Preventing alloimmune rejection using circular RNA FSCN1-silenced dendritic cells in heart transplantation.

BACKGROUND: While heart transplantation is used as a standard treatment for heart failure, transplant rejection continues to pose a challenge. Recent evidence has shown that circular RNA (circRNA) is a new type of gene regulator in cell development. Our aim was to demonstrate that treatment with tolerogenic dendritic cells (Tol-DCs) generated by circular RNA FSCN1 (circFSCN1) silencing could prevent alloimmune rejection and prolong heart graft survival in heart transplantation. METHODS: Bone marrow-derived DCs were transfected with circFSCN1 siRNA in vitro. The circFSCN1 level was measured by qRT-PCR. DC maturation was determined by flow cytometry. Mixed lymphocyte reactions (MLRs) were conducted to assess the function of DCs to activate T cells and to generate regulatory T cells (Tregs). In situ RNA hybridization and fluorescent microscopy were performed to detect the distribution of circFSCN1 in DCs. A heterotopic allogeneic murine heart transplantation was conducted where recipients were pre-treated with donor derived circFSCN1-silenced Tol-DCs. Heartbeat was monitored to assess immune rejection. RESULTS: Exonic circFSCN1 was highly expressed in the cytoplasm of mature DCs. Knockdown of circFSCN1 using siRNA arrested DCs at an immature state, impaired DC's ability to activate T cells and enhanced Treg generation. Treatment with circFSCN1-silenced Tol-DCs prevented alloimmune rejection, prolonged allograft survival, reduced fibrosis, and induced Tregs in vivo. CONCLUSIONS: Knockdown of circFSCN1 induces Tol-DCs and treatment with these Tol-DCs prevents alloimmune rejection and prolongs allograft survival. This is a promising therapeutic target to combat transplant rejection in heart transplantation and increases our understanding of circRNA in the immune system.

Coordinative mediation of the response to alarm pheromones by three odorant binding proteins in the green peach aphid Myzus persicae.

Odorant binding proteins (OBPs) play an essential role for insect chemosensation in insect peripheral nervous systems of antennae. Each antennal sensilla contains more than one OBP at high concentrations but the interactions and cooperation between co-localized OBPs are rarely reported. In present study, we cloned, expressed and purified eight OBPs of the green peach aphid Myzus persicae. The effects of knocking down the expression of these OBP genes by RNAi on the electrophysiological and behavioural responses of M. persicae to the aphid alarm pheromone, (E)-ß-farnesene (EßF) were investigated. The results showed that the aphids could still be repelled by EßF when the expression of each of three OBP genes was individually knocked down. However, the simultaneous knockdown of MperOBP3/7/9 expression significantly reduced the electrophysiological response and the repellent behaviours of M. persicae to EßF than the single OBP gene knockdown (P < 0.05). Rather than a normal saturation binding curve of individual OBP, the binding curve of MperOBP3/7/9 is bell-shaped with a higher affinity for the fluorescent probe N-phenyl-1-naphthylamine (1-NPN). The competitive binding assays confirmed that MperOBP3, MperOBP7, MperOBP9 and MperOBP3/7/9 mixture exhibited a stronger binding affinity for EßF, than for sex pheromones and plant volatiles with a dissociation constant of 2.5 µM, 1.1 µM, 3.9 µM and 1.0 µM, respectively. The competitive binding curve of MperOBP3/7/9 mixture to EßF is shallow without bottom plateau, suggesting a conformational change and a rapid dissociation after the displacement of all 1-NPN (in vivo after the saturation binding of all OBPs by EßF). The interaction between OBPs and formation of a heterogeneous unit may facilitate the delivery EßF to the OR at electrophysiological and behavioural levels during insect odorant signal transduction thus mediate M. persicae response to the alarm pheromone EßF.

Two highly related odorant receptors specifically detect α-bile acid pheromones in sea lamprey (Petromyzon marinus).

