Odorant Receptor Gene Expression Research Articles

Epigenetic programming of stochastic olfactory receptor choice.

The mammalian sense of smell relies upon a vast array of receptor proteins to detect odorant compounds present in the environment. The proper deployment of these receptor proteins in olfactory sensory neurons is orchestrated by a suite of epigenetic processes that remodel the olfactory genes in differentiating neuronal progenitors. The goal of this review is to elucidate the central role of gene regulatory processes acting in neuronal progenitors of olfactory sensory neurons that lead to a singular expression of an odorant receptor in mature olfactory sensory neurons. We begin by describing the principal features of odorant receptor gene expression in mature olfactory sensory neurons. Next, we delineate our current understanding of how these features emerge from multiple gene regulatory mechanisms acting in neuronal progenitors. Finally, we close by discussing the key gaps in our understanding of how these regulatory mechanisms work and how they interact with each other over the course of differentiation.

Screening for odorant receptor genes expressed in Aedes aegypti involved in host-seeking, blood-feeding and oviposition behaviors

BackgroundAedes aegypti is one of the most important vectors of zoonotic diseases worldwide, and its survival and reproductive processes depend heavily on its olfactory system. In this study, the expression levels of all odorant receptor (OR) genes of Ae. aegypti were explored during different physiological periods to identify olfactory genes that may be associated with mosquito blood-feeding and the search for oviposition sites.MethodsFour experimental groups, consisting of Ae. aegypti males, pre-blood-feeding females, post-blood-feeding females and post-oviposition females, were established. A total of 114 pairs of primers targeting all messenger RNA encoded by OR genes were designed based on the whole genome of Ae. aegypti. The expression of OR genes was evaluated by real-time fluorescence quantitative PCR for relative quantification and the comparison of differences between groups.ResultsA total of 53 differentially expressed OR genes were identified between males and females in Ae. aegypti antennae. Also, eight, eight and 13 differentially expressed OR genes were identified in pre- versus post-blood-feeding females, in pre- versus post-oviposition females and in post-blood-feeding versus post-oviposition females, respectively. In addition, 16 OR genes were significantly differentially expressed in multiple physiological periods of the mosquitoes.ConclusionsA large number of ORs with significant intergroup differences and high expression levels were screened in this study. Some of these genes are reported for the first time, providing possible targets for the development of mosquito control pathways based on the olfactory system.Graphical

Bcl11b is required for proper odorant receptor expression in the mouse septal organ.

Individual olfactory sensory neurons (OSNs) in the mouse main olfactory epithelium express a single odorant receptor (OR) gene from the repertoire of either class I or class II ORs. The transcription factor Bcl11b determines the OR class to be expressed in OSNs. The septal organ (SO), a small neuroepithelium located at the ventral base of the nasal septum, is considered as an olfactory subsystem because it expresses a specific subset of ORs. However, the mechanisms underlying the generation and differentiation of SO-OSN remain unknown. In the present study, we show that the generation and differentiation of SO-OSN employ the same genetic pathway as in the OSN lineage, which is initiated by the neuronal fate determinant factor Ascl1. Additionally, the key role of Bcl11b in the SO is demonstrated by the abnormal phenotypes of Bcl11b-deficient mice: significant reduction in the expression of OR genes and in the number of mature SO-OSNs. Although SO-OSNs are specified to express a subset of class II OR genes in wild-type mice, the Bcl11b deletion led to the expression of class I OR genes, while the expression of class II OR genes was significantly decreased, with one exception of Olfr15. These results indicate that Bcl11b is necessary for proper OR expression in SO-OSNs.

