees also exhibit profound differences in their sensitivity to different neonicotinoids, and we identify CYP9Q4 as a functional ortholog of honeybee CYP9Q3 and a key metabolic determinant of neonicotinoid sensitivity in this species. Our results demonstrate that bee pollinators are equipped with biochemical defense systems that define their sensitivity to insecticides and this knowledge can be leveraged to safeguard bee health.

A homology model of the housefly voltage-gated sodium channel was developed to predict the location of binding sites for the insecticides fenvalerate, a synthetic pyrethroid, and DDT an early generation organochlorine. The model successfully addresses the state-dependent affinity of pyrethroid insecticides, their mechanism of action and the role of mutations in the channel that are known to confer insecticide resistance. The sodium channel was modelled in an open conformation with the insecticide-binding site located in a hydrophobic cavity delimited by the domain II S4-S5 linker and the IIS5 and IIIS6 helices. The binding cavity is predicted to be accessible to the lipid bilayer and therefore to lipid-soluble insecticides. The binding of insecticides and the consequent formation of binding contacts across different channel elements could stabilize the channel when in an open state, which is consistent with the prolonged sodium tail currents induced by pyrethroids and DDT. In the closed state, the predicted alternative positioning of the domain II S4-S5 linker would result in disruption of pyrethroid-binding contacts, consistent with the observation that pyrethroids have their highest affinity for the open channel. The model also predicts a key role for the IIS5 and IIIS6 helices in insecticide binding. Some of the residues on the helices that form the putative binding contacts are not conserved between arthropod and non-arthropod species, which is consistent with their contribution to insecticide species selectivity. Additional binding contacts on the II S4-S5 linker can explain the higher potency of pyrethroid insecticides compared with DDT.