Pheromones play critical roles in habitat identification and reproductive behavior synchronization in the sea lamprey (Petromyzon marinus). The bile acid 3-keto petromyzonol sulfate (3kPZS) is a major component of the sex pheromone mixture from male sea lamprey that induces specific olfactory and behavioral responses in conspecific individuals. Olfactory receptors interact directly with pheromones, which is the first step in their detection, but identifying the cognate receptors of specific pheromones is often challenging. Here, we deorphanized two highly related odorant receptors (ORs), OR320a and OR320b, of P. marinus that respond to 3kPZS. In a heterologous expression system coupled to a cAMP-responsive CRE-luciferase, OR320a and OR320b specifically responded to C24 5α-bile acids, and both receptors were activated by the same set of 3kPZS analogs. OR320a displayed larger responses to all 3kPZS analogs than did OR320b. This difference appeared to be largely determined by a single amino acid residue, Cys-792.56, the C-terminal sixth residue relative to the most conserved residue in the second transmembrane domain (2.56) of OR320a. This region of TM2 residues 2.56-2.60 apparently is critical for the detection of steroid compounds by odorant receptors in lamprey, zebrafish, and humans. Finally, we identified OR320 orthologs in Japanese lamprey (Lethenteron camtschaticum), suggesting that the OR320 family may be widely present in lamprey species and that OR320 may be under purifying selection. Our results provide a system to examine the origin of olfactory steroid detection in vertebrates and to define a highly conserved molecular mechanism for steroid-ligand detection by G protein-coupled receptors.

Molecular characterization and volatile binding properties of pheromone binding proteins and general odorant binding proteins in Conogethes pinicolalis (Lepidoptera: Crambidae).

Conogethes pinicolalis (Lepidoptera: Crambidae), a major pine pest, possesses a sensitive olfactory system to locate its host. Pheromone binding proteins (PBPs) and general odorant binding proteins (GOBPs) are two types of proteins involved in the process. In this work, we used phylogenetic analysis, gene expression, fluorescence competitive binding assay, and molecular docking to characterize PBPs and GOBPs in C. pinicolalis. The phylogenetic unrooted tree revealed the C. pinicolalis GOBPs and PBPs amino acid sequences showed very close relation with Conogethes punctiferalis (yellow peach moth). Meanwhile, both the PBPs and GOBPs were specifically expressed in the antennae. Binding affinities of PBPs and GOBPs to 19 volatile compounds were tested. PBP2 shows the strongest binding to E10-16: Ald with a Ki value of 0.28/1.66 µM; GOBP1 to Z10-16:Ald with a Ki value of 3.11/4.24 µM. Furthermore, molecular docking reveals potential active sites in PBP2 and GOBP1 to interact with most of the tested volatiles. Finally, we demonstrate that PBP2 and GOBP1 are the dominant genes in their respective families using several different assays. In conclusion, PBP2 and GOBP1 genes may play similar roles in detecting and transporting sex pheromones and host plant volatiles in C. pinicolalis.

The olfactory G protein-coupled receptor (Olfr-78/OR51E2) modulates the intestinal response to colitis.

Olfactory receptor-78 (Olfr-78) is a recently identified G protein-coupled receptor activated by short-chain fatty acids acetate and propionate. A suggested role for this receptor exists in the prostate where it may influence chronic inflammatory response leading to intraepithelial neoplasia. Olfr-78 has also been shown to be expressed in mouse colon. Short-chain fatty acids and their receptors are well known to modulate inflammation in the gut. Considering this possibility, we first explored if colitis regulated Olfr-78 expression in the gut, where we observed a significant reduction in the expression of Olfr-78 transcript in mouse models of dextran sodium sulfate (DSS)- and 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis. To more directly test this, mice deficient in Olfr-78 were administered with DSS in water for 7 days and were found to have increased expression of IL-1ß and inflammatory signs in colon compared with control mice. Next, we explored the expression of its human counterpart olfactory receptor family 51, subfamily E, member 2 (OR51E2) in human intestinal samples and observed that it was in fact also expressed in human colon samples. RNA sequence analysis revealed significant changes in the genes involved in infection, immunity, inflammation, and colorectal cancer between wild-type and Olfr-78 knockout mice. Collectively, our findings show that Olfr-78 is highly expressed in colon and downregulated in DSS- and TNBS-induced colitis, and DSS-treated Olfr-78 null mice had increased colonic expression of cytokine RNA levels, suggesting a potential role for this receptor in intestinal inflammation. Future investigations are needed to understand how Olfr-78/OR51E2 in both mouse and human intestine modulates gastrointestinal pathophysiology.