Identification and phylogenetics of Spodoptera frugiperda chemosensory proteins based on antennal transcriptome data

Understanding the interaction between the insect olfactory system and the environment is crucial for fully explaining the molecular mechanisms underlying insect behavior, and providing new strategies for integrated pest management. Although there is good evidence that olfactory proteins play a vital role in mediating insect behaviors, the olfactory mechanism of insects remains poorly understood. We identified a total of 71 chemosensory genes; 25 odorant-binding proteins (OBPs), 27 odorant receptors (ORs), 8 ionotropic receptors (IRs), 8 chemosensory proteins (CSPs) and 3 sensory neuron membrane proteins (SNMPs), in the antennae of male and female fall armyworms, Spodoptera frugiperda, an invasive global pest that causes significant economic damage worldwide. We used differential gene expression (DGE) and fragments per kilobase per million fragments (FPKM) values to compare the transcript levels of candidate chemosensory genes, and qRT-PCR to compare the expression levels of the OR gene, in male and female antennae. The expression of candidate OR genes in male and female antennae was consistent with the DGE data, and the expression of the SfruCL4419.Contig1-All and SfruUnigene1070-All genes was sex-biased. These results not only provide new information on the olfactory mechanism of S. frugiperda, and insects in general, but also suggest new gene targets for pest control.

Making sense of our sense of smell

Evolutionary Biology Human olfactory sensory neurons may have adapted specifically to our ecological niche. Saraiva et al. used an RNA sequencing–based approach to assess the diversity of these neurons across mammalian evolution. Full nasal biopsies were conducted on mice, rats, dogs, marmosets, macaques, and humans to catalog and quantify neuronal subtypes. The sensitivity of smell in mammals may have been fine-tuned by selective pressures that modified the diversity of neuronal odorant receptor gene expression. The most abundant neuronal subtypes were those tuned to detect odors more ecologically relevant for specific species, such as pheromones or other communicable odorants. Sci. Adv. 10.1126/sciadv.aax0396 (2019).

Transcriptomics reveal the molecular underpinnings of chemosensory proteins in Chlorops oryzae

BackgroundChemosensory proteins are a family of insect-specific chemical sensors that sense specific chemical cues and regulate insect behavior. Chemosensory proteins have been identified and analyzed in many insect species, such as Drosophila melanogaster, Bactrocera dorsalis and Calliphora stygia. This research has revealed that these proteins play a crucial role in insect orientation, predation and oviposition. However, little is known about the chemosensory proteins of Chlorops oryzae, a major pest of rice crops throughout Asia.ResultsComparative transcription analysis of the genes of Chlorops oryzae larvae, pupae and adults identified a total of 104 chemosensory genes, including 25 odorant receptors (ORs), 26 odorant-binding proteins (OBPs), 19 ionotropic receptors (IRs), 23 gustatory receptors (GRs) and 11 sensory neuron membrane proteins (SNMPs). The sequences of these candidate chemosensory genes were confirmed and used to construct phylogenetic trees. Quantitative real-time PCR (qRT-PCR) confirmed that the expression of candidate OR genes in different developmental stages was consistent with the fragments per kilobase per million fragments (FPKM) values of differentially expressed genes (DEGs).ConclusionsThe identification of chemosensory genes in C. oryzae provides a foundation for the investigation of the function of chemosensory proteins in this species, which, in turn, could allow the development of new, improved methods of controlling this pest.

Chemosensory genes in the antennal transcriptome of two syrphid species,Episyrphus balteatusandEupeodes corollae (Diptera: Syrphidae)

