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Molecular, anatomical, and developmental properties of the peripheral olfactory system in D. melanogaster. (A) Schematic of the two main insect olfactory receptor families. Odorant receptors (Ors) are seven transmembrane domain proteins that form heteromeric odor-gated ion channels composed of subunits of a ligand-specific (“tuning”) receptor and a co-receptor, Orco. Ionotropic receptors (Irs) are distantly related to ionotropic glutamate receptors, and function as odor-gated channel complexes composed of tuning Ir subunits and co-receptors (Ir8a or Ir25a). (B) Left: schematic of the D. melanogaster head (facing left) illustrating the main olfactory organs (antennae and maxillary palps, gray shading) and connectivity of two populations of olfactory sensory neurons (OSNs) to the antennal lobe in the brain. Right: schematic of the antenna, which is covered with diverse classes of sensory sensilla; the cellular organization of one sensillum, housing two OSNs, is shown on the far right (see text). (C) Left: schematic of the larval antennal imaginal disc, showing the concentric arcs of cells where different sensory organ precursors (SOP) are born. Amos- and Atonal-positive arcs give rise to OSN lineages expressing Ors and Irs, respectively, while other patterning determinants (not shown) are thought to specify SOP identity for different sensilla subtypes. Right: a simplified developmental lineage of an SOP producing a sensillum class with two OSNs. Two other potential neurons are removed by programmed cell death (PCD). Delta/Notch signaling determines the asymmetry of cell divisions, while many other patterning factors (not shown) are involved in specifying cell identity, encompassing both receptor expression and glomerular targeting of different OSNs (see text).
Source publication
The singular expression of insect olfactory receptors in specific populations of olfactory sensory neurons is fundamental to the encoding of odors in patterns of neuronal activity in the brain. How a receptor gene is selected, from among a large repertoire in the genome, to be expressed in a particular neuron is an outstanding question. Focusing on...
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Citations
... Although snRNA-seq enables unified profiling of all mosquito tissues, nuclear transcriptomes may not fully reflect cytoplasmic mRNA levels and provide no insights into protein expression 171 . This is particularly relevant for chemoreceptor co-expression studies, where post-transcriptional regulation could affect final receptor composition 172 . In addition, other detected transcripts could be untranslated, as seen in the recent work looking at ORs in the clonal raider ant Ooceraea biroi 173 . ...
The female mosquito’s remarkable ability to hunt humans and transmit pathogens relies on her unique biology. Here, we present the Mosquito Cell Atlas (MCA), a comprehensive single-nucleus RNA sequencing dataset of more than 367,000 nuclei from 19 dissected tissues of adult female and male Aedes aegypti , providing cellular-level resolution of mosquito biology. We identify novel cell types and expand our understanding of sensory neuron organization of chemoreceptors to all sensory tissues. Our analysis uncovers male-specific cells and sexually dimorphic gene expression in the antenna and brain. In female mosquitoes, we find that glial cells in the brain, rather than neurons, undergo the most extensive transcriptional changes following blood feeding. Our findings provide insights into the cellular basis of mosquito behavior and sexual dimorphism. The MCA aims to serve as a resource for the vector biology community, enabling systematic investigation of cell-type specific expression across all mosquito tissues.
... At the molecular level, odorants are detected by specific olfactory receptors (ORs) located on the dendrites of the OSNs. Generally, each insect OSN expresses only one type of OR, along with the broadly expressed odorant receptor co-receptor (Orco) (Mika and Benton 2021). ORs have an inverted seven-transmembrane topology compared to the G-protein-coupled receptors found in vertebrates (Benton et al. 2007). ...