Molecular cloning and comparative analysis of transcripts encoding chemosensory proteins from two plant bugs, Lygus lineolaris and Lygus hesperus.

Chemosensory proteins (CSPs) are soluble carrier proteins typically characterized by a six-helix bundle structure joined by two disulfide bridges and a conserved Cys spacing pattern (C1-X6-8 -C2-X16-21 -C3-X2 -C4). CSPs are functionally diverse with reported roles in chemosensation, immunity, development, and resistance. To expand our molecular understanding of CSP function in plant bugs, we used recently developed transcriptomic resources for Lygus lineolaris and Lygus hesperus to identify 17 and 14 CSP-like sequences, respectively. The Lygus CSPs are orthologous and share significant sequence identity with previously annotated CSPs. Three of the CSPs are predicted to deviate from the typical CSP structure with either five or seven helical segments rather than six. The seven helix CSP is further differentiated by an atypical C3-X3 -C4 Cys spacing motif. Reverse transcriptase PCR-based profiling of CSP transcript abundance in adult L. lineolaris tissues revealed broad expression for most of the CSPs with antenna specific expression limited to a subset of the CSPs. Comparative sequence analyses and homology modeling suggest that variations in the amino acids that comprise the Lygus CSP binding pockets affect the size and nature of the ligands accommodated.

Activation of PSGR with ß-ionone suppresses prostate cancer progression by blocking androgen receptor nuclear translocation.

The prostate-specific G protein-coupled receptor (PSGR) is a class A G protein-coupled receptor (GPCR) that is specifically expressed in prostate epithelial cells, and its expression has been linked to prostate cancer (PCa) progression. Here, we show that activation of PSGR with its ligand ß-ionone, an end-ring analog of ß-carotenoid, can suppress PCa cell growth both in vitro and in vivo model. Dissection of the mechanism underlying this relationship reveals that activation of PSGR by ß-ionone suppresses AR nuclear translocation via phosphorylation of AR at residue Ser650 by p38 and JNK, which leads to the suppression of AR transactivation, further suppressing PCa cell growth. Overall, we link a cancer cell-specific GPCR with the nuclear AR and show that targeting PSGR can provide us a new target to combat PCa better.

Reversing Behavioral, Neuroanatomical, and Germline Influences of Intergenerational Stress.

BACKGROUND: Stressors affect populations exposed to them as well as offspring. Strategies preventing the intergenerational propagation of effects of stress would benefit public health. Olfactory cue-based fear conditioning provides a framework to address this issue. METHODS: We 1) exposed adult male mice to an odor, acetophenone (Ace) or Lyral (parental generation [F0]-Exposed), 2) trained mice to associate these odors with mild foot shocks (F0-Trained), and 3) trained mice to associate these odors with mild foot shocks and then extinguished their fear toward these odors with odor-only presentations (F0-Extinguished). We then examined sensitivity of future generation (F1) offspring to these odors, expression of M71 odorant (Ace-responsive) and MOR23 odorant (Lyral-responsive) receptor-expressing cell populations in F1 offspring, and DNA methylation at genes encoding the Ace- (Olfr151, Olfr160) and Lyral- (Olfr16) responsive receptors in F0 sperm. RESULTS: Extinguishing fear toward Ace or Lyral of F0 male mice (F0-Extinguished) that had been fear conditioned with Ace or Lyral, respectively, results in F1-Extinguished offspring that do not demonstrate behavioral sensitivity to Ace or Lyral, respectively, and do not have enhanced representation for M71 or MOR23 odorant receptors in the olfactory system, as is observed in F1-Trained-Ace or F1-Trained-Lyral cohorts, respectively. The promoters of genes encoding Olfr151 and Olfr160 receptors are less methylated in F0-Trained-Ace sperm compared with F0-Exposed-Ace sperm. The Olfr16 promoter is less methylated in F0-Trained-Lyral sperm compared with F0-Exposed-Lyral sperm, and F0-Extinguished-Lyral sperm have methylation levels comparable to F0-Exposed-Lyral sperm. CONCLUSIONS: Our study demonstrates the potential of using extinction-based behavioral strategies to reverse influences of parental stress in offspring and in the parental germline.