BackgroundPredatory syrphid larvae are an important natural enemy of aphids in cotton agro-ecosystems in China. Their behaviors in prey foraging, localization and oviposition greatly rely on the perception of chemical cues. As a first step to better understand syrphid olfaction at the molecular level, we have performed a systematic identification of their major chemosensory genes.ResultsMale and female antennal transcriptomes of Episyrphus balteatus and Eupeodes corollae were sequenced and assembled using Illumina HiSeq2000 technology. A total of 154 chemosensory genes in E. balteatus transcriptome, including candidate 51 odorant receptors (ORs), 32 ionotropic receptors (IRs), 14 gustatory receptors (GRs), 49 odorant-binding proteins (OBPs), 6 chemosensory proteins (CSPs) and 2 sensory neuron membrane proteins (SNMPs) were identified. In E. corollae transcriptome, we identified 134 genes including 42 ORs, 23 IRs, 16 GRs, 44 OBPs, 7 CSPs and 2 SNMPs. We have provided full-length sequences of the highly conserved co-receptor Orco, IR8a/25a family and carbon dioxide gustatory receptor in both syrphid species. The expression of candidate OR genes in the two syrphid species was evaluated by semi-quantitative reverse transcription PCR. There were no significant differences of transcript abundances in the respective male and female antenna, which is consistent with differentially expressed genes (DEGs) analysis using the FPKM value. The sequences of candidate chemosensory genes were confirmed and phylogenetic analysis was performed.ConclusionsThis research comprehensively analyzed and identified many novel candidate chemosensory genes regarding syrphid olfaction. It provides an opportunity for understanding how syrphid insects use chemical cues to conduct their behaviors among tritrophic interactions of plants, herbivorous insects, and natural enemies in agricultural ecosystems.

Regulatory Features for Odorant Receptor Genes in the Mouse Genome.

The odorant receptor genes, seven transmembrane receptor genes constituting the vastest mammalian gene multifamily, are expressed monogenically and monoallelicaly in each sensory neuron in the olfactory epithelium. This characteristic, often referred to as the one neuron–one receptor rule, is driven by mostly uncharacterized molecular dynamics, generally named odorant receptor gene choice. Much attention has been paid by the scientific community to the identification of sequences regulating the expression of odorant receptor genes within their loci, where related genes are usually arranged in genomic clusters. A number of studies identified transcription factor binding sites on odorant receptor promoter sequences. Similar binding sites were also found on a number of enhancers that regulate in cis their transcription, but have been proposed to form interchromosomal networks. Odorant receptor gene choice seems to occur via the local removal of strongly repressive epigenetic markings, put in place during the maturation of the sensory neuron on each odorant receptor locus. Here we review the fast-changing state of art for the study of regulatory features for odorant receptor genes.

Monogenic and Monoallelic Expression of Odorant Receptors

Odorant receptors (ORs) belong to a large gene family of rhodopsin-like G protein-coupled receptors (GPCRs). The mouse OR gene family is composed of ∼1000 OR genes, and the human OR gene family is composed of ∼400 OR genes. The OR genes are spread throughout the genome, and can be found in clusters or as solitary genes in almost all chromosomes. These chemosensory GPCRs are expressed in highly specialized cells, the olfactory sensory neurons of the nose. Each one of these neurons expresses a single OR gene out of the complete repertoire of genes. In addition, only one of the two homologous alleles of the chosen OR gene, the maternal or the paternal, is expressed per neuron. Here we review recent findings that help to elucidate the mechanisms underlying monogenic and monoallelic expression of OR genes.

Identification and expression pattern of candidate olfactory genes in Chrysoperla sinica by antennal transcriptome analysis

Chrysoperla sinica is one of the most prominent natural enemies of many agricultural pests. Host seeking in insects is strongly mediated by olfaction. Understanding the sophisticated olfactory system of insect antennae is crucial for studying the physiological bases of olfaction and could also help enhance the effectiveness of C. sinica in biological control. Obtaining olfactory genes is a research priority for investigating the olfactory system in this species. However, no olfaction sequence information is available for C. sinica. Consequently, we sequenced female- and male-antennae transcriptome of C. sinica. Many candidate chemosensory genes were identified, including 12 odorant-binding proteins (OBPs), 19 chemosensory proteins (CSPs), 37 odorant receptors (ORs), and 64 ionotropic receptors from C. sinica. The expression patterns of 12 OBPs, 19 CSPs and 37 ORs were determined by RT-PCR, and demonstrated antennae-dominantly expression of most OBP and OR genes. Our finding provided large scale genes for further investigation on the olfactory system of C. sinica at the molecular level.