Eurasian spruce bark beetle Ips typographus, a natural part of forest ecosystems, is a major threat to Norway spruce forests during outbreaks. Olfaction plays a crucial role in the survival and range expansion of these beetles, amid forest disturbances and climate change. As the current management strategies are suboptimal for controlling outbreaks, the reverse chemical ecology approaches based on pheromone receptors offer promising alternatives. While the search for pheromone receptors is in progress, recently found chromosomal inversions indicates signs of adaptation in this species. Our attempts to characterise one of the highly expressed odorant receptors, ItypOR33, located in an inversion, led to the discovery of polymorphic variants distributed with similar frequency across 18 European populations. Deorphanizing ItypOR33 and its variant ItypOR33a using the Drosophila empty-neuron system (DeNS) revealed ItypOR33 tuned to amitinol, a heterospecific pheromone component in Ips spp., whereas its variant tuned to (S)-(−)-ipsenol, a conspecific pheromone component of I. typographus. The in silico approaches revealed the structural basis of variations by predicting putative ligand-binding sites, tunnels and ligand-receptor interactions. However, no sex-specific differences were found in the ItypOR33 expression, and its ligand amitinol elicited behavioural and electrophysiolog-ical responses. Reporting population-level functional olfactory polymorphisms for the first time in a non-model organism-bark beetles, provides key evidence for further exploring their survival and adaptation in forests. Additionally, these findings indicate potential long-term complexities of managing bark beetles in forests.
... In insects and mammals, it is generally accepted idea that a single OSN expresses only one specific OR gene out of hundreds of receptor genes in the genome 17,18 . Recently, researchers have found some exceptions to this 'singular odorant receptor expression' theory in several holometabolous insect species, including fruit flies, mosquitoes, and ants [19][20][21][22] . ...
Periplaneta cockroaches use periplanone analogs as female sex pheromones to attract males. We previously identified two periplanone receptor genes, PameOR1 and PameOR2, in the American cockroach Periplaneta americana. Here, we report the identification of PameOR1-like, an additional olfactory receptor resembling PameOR1 in P. americana. PameOR1-like showed high-level sequence similarity to PameOR1, and is preferentially expressed in the male antennae. Quantitative expression analysis revealed that, in the adult male antennae, the expression level of PameOR1 is 2.4-fold higher than that of PameOR1-like. Fluorescent in situ hybridization revealed that PameOR1 and PameOR1-like are co-expressed in the periplanone-A-responsive sensory neurons within the single walled-B sensilla on the male adult antennae. These data support the idea that PameOR1 and PameOR1-like are generated by a recent gene duplication event and play a redundant function in sex pheromone reception in P. americana.
... The axons of these neurons project to the antennal lobe (AL) in the brain, where they form glomeruli, discrete spherical structures that serve as processing units. Each OSN generally expresses a single olfactory receptor (OR)together with the ubiquitous co-receptorwhich defines its response profile to odorant stimuli (15,16). The OR expression also dictates the specific glomerular target of each neuron, establishing a nearly oneto-one correspondence between ORs and glomeruli (17). ...
Ants are a monophyletic but diverse group of social insects whose heightened olfactory ability has been crucial to their evolutionary success. Their complex olfactory system is believed to have evolved due to the expansion of a specialized olfactory subsystem and the associated clade of olfactory receptors. Specifically, ants exhibit specialized antennal hairs known as basiconic sensilla, whose neurons project to a distinctive cluster of numerous, small glomeruli in their antennal lobes. This adaptation is believed to be linked to their social lifestyle, enabling the detection of recognition cues like cuticular hydrocarbons (CHCs), which are essential for nestmate recognition and maintaining colony cohesion. However, our understanding of the ant olfactory system remains incomplete, lacking evolutionary context and phylogenetic breadth, which leaves the complexity in their most recent common ancestor uncertain. We thus conducted a comparative study of neuroanatomical traits across the phylogeny of the Formicidae. Our findings reveal a common blueprint for the ant olfactory pathway, alongside lineage-specific adaptations. This highlights a dynamic evolution, particularly for the CHC-related subsystem. Ancestral trait reconstructions indicate that olfactory sophistication predates the most recent common ancestor of ants. Additionally, we found that the chemical complexity of species-specific recognition cues is associated with neuronal investment within the olfactory system. Lastly, behavioral experiments on anatomically divergent ant species show that, despite variation in neuroanatomical traits, ants consistently discriminate nestmates from non-nestmates. This suggests that the evolution of ants' olfactory system integrates sensory adaptations to diverse chemical environments, facilitating communication, a key to social behaviors.
... In insects and mammals, it is generally accepted idea that a single OSN expresses only one speci c OR gene out of hundreds of receptor genes in the genome [17,18] . Recently, researchers have found some exceptions to this 'singular odorant receptor expression' theory in several insect species [19][20][21][22] . ...