Functional characterization of odorant-binding proteins from the scarab beetle Holotrichia oblita based on semiochemical-induced expression alteration and gene silencing.

With the advent of next-generation sequencing, it is now possible to rapidly identify the entire repertoire of olfactory genes likely to be involved in chemical communication of an insect species. It remains, however, a challenge to identify olfactory proteins, such as odorant receptors and odorant-binding proteins (OBPs), vis-à-vis the odorants they detect. It has been reported that exposing the olfactory system to a physiologically relevant odorant alters the transcript levels of odorant receptor(s) involved in the detection of the tested odorant. We applied this paradigm in an attempt to identify putative OBPs from the scarab beetle Holotrichia oblita involved in the reception of plant-derived kairomones. Twenty-nine OBP genes were identified in the H. oblita transcriptome, 20 of which were enriched in antennae compared with nonolfactory tissues. Of these, 2 OBP genes, HoblOBP13 and HoblOBP9, were upregulated upon exposure to one of the female attractants (E)-2-hexenol and phenethyl alcohol; none of the OBP transcripts changed upon exposure to methyl anthranilate, which does not attract H. oblita females. Binding assays showed that HoblOBP13 and HoblOBP9 have high affinity for (E)-2-hexenol and phenethyl alcohol, respectively. RNAi treatment showed that transcripts of both HoblOBP13 and HoblOBP9 declined in a time-course manner 24-72 h postinjection. OBP-dsRNA-treated female beetles showed significantly lower attraction to (E)-2-hexenol and phenethyl alcohol than did water-injected beetles and those treated with GFP-dsRNA. We, therefore, concluded that HoblOBP13 and HoblOBP9 are essential for H. oblita reception of the plant-derived kairomones (E)-2-hexenol and phenethyl alcohol.

Identification and expression analysis of putative chemoreception genes from Cyrtorhinus lividipennis (Hemiptera: Miridae) antennal transcriptome.

Cyrtorhinus lividipennis Reuter (Hemiptera: Miridae) is an important egg predator of planthoppers which are destructive rice pests. The chemosensory genes in the mirid antennae play important roles in mating and prey-seeking behaviors. To gain a better understanding of the olfaction of C. lividipennis, we sequenced the antennal transcriptomes of the predator to identify the key olfaction genes. We identified 18 odorant binding proteins (OBPs), 12 chemosensory proteins (CSPs), 1 Niemann-Pick C2 protein (NPC2), 15 odorant receptors (ORs), 6 ionotropic receptors (IRs), 3 gustatory receptors (GRs) and 3 sensory neuron membrane proteins (SNMPs). Quantitative real-time PCR results showed that the relative transcript levels of three ClivORs (ClivOR6, 7 and 14) in the female antennae were 3 to 6 folds higher than that in the male antennae, indicating these genes were more related to oviposition site selection. The relative transcript levels of ClivCSP8 and ClivOR11 were 2.6 and 2.7 times higher in the male antennae than that of the female, respectively, indicating that these genes might be involved in mate searching. Moreover, the responses of dsorco treated predators to volatiles emitted from infested rice were significantly reduced, indicating these volatiles might serve as crucial cues in the host searching of C. lividipennis.

Virus-Mediated Overexpression of Vomeronasal Receptors and Functional Assessment by Live-Cell Calcium Imaging.

