In the moth Agrotis ipsilon, the behavioral response of males to the female-emitted sex pheromone increases throughout adult life and following a prior exposure to sex pheromone whereas it is temporally inhibited after the onset of mating. This behavioral flexibility is paralleled with changes in neuronal sensitivity to pheromone signal within the primary olfactory centers, the antennal lobes. In the present study, we tested the hypothesis that neuroligins, postsynaptic transmembrane proteins known to act as mediators of neuronal remodeling, are involved in the olfactory modulation in A. ipsilon males. We cloned a full-length cDNA encoding neuroligin 1 which is expressed predominantly in brain and especially in antennal lobes. The level of neuroligin 1 expression in antennal lobes gradually raised from day-2 until day-4 of adult life as well as at 24 h, 48 h and 72 h following pre-exposure to sex pheromone and the temporal dynamic of these changes correlated with increased sex pheromone responsiveness. By contrast, there was no significant variation in antennal lobe neuroligin 1 expression during the post-mating refractory period. Taken together, these results highlighted that age- and odor experience-related increase in sex pheromone responsiveness is linked to the overexpression of neuroligin 1 in antennal lobes, thus suggesting a potential role played by this postsynaptic cell-adhesion molecule in mediating the plasticity of central olfactory system in A. ipsilon.
Title: Expression and modulation of neuroligin and neurexin in the olfactory organ of the cotton leaf worm Spodoptera littoralis Durand N, Chertemps T, Bozzolan F, Maibeche M Ref: Insect Sci, 24:210, 2017 : PubMed
Carboxylesterases are enzymes widely distributed within living organisms. In insects, they have been mainly involved in dietary metabolism and detoxification function. Interestingly, several members of this family called carboxylesterase-like adhesion molecules (CLAMs) have lost their catalytic properties and are mainly involved in neuro/developmental functions. CLAMs include gliotactins, neurotactins, glutactins, and neuroligins. The latter have for binding partner the neurexin. In insects, the function of these proteins has been mainly studied in Drosophila central nervous system or neuromuscular junction. Some studies suggested a role of neuroligins and neurexin in sensory processing but CLAM expression within sensory systems has not been investigated. Here, we reported the identification of 5 putative CLAMs expressed in the olfactory system of the model pest insect Spodoptera littoralis. One neuroligin, Slnlg4-yll and its putative binding partner neurexin SlnrxI were the most expressed in the antennae and were surprisingly associated with olfactory sensilla. In addition, both transcripts were upregulated in male antennae after mating, known to modulate the sensitivity of the peripheral olfactory system in S. littoralis, suggesting that these molecules could be involved in sensory plasticity.
Title: Neofunctionalization of Juvenile Hormone Esterase Duplication in Drosophila as an odorant-degrading enzyme towards food odorants Steiner C, Bozzolan F, Montagne N, Maibeche M, Chertemps T Ref: Sci Rep, 7:12629, 2017 : PubMed
Odorant degrading enzymes (ODEs) are thought to be responsible, at least in part, for olfactory signal termination in the chemosensory system by rapid degradation of odorants in the vicinity of the receptors. A carboxylesterase, specifically expressed in Drosophila antennae, called "juvenile hormone esterase duplication (JHEdup)" has been previously reported to hydrolyse different fruit esters in vitro. Here we functionally characterize JHEdup in vivo. We show that the jhedup gene is highly expressed in large basiconic sensilla that have been reported to detect several food esters. An electrophysiological analysis demonstrates that ab1A olfactory neurons of jhedup mutant flies exhibit an increased response to certain food acetates. Furthermore, mutant flies show a higher sensitivity towards the same odorants in behavioural assays. A phylogenetic analysis reveals that jhedup arose as a duplication of the juvenile hormone esterase gene during the evolution of Diptera, most likely in the ancestor of Schizophora, and has been conserved in all the 12 sequenced Drosophila species. Jhedup exhibits also an olfactory-predominant expression pattern in other Drosophila species. Our results support the implication of JHEdup in the degradation of food odorants in D. melanogaster and propose a neofunctionalization of this enzyme as a bona fide ODE in Drosophilids.