Patterns of expression of odorant receptor genes in a Chagas disease vector

Rhodnius prolixus is a triatomine bug acting as a relevant vector of Chagas disease for which the genome sequence has been recently made available. Based on this information, a set of olfactory (ORs) and ionotropic receptor (IRs) genes potentially related to olfactory processes was characterized, and the expression patterns along bug development and in different structures potentially involved in promoting chemosensory-mediated behaviors were studied. For this, diverse bioinformatic procedures were used to validate gene models analyzing their structural and functional features and designing specific primers. Evolutionary relationships among R. prolixus olfactory coreceptors (RproOrco, RproIR25a, RproIR8a and RproIR76b) and their orthologues from other insects were shown to have mostly good bootstrap support values in phylogenetic trees. Moreover, antennal expression was confirmed for most genes included in the study. Both ORs and IRs showed antennal expression along the whole development of bugs of this species, with few exceptional receptors showing gradually increasing expression or expression restricted to the antennae of adult bugs. Finally, the expression of most of the selected genes was confirmed in other structures, such as rostri, tarsi, tibial pads and genitalia, which are potentially involved in promoting chemosensory-mediated behaviors. These results are discussed in terms of their relevance to advance in the understanding of the molecular bases of triatomine behavior.

Co-expression of six tightly clustered odorant receptor genes in the antenna of the malaria mosquito Anopheles gambiae

The behavior of female malaria mosquitoes, Anopheles gambiae, especially seeking out blood hosts or selecting oviposition sites, highly depends on the detection of relevant odorants by their sense of smell. This is mediated by olfactory sensory neurons (OSNs) which express distinct odorant receptor (OR) types. In the genome of A. gambiae 76 genes have been annotated to encode putative odorant receptors and the majority of these AgOR genes are arranged in clusters. To assess whether clustered AgOR genes are expressed in a characteristic manner we explored the topographic expression pattern of six tightly adjoined AgOR genes in the female antenna. Whole mount fluorescence in situ hybridization experiments were performed to visualize the olfactory neurons which express a distinct AgOR type in order to determine the number and the distribution of the cells. We found that within the thirteen antennal segments about 75 cells contain mRNA for the four receptor types AgOR13, AgOR15, AgOR17 and AgOR55. Moreover, about half of these cells also transcribe mRNA for the subtypes AgOR16 and AgOR47. Subsequent RT-PCR experiments with primer pairs spanning the coding regions of adjacent AgOR genes revealed the existence of polycistronic mRNA. This result indicates that individual genes were not transcribed but mRNA was comprised of coding sequence from several genes within the studied cluster. Taken together, the data indicate a unique principle for the expression of odorant receptor genes arranged in a large cluster and suggest that the corresponding olfactory neurons are endowed with a distinct set of odorant receptor types.

Nuclear architecture and gene silencing in olfactory sensory neurons.

Odorants are discriminated by hundreds of odorant receptor (OR) genes, which are dispersed throughout the mammalian genome. The OR genes are expressed in a highly specialized type of cell, the olfactory sensory neuron. Each one of these neurons expresses one of the 2 alleles from one single OR gene type. The mechanisms underlying OR gene expression are unclear. Here we describe recent work demonstrating that the olfactory sensory neuron shows a particular nuclear architecture, and that the genomic OR loci are colocalized in silencing heterochromatin compartments within the nucleus. These discoveries highlight the important role played by epigenetic modifications and nuclear genome organization in the regulation of OR gene expression.

Silencing of Odorant Receptor Genes by G Protein βγ Signaling Ensures the Expression of One Odorant Receptor per Olfactory Sensory Neuron

Olfactory sensory neurons express just one out of a possible ∼ 1,000 odorant receptor genes, reflecting an exquisite mode of gene regulation. In one model, once an odorant receptor is chosen for expression, other receptor genes are suppressed by a negative feedback mechanism, ensuring a stable functional identity of the sensory neuron for the lifetime of the cell. The signal transduction mechanism subserving odorant receptor gene silencing remains obscure, however. Here, we demonstrate in the zebrafish that odorant receptor gene silencing is dependent on receptor activity. Moreover, we show that signaling through G protein βγ subunits is both necessary and sufficient to suppress the expression of odorant receptor genes and likely acts through histone methylation to maintain the silenced odorant receptor genes in transcriptionally inactive heterochromatin. These results link receptor activity with the epigenetic mechanisms responsible for ensuring the expression of one odorant receptor per olfactory sensory neuron.