Periplaneta cockroaches use periplanone analogs as female sex pheromones to attract males. We previously identified two periplanone receptor genes, PameOR1 and PameOR2 , in the American cockroach Periplaneta americana . Here, we report the identification of PameOR1-like , an additional olfactory receptor resembling PameOR1 in P. americana . PameOR1-like showed high-level sequence similarity to PameOR1 , and is preferentially expressed in the male antennae. Quantitative expression analysis revealed that, in the adult male antennae, the expression level of PameOR1 is 2.4-fold higher than that of PameOR1-like . Fluorescent in situ hybridization (FISH) revealed that PameOR1 and PameOR1-like are expressed in the PA-responsive sensory neurons (PA-SNs) within the single walled -B ( sw -B) sensilla on the male adult antennae. These data support the idea that PameOR1 and PameOR1-like are generated by a recent gene duplication event and play a redundant function in sex pheromone reception in P. americana .
... Differential transcript expressions and volcano plots were plotted using Microsoft Excel (Redmond, WA, USA). Only transcripts with a TMP expression ≥ 5 were used for downstream analysis of sensory receptors as protein-coding transcripts expressed above a low expression threshold are more likely to be biologically relevant to fly olfaction [52]. ...
Background
Lucilia cuprina (Wiedemann, 1830) (Diptera: Calliphoridae) is the main causative agent of flystrike of sheep in Australia and New Zealand. Female flies lay eggs in an open wound or natural orifice, and the developing larvae eat the host’s tissues, a condition called myiasis. To improve our understanding of host-seeking behavior, we quantified gene expression in male and female antennae based on their behavior.
Methods
A spatial olfactometer was used to evaluate the olfactory response of L. cuprina mated males and gravid females to fresh or rotting beef. Antennal RNA-Seq analysis was used to identify sensory receptors differentially expressed between groups.
Results
Lucilia cuprina females were more attracted to rotten compared to fresh beef (> fivefold increase). However, males and some females did not respond to either type of beef. RNA-Seq analysis was performed on antennae dissected from attracted females, non-attracted females and males. Transcripts encoding sensory receptors from 11 gene families were identified above a threshold (≥ 5 transcript per million) including 49 ATP-binding cassette transporters (ABCs), two ammonium transporters (AMTs), 37 odorant receptors (ORs), 16 ionotropic receptors (IRs), 5 gustatory receptors (GRs), 22 odorant-binding proteins (OBPs), 9 CD36-sensory neuron membrane proteins (CD36/SNMPs), 4 chemosensory proteins (CSPs), 4 myeloid lipid-recognition (ML) and Niemann-Pick C2 disease proteins (ML/NPC2), 2 pickpocket receptors (PPKs) and 3 transient receptor potential channels (TRPs). Differential expression analyses identified sex-biased sensory receptors.
Conclusions
We identified sensory receptors that were differentially expressed between the antennae of both sexes and hence may be associated with host detection by female flies. The most promising for future investigations were as follows: an odorant receptor (LcupOR46) which is female-biased in L. cuprina and Cochliomyia hominivorax Coquerel, 1858; an ABC transporter (ABC G23.1) that was the sole sensory receptor upregulated in the antennae of females attracted to rotting beef compared to non-attracted females; a female-biased ammonia transporter (AMT_Rh50), which was previously associated with ammonium detection in Drosophila melanogaster Meigen, 1830. This is the first report suggesting a possible role for ABC transporters in L. cuprina olfaction and potentially in other insects.
Graphical Abstract
... Therefore, additional factors must underlie the differential susceptibility of tuning receptors to transcript loss in the absence of family-specific co-receptors. Recent work has revealed repressive transcriptional interactions between some odorant tuning receptors in insects, one of many mechanisms that support the expression of a singular tuning receptor in a particular olfactory neuron class (Jafari et al., 2021;Mika and Benton, 2021;Maguire et al., 2022). It will be interesting to determine if there are also transcriptional interactions between odorant tuning receptors and their co-receptors. ...