The mammalian vomeronasal organ (VNO) detects and transduces molecular cues emitted by other individuals that influence social behaviors such as mating and aggression. The detection of these chemosignals involves recognition of specific ligands by dedicated G protein-coupled receptors. Here, we describe recent methodological advances using a herpes virus-based amplicon delivery system to overexpress vomeronasal receptor genes in native, dissociated VNO neurons and to characterize corresponding cell responses to potential ligands through Ca2+ imaging. This methodology enables us to analyze the response patterns of single vomeronasal receptors to a large number of chemosensory stimuli.

A receptor-neuron correlate for the detection of attractive plant volatiles in Helicoverpa assulta (Lepidoptera: Noctuidae).

Plant volatiles are vital cues in the location of hosts for feeding and oviposition for Lepidoptera moths. The noctuid Helicoverpa assulta is a typical polyphagous moth, regarded as a good model for studying the olfactory reception of plant volatiles. In this study, four full-length genes encoding odorant receptors HassOR24, HassOR40, HassOR41, and HassOR55 expressed in antenna in H. assulta were functionally characterized. The highly expressed HassOR40 was narrowly tuned to a few structurally-related plant volatiles: geranyl acetate, geraniol and nerolidol. By systematically analyzing responses of single neuron in both trichoid sensilla and basiconic sensilla using single sensillum recording, the specific neuron B in one type of short trichoid sensilla was found to be mainly activated by the same chemicals as HassOR40 with high sensitivity, and with no significant difference between male and female neurons. Thus, a clear "receptor-neuron" relationship in H. assulta was demonstrated here, suggesting that HassOR40/HassOrco are expressed in neuron B of short trichoid sensilla. The active tobacco volatile nerolidol, recognized by this receptor-neuron line, elicits significant behavioral attraction of both sexes in H. assulta adults. The results indicate that we identified a receptor-neuron route for the peripheral coding of a behaviorally relevant host volatile in H. assulta.

Odorant-binding protein-based identification of natural spatial repellents for the African malaria mosquito Anopheles gambiae.

There is increasing interest in the development of effective mosquito repellents of natural origin to reduce transmission of diseases such as malaria and yellow fever. To achieve this we have employed an in vitro competition assay involving odorant-binding proteins (OBPs) of the malaria mosquito, Anopheles gambiae, with a predominantly female expression bias to identify plant essential oils (EOs) containing bioactive compounds that target mosquito olfactory function. EOs and their fractions capable of binding to such OBPs displayed repellence against female mosquitoes in a laboratory repellent assay. Repellent EOs were subjected to gas chromatographic analysis linked to antennogram (EAG) recordings from female A. gambiae to identify the biologically active constituents. Among these compounds cumin alcohol, carvacrol, ethyl cinnamate and butyl cinnamate proved as effective as DEET at an equivalent dose in the repellent assay, and combinations of carvacrol with either butyl cinnamate or cumin alcohol proved to be significantly more effective than DEET in the assay. When tested as spatial repellents in experimental shelters housing sleeping humans in northern Nigeria a binary mixture of carvacrol plus cumin alcohol caused mosquitoes to leave shelters in significantly higher numbers to those induced by DEET in female Anopheles spp. and in numbers equivalent to that of DEET in Culex spp. mosquitoes. These findings indicate an approach for the identification of biologically active molecules of natural origin serving as repellents for mosquitoes.

Nanodisc-Based Bioelectronic Nose Using Olfactory Receptor Produced in Escherichia coli for the Assessment of the Death-Associated Odor Cadaverine.

Cadaverine (CV), a death-associated odor, is an important target molecule for various sensor applications, including the evaluation of food spoilage. In this study, we developed an oriented nanodisc (ND)-functionalized bioelectronic nose (ONBN), based on carbon nanotube transistors and nanodiscs embedded with an olfactory receptor produced in Escherichia coli (E. coli) for detection of CV. To fabricate ONBN devices, a trace-amine-associated receptor 13c (TAAR13c) binding to CV was produced in E. coli, purified, reconstituted into NDs, and assembled, in the desired orientation, onto a carbon- nanotube-based field-effect transistor with floating electrodes. The ONBN showed high performance in terms of sensitivity and selectivity. Moreover, the ONBN was used to measure CV in diverse real-food samples for the determination of food freshness. These results indicate ONBN devices can be utilized to evaluate the quality of food samples quantitatively, which should enable versatile practical applications such as food safety and preservative development. Moreover, the ONBN could provide a useful tool for detection of corpses, which could be practically used in disaster responses.

