Functional characterization of a sex pheromone receptor in the pest moth Spodoptera littoralis by heterologous expression in Drosophila

Abstract

Moth sex pheromone communication is recognised as a long-standing model for insect olfaction studies, and a widespread knowledge has been accumulated on this subject thanks to numerous chemical, electrophysiological and behavioural studies. A key step has been the identification of candidate sex pheromone receptors, opening new routes to understanding the specificity and sensitivity of this communication system, but only few of these receptors have as yet been functionally characterised. In this context, we aim at unravelling the molecular bases of pheromone reception in the noctuid moth Spodoptera littoralis. Taking advantage of a collection of antennal-expressed sequence tags, we previously identified three fragments of candidate pheromone receptors in this species. Here, we report full-length cloning of one of these receptors, named SlitOR6. Both sequence and expression pattern analyses were consistent with its annotation as a pheromone receptor, which we further confirmed by functional characterization. Using Drosophila antennae as a heterologous expression system, we identified a single component of the pheromone blend of S. littoralis, (Z,E)-9,12-tetradecadienyl acetate, as the ligand of SlitOR6. Two strategies were employed: (i) expressing SlitOR6 in the majority of Drosophila olfactory neurons, in addition to endogenous receptors, and monitoring the responses to pheromone stimuli by electroantennography; (ii) replacing the Drosophila pheromone receptor OR67d with SlitOR6 and monitoring the response by single sensillum recordings. Results were fully congruent and responses to (Z,E)-9,12-tetradecadienyl acetate were highly specific in both heterologous systems. This approach appears to be efficient and reliable for studying moth pheromone receptors in an in vivo context.

A predicted sex pheromone receptor of codling moth Cydia pomonella detects the plant volatile pear ester

“…CpomOR3 was almost equally sensitive when expressed in trichoid T1 and basiconic ab3 sensilla, except at the highest dose of pear ester (Figure 4). Interestingly, the pheromone receptors BmorOR1 of silkmoth B. mori and SlitOR6 of cotton leafworm moth S. littoralis were more sensitive when expressed in T1 than in ab3 sensilla (Syed et al, 2010;Montagné et al, 2012). This indicates that T1 sensilla, containing an important PR partner, the sensory neuron membrane protein (Benton et al, 2007), are more adapted for correct PR functioning, whereas plant odorant ORs function equally well in T1 or ab3.…”

“…Amino acid sequences of CpomORs clustering in the candidate PR clade (Bengtsson et al, 2012) were included in a dataset together with sequences of candidate PRs from the following Lepidoptera: Antheraea polyphemus (Forstner et al, 2009), Bombyx mori (Nakagawa et al, 2005), Danaus plexippus (Zhan et al, 2011), Diaphania indica (Mitsuno et al, 2008), Epiphyas postvittana (Jordan et al, 2009), Heliconius melpomene (Heliconius Genome Consortium, 2012), Helicoverpa armigera (Liu et al, 2012), Heliothis virescens (Grosse-Wilde et al, 2007;Wang et al, 2010), Manduca sexta (Grosse-Wilde et al, 2010), Mythimna separata (Mitsuno et al, 2008), Ostrinia furnacalis (Miura et al, 2010;Leary et al, 2012), O. nubilalis (Wanner et al, 2010;Leary et al, 2012), O. scapulalis (Miura et al, 2009(Miura et al, , 2010, Plutella xylostella (Mitsuno et al, 2008;Sun et al, 2013), Spodoptera exigua and S. littoralis (Legeai et al, 2011;Montagné et al, 2012). Sequences from B. mori (BmorOR6) and H. melpomene (HmelOR5, 6, and 7) were also included in the dataset as external groups, since they belong to the sister group to the PR clade (Poivet et al, 2013).…”

“…While the empty neuron system has been used mainly to functionally characterize general odorant receptors, pheromone receptors may respond more strongly when expressed in T1 rather than in ab3A (Syed et al, 2010;Montagné et al, 2012).…”

“…The OR repertoire of Drosophila has been studied exhaustively (Hallem et al, 2004;Kreher et al, 2005;Hallem and Carlson, 2006) and current research aims at other insect groups. For moths, a number of ORs and PRs have been identified and functionally characterized, using various heterologous expression systems, including human embryonic kidney (HEK) cells (Grosse-Wilde et al, 2007), Xenopus oocytes (Sakurai et al, 2004;Nakagawa et al, 2005;Jiang et al, 2014), Sf9, a cell line derived from fall armyworm Spodoptera frugiperda ovaries (Jordan et al, 2009), and Drosophila OSNs (Syed et al, 2010;Montagné et al, 2012), which is an in vivo antennal expression approach.…”

A Conserved Odorant Receptor Detects the Same 1-Indanone Analogs in a Tortricid and a Noctuid Moth

“…A number of lepidopteran OR gene repertoires have been described, following genome and transcriptome sequencing (Jordan et al, 2009;Grosse-Wilde et al, 2011;Montagné et al, 2012Montagné et al, , 2014Cao et al, 2014;Gu et al, 2014;Liu et al, 2014;Corcoran et al, 2015;Yang et al, 2015;Zhang et al, 2015a,b). In our own transcriptome sequence analyses of the antennae of the codling moth (Cydia pomonella: Tortricidae; Bengtsson et al, 2012) and the cotton leafworm (Spodoptera littoralis: Noctuidae; Legeai et al, 2011;Jacquin-Joly et al, 2012;Poivet et al, 2013) we have identified one OR (OR19) with relatively high sequence similarity in both species.…”