Pesticides have long been used as the main solution to limit agricultural pests, but their widespread use resulted in chronic or diffuse environmental pollutions, development of insect resistances, and biodiversity reduction. The effects of low residual doses of these chemical products on organisms that affect both targeted species (crop pests) but also beneficial insects became a major concern, particularly because low doses of pesticides can induce unexpected positive--also called hermetic--effects on insects, leading to surges in pest population growth at greater rate than what would have been observed without pesticide application. The present study aimed to examine the effects of sublethal doses of deltamethrin, one of the most used synthetic pyrethroids, known to present a residual activity and persistence in the environment, on the peripheral olfactory system and sexual behavior of a major pest insect, the cotton leafworm Spodoptera littoralis. We highlighted here a hormetic effect of sublethal dose of deltamethrin on the male responses to sex pheromone, without any modification of their response to host-plant odorants. We also identified several antennal actors potentially involved in this hormetic effect and in the antennal detoxification or antennal stress response of/to deltamethrin exposure.
The metabolism of volatile signal molecules by odorant degrading enzymes (ODEs) is crucial to the ongoing sensitivity and specificity of chemoreception in various insects, and a few specific esterases, cytochrome P450s, glutathione S-transferases (GSTs) and UDP-glycosyltransferases (UGTs) have previously been implicated in this process. Significant progress has been made in characterizing ODEs in Lepidoptera but very little is known about them in Diptera, including in Drosophila melanogaster, a major insect model. We have therefore carried out a transcriptomic analysis of the antennae of D. melanogaster in order to identify candidate ODEs. Virgin male and female and mated female antennal transcriptomes were determined by RNAseq. As with the Lepidoptera, we found that many esterases, cytochrome P450 enzymes, GSTs and UGTs are expressed in D. melanogaster antennae. As olfactory genes generally show selective expression in the antennae, a comparison to previously published transcriptomes for other tissues has been performed, showing preferential expression in the antennae for one esterase, JHEdup, one cytochrome P450, CYP308a1, and one GST, GSTE4. These largely uncharacterized enzymes are now prime candidates for ODE functions. JHEdup was expressed heterologously and found to have high catalytic activity against a chemically diverse group of known ester odorants for this species. This is a finding consistent with an ODE although it might suggest a general role in clearing several odorants rather than a specific role in clearing a particular odorant. Our findings do not preclude the possibility of odorant degrading functions for other antennally expressed esterases, P450s, GSTs and UGTs but, if so, they suggest that these enzymes also have additional functions in other tissues.
Mast syndrome is a complicated form of human hereditary spastic paraplegias, caused by a mutation in the gene acid cluster protein 33, which encodes a protein designated as "maspardin." Maspardin presents similarity to the alpha/beta-hydrolase superfamily, but might lack enzymatic activity and rather be involved in protein-protein interactions. Association with the vesicles of the endosomal network also suggested that maspardin may be involved in the sorting and/or trafficking of molecules in the endosomal pathway, a crucial process for maintenance of neuron health. Despite a high conservation in living organisms, studies of maspardin in other animal species than mammals were lacking. In the cotton armyworm Spodoptera littoralis, an insect pest model, analysis of an expressed sequence tag collection from antenna, the olfactory organ, has allowed identifying a maspardin homolog (SlMasp). We have investigated SlMasp tissue distribution and temporal expression by PCR and in situ hybridization techniques. Noteworthy, we found that maspardin was highly expressed in antennae and associated with the structures specialized in odorant detection. We have, in addition, identified maspardin sequences in numerous "nonmammalian" species and described here their phylogenetic analysis in the context of metazoan diversity. We observed a strong conservation of maspardin in metazoans, with surprisingly two independent losses of this gene in two relatively distant ecdysozoan taxa that include major model organisms, i.e., dipterans and nematodes.