Cooperative interactions enable singular olfactory receptor expression in mouse olfactory neurons.

The monogenic and monoallelic expression of only one out of >1000 mouse olfactory receptor (ORs) genes requires the formation of large heterochromatic chromatin domains that sequester the OR gene clusters. Within these domains, intergenic transcriptional enhancers evade heterochromatic silencing and converge into interchromosomal hubs that assemble over the transcriptionally active OR. The significance of this nuclear organization in OR choice remains elusive. Here, we show that transcription factors Lhx2 and Ebf specify OR enhancers by binding in a functionally cooperative fashion to stereotypically spaced motifs that defy heterochromatin. Specific displacement of Lhx2 and Ebf from OR enhancers resulted in pervasive, long-range, and trans downregulation of OR transcription, whereas pre-assembly of a multi-enhancer hub increased the frequency of OR choice in cis. Our data provide genetic support for the requirement and sufficiency of interchromosomal interactions in singular OR choice and generate general regulatory principles for stochastic, mutually exclusive gene expression programs.

Biophysical and functional characterization of the human olfactory receptor OR1A1 expressed in a mammalian inducible cell line.

Olfactory receptors (ORs) play a crucial role in detecting the odorant molecules present in the surrounding environment. These receptors, which belong to class A G-protein-coupled receptors, constitute the largest transmembrane protein family in the human genome. Functional studies showed that the OR family includes members that are able to respond to a large set of odorants and members that are activated by a relatively small number of related odorants. To understand the molecular mechanisms that govern the receptor-ligand interactions, we overexpressed the human OR hOR1A1 in a stable tetracycline-inducible HEK293S cell line. This receptor was engineered by inserting a C-terminal rho1D4 epitope tag and an N-terminal FLAG epitope tag to allow its purification and detection. The functional activity of the FLAG-rho1D4-tagged hOR1A1 in heterologous HEK293S cells was analysed using a real-time cAMP assay. A two-step purification using monoclonal anti-FLAG immunoaffinity purification and gel filtration was then employed to purify the detergent-solubilized receptor. A size exclusion chromatography-multi-angle light scattering analysis showed the presence of monomeric and dimeric forms of FLAG-rho1D4-tagged hOR1A1. The amounts of the monomeric and dimeric forms purified from sixty T175 flasks were approximately 1.6 and 1.1 mg, respectively. The circular dichroism analysis showed that the purified receptor was properly folded. Ligand binding was quantified using an intrinsic tryptophan fluorescence assay and revealed that the detergent-solubilized FLAG-rho1D4-tagged hOR1A1 bound its cognate odorant, dihydrojasmone, with an affinity in the micromolar range. These results pave the way for future crystallographic and NMR studies.

Olfactory receptor pseudo-pseudogenes.

Pseudogenes are generally considered to be non-functional DNA sequences that arise through nonsense or frame-shift mutations of protein-coding genes. Although certain pseudogene-derived RNAs have regulatory roles, and some pseudogene fragments are translated, no clear functions for pseudogene-derived proteins are known. Olfactory receptor families contain many pseudogenes, which reflect low selection pressures on loci no longer relevant to the fitness of a species. Here we report the characterization of a pseudogene in the chemosensory variant ionotropic glutamate receptor repertoire of Drosophila sechellia, an insect endemic to the Seychelles that feeds almost exclusively on the ripe fruit of Morinda citrifolia. This locus, D. sechellia Ir75a, bears a premature termination codon (PTC) that appears to be fixed in the population. However, D. sechellia Ir75a encodes a functional receptor, owing to efficient translational read-through of the PTC. Read-through is detected only in neurons and is independent of the type of termination codon, but depends on the sequence downstream of the PTC. Furthermore, although the intact Drosophila melanogaster Ir75a orthologue detects acetic acid-a chemical cue important for locating fermenting food found only at trace levels in Morinda fruit-D. sechellia Ir75a has evolved distinct odour-tuning properties through amino-acid changes in its ligand-binding domain. We identify functional PTC-containing loci within different olfactory receptor repertoires and species, suggesting that such 'pseudo-pseudogenes' could represent a widespread phenomenon.