Temporal patterns of odorant receptor gene expression in adult and aged mice

In the mouse, the sense of smell relies predominantly on the expression of ~1200 odorant receptor (OR) genes in the main olfactory epithelium (MOE). Each mature olfactory sensory neuron (OSN) in the MOE is thought to express just one of these OR genes; conversely, an OR gene is expressed in thousands to tens of thousands of OSNs per mouse. Here, we have characterized temporal patterns of OR gene expression in a cohort of inbred C57BL6/N mice from the Aged Rodent Colonies of the National Institute on Aging. We applied the NanoString multiplex platform to quantify RNA abundance for 531 OR genes in whole olfactory mucosa (WOM) tissue samples. The five study groups were females aged 2, 6, 12, 18, and 31 months (mo). We classified the 531 temporal patterns using a step-down quadratic regression method for time course analysis. The majority of OR genes (58.4%) are classified as flat: there is no significant difference from a horizontal line within this time window. There are 32.8% of OR genes with a downward profile, 7.2% with an upward profile, and 1.7% with a convex or concave profile. But the magnitude of these decreases and increases tends to be small: only 4.3% of OR genes are differentially expressed (DE) at 31 mo compared to 2 mo. Interestingly, the variances of NanoString counts for individual OR genes are homogeneous among the age groups. Our analyses of these 15,930 OR gene expression data of C57BL6/N mice that were raised and housed under well-controlled conditions indicate that OR gene expression at the MOE level is intrinsically stable.

Activity-Dependent Modulation of Odorant Receptor Gene Expression in the Mouse Olfactory Epithelium

Activity plays critical roles in development and maintenance of the olfactory system, which undergoes considerable neurogenesis throughout life. In the mouse olfactory epithelium, each olfactory sensory neuron (OSN) stably expresses a single odorant receptor (OR) type out of a repertoire of ∼1200 and the OSNs with the same OR identity are distributed within one of the few broadly-defined zones. However, it remains elusive whether and how activity modulates such OR expression patterns. Here we addressed this question by investigating OR gene expression via in situ hybridization when sensory experience or neuronal excitability is manipulated. We first examined the expression patterns of fifteen OR genes in mice which underwent neonatal, unilateral naris closure. After four-week occlusion, the cell density in the closed (sensory-deprived) side was significantly lower (for four ORs), similar (for three ORs), or significantly higher (for eight ORs) as compared to that in the open (over-stimulated) side, suggesting that sensory inputs have differential effects on OSNs expressing different OR genes. We next examined the expression patterns of seven OR genes in transgenic mice in which mature OSNs had reduced neuronal excitability. Neuronal silencing led to a significant reduction in the cell density for most OR genes tested and thinner olfactory epithelium with an increased density of apoptotic cells. These results suggest that sensory experience plays important roles in shaping OR gene expression patterns and the neuronal activity is critical for survival of OSNs.

Expression of odorant receptor family, type 2 OR in the aquatic olfactory cavity of amphibian frog Xenopus tropicalis.

Recent genome wide in silico analyses discovered a new family (type 2 or family H) of odorant receptors (ORs) in teleost fish and frogs. However, since there is no evidence of the expression of these novel OR genes in olfactory sensory neurons (OSN), it remains unknown if type 2 ORs (OR2) function as odorant receptors. In this study, we examined expression of OR2 genes in the frog Xenopus tropicalis. The overall gene expression pattern is highly complex and differs depending on the gene and developmental stage. RT-PCR analysis in larvae showed that all of the OR2η genes we identified were expressed in the peripheral olfactory system and some were detected in the brain and skin. Whole mount in situ hybridization of the larval olfactory cavity confirmed that at least two OR2η genes so far tested are expressed in the OSN. Because tadpoles are aquatic animals, OR2η genes are probably involved in aquatic olfaction. In adults, OR2η genes are expressed in the nose, brain, and testes to different degrees depending on the genes. OR2η expression in the olfactory system is restricted to the medium cavity, which participates in the detection of water-soluble odorants, suggesting that OR2ηs function as receptors for water-soluble odorants. Moreover, the fact that several OR2ηs are significantly expressed in non-olfactory organs suggests unknown roles in a range of biological processes other than putative odorant receptor functions.