Insects detect odorants using two large families of heteromeric receptors, the Odorant Receptors (ORs) and Ionotropic Receptors (IRs). Most OR and IR genes encode odorant-binding “tuning” subunits, whereas four ( Orco , Ir8a , Ir25a , and Ir76b ) encode co-receptor subunits required for receptor function. Olfactory neurons are thought to degenerate in the absence of Orco in ants and bees, and limited data suggest this may happen to some olfactory neurons in Drosophila fruit flies as well. Here, we thoroughly examined the role of co-receptors on olfactory neuron survival in Drosophila . Leveraging knowledge that olfactory neuron classes are defined by the expression of different tuning receptors, we used tuning receptor expression in antennal transcriptomes as a proxy for the survival of distinct olfactory neuron classes. Consistent with olfactory neuron degeneration, expression of many OR-family tuning receptors is decreased in Orco mutants relative to controls, and transcript loss is progressive with age. The effects of Orco are highly receptor-dependent, with expression of some receptor transcripts nearly eliminated and others unaffected. Surprisingly, further studies revealed that olfactory neuron classes with reduced tuning receptor expression generally survive in Orco mutant flies. Furthermore, there is little apoptosis or neuronal loss in the antenna of these flies. We went on to investigate the effects of IR family co-receptor mutants using similar approaches and found that expression of IR tuning receptors is decreased in the absence of Ir8a and Ir25a , but not Ir76b . As in Orco mutants, Ir8a-dependent olfactory neurons mostly endure despite near-absent expression of associated tuning receptors. Finally, we used differential expression analysis to identify other antennal genes whose expression is changed in IR and OR co-receptor mutants. Taken together, our data indicate that odorant co-receptors are necessary for maintaining expression of many tuning receptors at the mRNA level. Further, most Drosophila olfactory neurons persist in OR and IR co-receptor mutants, suggesting that the impact of co-receptors on neuronal survival may vary across insect species.
... In insects, the olfactory system detects odorants, essential for feeding, mating, and avoiding hostile environments and toxic substances, and its rapid evolution suggests it is involved in fast adaptation to changing environments (Zhou, 2010). Since most odorants are small hydrophobic molecules (odorants and pheromones), they enter the antenna or palp sensillum and need to be transported by odorant-binding proteins to odorant receptors (ORs), to which they bind directly or complexed with an Obp (Zhou, 2010;Mika and Benton, 2021). Among the reported large families of odorantbinding and odorant receptor proteins (Zhou, 2010;Zhang et al., 2017;Mitchell et al., 2020;Tanaka et al., 2022), we gained information on the transcriptional behaviour of two odorant-binding protein (Obp.g1.i1 and Obp.g2.i1) and two odorant receptor protein genes (OR.g2.i1 and OR.g3.i1). ...
In phytophagous insects, adaptation to a new host is a dynamic process, in which early and later steps may be underpinned by different features of the insect genome. Here, we tested the hypothesis that early steps of this process are underpinned by a shift in gene expression patterns. We set up a short-term artificial selection experiment (10 generations) for the use of an alternative host (Cicer arietinum) on populations of the bean beetle Zabrotes subfasciatus. Using Illumina sequencing on young adult females, we show the selected populations differ in the expression of genes associated to stimuli, signalling, and developmental processes. Particularly, the “C. arietinum” population shows upregulation of histone methylation genes, which may constitute a strategy for fine-tuning the insect global gene expression network. Using qPCR on body regions, we demonstrated that the “Phaseolus vulgaris” population upregulates the genes polygalacturonase and egalitarian and that the expression of an odorant receptor transcript variant changes over generations. Moreover, in this population we detected the existence of vitellogenin (Vg) variants in both males and females, possibly harbouring canonical reproductive function in females and extracellular unknown functions in males. This study provides the basis for future genomic investigations seeking to shed light on the nature of the proximate mechanisms involved in promoting differential gene expression associated to insect development and adaptation to new hosts.
... 1,8 The insect chemosensory system is conceptually similar, even though insect chemoreceptors belong to different receptor classes. 1 Insect ORs are seven-pass transmembrane, ligand-gated ion channels that consist of conserved co-receptor subunits (Orco) along with one of several tuning OR subunits (OrX). [14][15][16][17][18][19][20] In addition, insect chemosensation employs gustatory receptors (GRs), which are also seven-pass transmembrane receptors, as well as ionotropic receptors (IRs), which are related to ionotropic glutamate receptors and likewise require co-receptors. 18,19 Receptor choice by OSNs is deterministic and based on transcription factor cascades. ...