BACKGROUND: Odorant-Degrading Enzymes (ODEs) are supposed to be involved in the signal inactivation step within the olfactory sensilla of insects by quickly removing odorant molecules from the vicinity of the olfactory receptors. Only three ODEs have been both identified at the molecular level and functionally characterized: two were specialized in the degradation of pheromone compounds and the last one was shown to degrade a plant odorant. METHODOLOGY: Previous work has shown that the antennae of the cotton leafworm Spodoptera littoralis, a worldwide pest of agricultural crops, express numerous candidate ODEs. We focused on an esterase overexpressed in males antennae, namely SlCXE7. We studied its expression patterns and tested its catalytic properties towards three odorants, i.e. the two female sex pheromone components and a green leaf volatile emitted by host plants. CONCLUSION: SlCXE7 expression was concomitant during development with male responsiveness to odorants and during adult scotophase with the period of male most active sexual behaviour. Furthermore, SlCXE7 transcription could be induced by male exposure to the main pheromone component, suggesting a role of Pheromone-Degrading Enzyme. Interestingly, recombinant SlCXE7 was able to efficiently hydrolyze the pheromone compounds but also the plant volatile, with a higher affinity for the pheromone than for the plant compound. In male antennae, SlCXE7 expression was associated with both long and short sensilla, tuned to sex pheromones or plant odours, respectively. Our results thus suggested that a same ODE could have a dual function depending of it sensillar localisation. Within the pheromone-sensitive sensilla, SlCXE7 may play a role in pheromone signal termination and in reduction of odorant background noise, whereas it could be involved in plant odorant inactivation within the short sensilla.
BACKGROUND: Carboxyl/cholinesterases (CCEs) are highly diversified in insects. These enzymes have a broad range of proposed functions, in neuro/developmental processes, dietary detoxification, insecticide resistance or hormone/pheromone degradation. As few functional data are available on purified or recombinant CCEs, the physiological role of most of these enzymes is unknown. Concerning their role in olfaction, only two CCEs able to metabolize sex pheromones have been functionally characterized in insects. These enzymes are only expressed in the male antennae, and secreted into the lumen of the pheromone-sensitive sensilla. CCEs able to hydrolyze other odorants than sex pheromones, such as plant volatiles, have not been identified. METHODOLOGY: In Spodoptera littoralis, a major crop pest, a diversity of antennal CCEs has been previously identified. We have employed here a combination of molecular biology, biochemistry and electrophysiology approaches to functionally characterize an intracellular CCE, SlCXE10, whose predominant expression in the olfactory sensilla suggested a role in olfaction. A recombinant protein was produced using the baculovirus system and we tested its catabolic properties towards a plant volatile and the sex pheromone components. CONCLUSION: We showed that SlCXE10 could efficiently hydrolyze a green leaf volatile and to a lesser extent the sex pheromone components. The transcript level in male antennae was also strongly induced by exposure to this plant odorant. In antennae, SlCXE10 expression was associated with sensilla responding to the sex pheromones and to plant odours. These results suggest that a CCE-based intracellular metabolism of odorants could occur in insect antennae, in addition to the extracellular metabolism occurring within the sensillar lumen. This is the first functional characterization of an Odorant-Degrading Enzyme active towards a host plant volatile.
Rapid degradation of odours after interaction with olfactory receptors is a critical step of the signal reception process. However, the implied mechanisms are still largely unknown in vertebrates as well as in insects. Involvement of odourant-degrading enzymes in odourant degradation within the antennae has been shown in some insect species and, in particular, esterases could play a key role in degradation of sex pheromones from Lepidoptera. Using a PCR-based strategy, we isolated cDNAs encoding two new esterases from two moths which used acetates as pheromone compounds: the Egyptian armyworm Spodoptera littoralis and the Mediterranean corn borer Sesamia nonagrioides. In antennae, both transcripts were clearly restricted to olfactory sensilla, suggesting their involvement in the degradation of odourant acetate components.