A conditional Orco requirement in the somatic cyst cells for maintaining spermatids in a tight bundle in Drosophila testis.

Odorant receptors (OR) heterodimerizes with the OR co-receptor (Orco), forming specific odorant-gated cation channels, which are key to odor reception at the olfactory sensory neurons (OSN). Mammalian ORs are expressed in many other tissues, including testis. However, their biological implications are yet to be fully ascertained. In the mosquito, Orco is localized along the sperm tail and is indicated to maintain fidelity. Here, we show that orco expresses in Drosophila testis. The levels are higher in the somatic cyst cells. The orco-null mutants are perfectly fertile at 25 degree C. At 28 degree C, the coiled spermatid bundles are severely disrupted. The loss of Orco also disrupts the actin cap, which forms inside the head cyst cell at the rostral ends of the spermatid nuclei after coiling, and plays a key role in preventing the abnormal release of spermatids from the cyst enclosure. Both the defects are rescued by the somatic cyst cell-specific expression of the UAS-orco transgene. These results highlight a novel role of Orco in the somatic tissue during sperm release.

Achieving diverse and monoallelic olfactory receptor selection through dual-objective optimization design.

Multiple-objective optimization is common in biological systems. In the mammalian olfactory system, each sensory neuron stochastically expresses only one out of up to thousands of olfactory receptor (OR) gene alleles; at the organism level, the types of expressed ORs need to be maximized. Existing models focus only on monoallele activation, and cannot explain recent observations in mutants, especially the reduced global diversity of expressed ORs in G9a/GLP knockouts. In this work we integrated existing information on OR expression, and constructed a comprehensive model that has all its components based on physical interactions. Analyzing the model reveals an evolutionarily optimized three-layer regulation mechanism, which includes zonal segregation, epigenetic barrier crossing coupled to a negative feedback loop that mechanistically differs from previous theoretical proposals, and a previously unidentified enhancer competition step. This model not only recapitulates monoallelic OR expression, but also elucidates how the olfactory system maximizes and maintains the diversity of OR expression, and has multiple predictions validated by existing experimental results. Through making an analogy to a physical system with thermally activated barrier crossing and comparative reverse engineering analyses, the study reveals that the olfactory receptor selection system is optimally designed, and particularly underscores cooperativity and synergy as a general design principle for multiobjective optimization in biology.

Gene set of chemosensory receptors in the polyembryonic endoparasitoid Macrocentrus cingulum.

Insects are extremely successful animals whose odor perception is very prominent due to their sophisticated olfactory system. The main chemosensory organ, antennae play a critical role in detecting odor in ambient environment before initiating appropriate behavioral responses. The antennal chemosensory receptor genes families have been suggested to be involved in olfactory signal transduction pathway as a sensory neuron response. The Macrocentrus cingulum is deployed successfully as a biological control agent for corn pest insects from the Lepidopteran genus Ostrinia. In this research, we assembled antennal transcriptomes of M. cingulum by using next generation sequencing to identify the major chemosensory receptors gene families. In total, 112 olfactory receptors candidates (79 odorant receptors, 20 gustatory receptors, and 13 ionotropic receptors) have been identified from the male and female antennal transcriptome. The sequences of all of these transcripts were confirmed by RT-PCR, and direct DNA sequencing. Expression profiles of gustatory receptors in olfactory and non-olfactory tissues were measured by RT-qPCR. The sex-specific and sex-biased chemoreceptors expression patterns suggested that they may have important functions in sense detection which behaviorally relevant to odor molecules. This reported result provides a comprehensive resource of the foundation in semiochemicals driven behaviors at molecular level in polyembryonic endoparasitoid.