Determinants of the Drosophila Odorant Receptor Pattern

In most olfactory systems studied to date, neurons that express the same odorant receptor (Or) gene are scattered across sensory epithelia, intermingled with neurons that express different Or genes. In Drosophila, olfactory sensilla that express the same Or gene are dispersed on the antenna and the maxillary palp. Here we show that Or identity is specified in a spatially stereotyped pattern by the cell-autonomous activity of the transcriptional regulators Engrailed and Dachshund. Olfactory sensilla then become highly motile and disperse beneath the epidermis. Thus, positional information and cell motility underlie the dispersed patterns of Drosophila Or gene expression.

Homeodomain binding motifs modulate the probability of odorant receptor gene choice in transgenic mice

Odorant receptor (OR) genes constitute with 1200 members the largest gene family in the mouse genome. A mature olfactory sensory neuron (OSN) is thought to express just one OR gene, and from one allele. The cell bodies of OSNs that express a given OR gene display a mosaic pattern within a particular region of the main olfactory epithelium. The mechanisms and cis-acting DNA elements that regulate the expression of one OR gene per OSN — OR gene choice — remain poorly understood. Here, we describe a reporter assay to identify minimal promoters for OR genes in transgenic mice, which are produced by the conventional method of pronuclear injection of DNA. The promoter transgenes are devoid of an OR coding sequence, and instead drive expression of the axonal marker tau-β-galactosidase. For four mouse OR genes (M71, M72, MOR23, and P3) and one human OR gene (hM72), a mosaic, OSN-specific pattern of reporter expression can be obtained in transgenic mice with contiguous DNA segments of only ~300bp that are centered around the transcription start site (TSS). The ~150bp region upstream of the TSS contains three conserved sequence motifs, including homeodomain (HD) binding sites. Such HD binding sites are also present in the H and P elements, DNA sequences that are known to strongly influence OR gene expression. When a 19mer encompassing a HD binding site from the P element is multimerized nine times and added upstream of a MOR23 minigene that contains the MOR23 coding region, we observe a dramatic increase in the number of transgene-expressing founders and lines and in the number of labeled OSNs. By contrast, a nine times multimerized 19mer with a mutant HD binding site does not have these effects. We hypothesize that HD binding sites in the H and P elements and in OR promoters modulate the probability of OR gene choice.

Differential expression of odorant receptor genes involved in the sexual isolation of two Heliothis moths

Moth sexual communication systems are highly diverse, but the mechanisms underlying their evolutionary diversification remain unclear. Recently, genes coding for odorant receptors (ORs) OR6, OR14, OR15 and OR16 have been genetically associated with species-specific male response to female pheromone blends in Heliothis virescens (Hv) and Heliothis subflexa (Hs). Quantitative real-time PCR analysis indicates that expression of HvOR6, HsOR6, HvOR14, HsOR14, HvOR15 and HsOR15 is male biased, which supports the hypothesis that they have a role in mediating female sex pheromone detection. The genes HvOR14, HvOR15 and HvOR16 are expressed at higher levels than their corresponding orthologues HsOR14, HsOR15 and HsOR16 in male antennae, while HvOR6 and HsOR6 transcripts are equally abundant in male antennae. The lack of higher expression of any of the receptor genes in H. subflexa antennae suggests that interspecific sequence differences, rather than gene regulation differences, underly the species-specific male response to pheromone components.