Probing insect odorant receptors with their cognate ligands: Insights into structural features

Abstract

Oodorant receptors (ORs) are essential for insect survival in the environment and thus are ideal molecular targets for the design of insect-inspired modern green chemicals to control populations of agricultural pests and insects of medical importance. Although insect ORs are known for more than a decade, their structural biology is still in its infancy. Here, we unravel the first structural features of ORs from the malaria mosquito, the Southern house mosquito and the silkworm moth. The second extracellular loops (ECL-2s) of their predicted structures are much longer than ECL-1s and ECL-3s. The 27 amino-acid-residue-long of the ECL-2s in mosquito and the 43 amino-acid-residue-long ECL2s in moth ORs are well-conserved. About one-third of the residues are identical, including 3-4 Pro residues. Thorough examination of well-conserved residues in these structures, by point mutation and functional assay with the Xenopus oocyte recording system, strongly suggest that these “loops” include three β-turns and some degree of folding. In the Southern house mosquito three Pro residues in ECL-2 are essential for full activation of the receptor, which is finely tuned to the oviposition attractant 3-methylindole. Additionally, the “corner residues” of prolines, including Gly, Tyr, and Leu are functionally important thus suggesting that turns are stabilized not only by backbone hydrogen bonds, but also by side-chain interactions. Examination of ECL-2s from a distant taxonomical group suggests these ECL-2 loops might be functionally important in all insect ORs. Two of the four Pro residues in the predicted ECL-2 of the bombykol receptor in the silkworm moth, BmorOR1, are essential for function. Experimental evidence indicates that these loops may not be specificity determinants, but they may form a cover to the yet-to-be-identified membrane embedded binding cavities of insect ORs.

Evolution of two receptors detecting the same pheromone compound in crop pest moths of the genus Spodoptera

“…There are experimental evidences that mutations in TM2, 3 and 4 domains affect ligand binding in Drosophila, mosquito or moth receptors (Nichols and Luetje, 2010;Pellegrino et al, 2011;Leary et al, 2012;Hughes et al, 2014) whereas mutations in TM5, 6 and 7 affect the ion channel function of a moth PR (Nakagawa et al, 2012). An in silico analysis of insect OR structural features also identified N-ter, EL2 and IL3 domains as the most evolutionary constrained (Hopf et al, 2015), which is consistent with their importance for the correct functioning of ORs (Benton et al, 2006;Jin et al, 2008;Xu and Leal, 2013). Further experimental validations are needed to confirm whether positive selection has indeed been a driving force for the diversification of moth PRs or not.…”

Molecular identification and sex distribution of two chemosensory receptor families in Athetis lepigone by antennal transcriptome analysis

“…Consistent with other moths (Bengtsson et al, 2012;Feng et al, 2015;Zhang et al, 2013Zhang et al, , 2015, the AlepORco gene also displayed a very high mRNA abundance. Many previous studies showed that ORco could act as a chaperone and dimerization partner for other ORs, forming a ligand gated ion channel to specific ligands (Krieger et al, 2003(Krieger et al, , 2005Nakagawa et al, 2005;Xu and Leal, 2013), and ORco was more highly conserved in insects than other ORs (Krieger et al, 2003;Leal, 2013;Robertson et al, 2003;Vogt, 2005), thus it required more ORco transcripts to activate different ORs. In addition, we identified 4 candidate PR genes by sex expression and physiologic analysis.…”

Differential expression of olfactory genes in the southern house mosquito and insights into unique odorant receptor gene isoforms

“…Antennae and legs dissected from blood-fed adult females were used for RNA-Seq as well as to generate templates for validation by qPCR and gene cloning. Deorphanization of a unique OR gene was performed with the Xenopus oocyte expression system (9,35). Oocytes expressing CquiOR95b along with CquiOrco were challenged with large panel of physiologically relevant odorants.…”