... [14][15][16][17][18][19][20] In addition, insect chemosensation employs gustatory receptors (GRs), which are also seven-pass transmembrane receptors, as well as ionotropic receptors (IRs), which are related to ionotropic glutamate receptors and likewise require co-receptors. 18,19 Receptor choice by OSNs is deterministic and based on transcription factor cascades. 17,19,21,22 However, most of what we know about gene regulation in insects comes from studies of Drosophila melanogaster, the canonical insect model in genetics and neuroscience. ...
... 18,19 Receptor choice by OSNs is deterministic and based on transcription factor cascades. 17,19,21,22 However, most of what we know about gene regulation in insects comes from studies of Drosophila melanogaster, the canonical insect model in genetics and neuroscience. 19,23 Although insect ORs, like those of mammals, are frequently arranged in genomic tandem arrays, [24][25][26][27][28][29] most of the 60 D. melanogaster ORs are singletons. ...
Insects and mammals have independently evolved odorant receptor genes that are arranged in large genomic tandem arrays. In mammals, each olfactory sensory neuron chooses to express a single receptor in a stochastic process that includes substantial chromatin rearrangements. Here, we show that ants, which have the largest odorant receptor repertoires among insects, employ a different mechanism to regulate gene expression from tandem arrays. Using single-nucleus RNA sequencing, we found that ant olfactory sensory neurons choose different transcription start sites along an array but then produce mRNA from many downstream genes. This can result in transcripts from dozens of receptors being present in a single nucleus. Such rampant receptor co-expression at first seems difficult to reconcile with the narrow tuning of the ant olfactory system. However, RNA fluorescence in situ hybridization showed that only mRNA from the most upstream transcribed odorant receptor seems to reach the cytoplasm where it can be translated into protein, whereas mRNA from downstream receptors gets sequestered in the nucleus. This implies that, despite the extensive co-expression of odorant receptor genes, each olfactory sensory neuron ultimately only produces one or very few functional receptors. Evolution has thus found different molecular solutions in insects and mammals to the convergent challenge of selecting small subsets of receptors from large odorant receptor repertoires.
... serine/threonine protein kinase) have been previously found to be under putative selection in the range expanding, introduced B. hypnorum in the United Kingdom (Huml et al., 2021), and we similarly find this gene within our candidate genes under selection ( of foraging (Crall et al., 2017), guidance to flower rewards (Kantsa et al., 2019) and even the choice of foraging pollen or nectar during windy conditions (Mountcastle et al., 2015). Notably, our candidate loci were annotated to gene functions involved in the olfactory system, such as odorant receptors (odorant receptor 4-like) (Mika & Benton, 2021) and the neuropeptide CCHamide-1 receptor (Farhan et al., 2013). In addition to foraging navigation, bumblebees rely on their olfactory system for learning about their environment, for example, navigating to their nest, communicating food resources within the colony and associating flower scent with nectar reward (Adam et al., 2022;Molet et al., 2009;Palottini et al., 2018). ...
Invasive species are predicted to adjust their morphological, physiological and life‐history traits to adapt to their non‐native environments. Although a loss of genetic variation during invasion may restrict local adaptation, introduced species often thrive in novel environments. Despite being founded by just a few individuals, Bombus terrestris (Hymenoptera: Apidae) has in less than 30 years successfully spread across the island of Tasmania (Australia), becoming abundant and competitive with native pollinators. We use RADseq to investigate what neutral and adaptive genetic processes associated with environmental and morphological variation allow B. terrestris to thrive as an invasive species in Tasmania. Given the widespread abundance of B. terrestris , we expected little genetic structure across Tasmania and weak signatures of environmental and morphological selection. We found high gene flow with low genetic diversity, although with significant isolation‐by‐distance and spatial variation in effective migration rates. Restricted migration was evident across the mid‐central region of Tasmania, corresponding to higher elevations, pastural land, low wind speeds and low precipitation seasonality. Tajima's D indicated a recent population expansion extending from the south to the north of the island. Selection signatures were found for loci in relation to precipitation, wind speed and wing loading. Candidate loci were annotated to genes with functions related to cuticle water retention and insect flight muscle stability. Understanding how a genetically impoverished invasive bumblebee has rapidly adapted to a novel island environment provides further understanding about the evolutionary processes that determine successful insect invasions, and the potential for invasive hymenopteran pollinators to spread globally.
