Novel odorant-binding proteins expressed in the taste tissue of the fly

A taste tissue cDNA library of the fleshfly Boettcherisca peregrina was screened with a subtracted cDNA probe enriched with taste-receptor-tissue-specific cDNA. Seven genes were identified with sequence similarity to insect odorant-binding protein (OBP) genes. The predicted amino acid sequences of the genes contain the putative signal peptide sequence at the N-terminal and most of them conserve the six cysteines common to known insect OBPs. These genes show a high degree of sequence divergence with approximately 20% amino acid identity. The most striking feature was that all seven of these genes are expressed mainly in the taste tissues, such as the labellum and tarsus, unlike the known insect OBP genes expressed in olfactory tissue. The predicted amino acid sequences had the highest degree of sequence similarity to the Drosophila melanogaster OBPs named pheromone binding protein-related proteins (PBPRPs). These gene products are here referred to as gustatory PBP-related proteins (GPBPRPs) 1-7. Homologous GPBPRP genes were found also in D. melanogaster by database search and are shown to be expressed in Drosophila taste tissues.     

Evolution of gene expression in the Drosophila olfactory system

Host plant shifts by phytophagous insects play a key role in insect evolution and plant ecology. Such shifts often involve major behavioral changes as the insects must acquire an attraction and/or lose the repulsion to the new host plant's odor and taste. The evolution of chemotactic behavior may be due, in part, to gene expression changes in the peripheral sensory system. To test this hypothesis, we compared gene expression in the olfactory organs of Drosophila sechellia, a narrow ecological specialist that feeds on the fruit of Morinda citrifolia, with its close relatives Drosophila simulans and Drosophila melanogaster, which feed on a wide variety of decaying plant matter. Using whole-genome microarrays and quantitative polymerase chain reaction, we surveyed the entire repertoire of Drosophila odorant receptors (ORs) and odorant-binding proteins (OBPs) expressed in the antennae. We found that the evolution of OR and OBP expression was accelerated in D. sechellia compared both with the genome average in that species and with the rate of OR and OBP evolution in the other species. However, some of the gene expression changes that correlate with D. sechellia's increased sensitivity to Morinda odorants may predate its divergence from D. simulans. Interspecific divergence of olfactory gene expression cannot be fully explained by changes in the relative abundance of different sensilla as some ORs and OBPs have evolved independently of other genes expressed in the same sensilla. A number of OR and OBP genes are upregulated in D. sechellia compared with its generalist relatives. These genes include Or22a, which likely responds to a key odorant of M. citrifolia, and several genes that are yet to be characterized in detail. Increased expression of these genes in D. sechellia may have contributed to the evolution of its unique chemotactic behavior.     

Identification of odorant-binding protein genes expressed in the antennae and the legs of the onion fly,Delia antiqua (Diptera: Anthomyiidae)

The onion fly, Delia antiqua (Meigen), is a pest specialized to the onion, Allium cepa L., and some other Allium plants. Host odorants play an important role in the attraction of D. antiqua adults and stimulation of oviposition in females. Odorant-binding proteins (OBPs) may serve as a first step in the perception of these chemical cues. In this study, to identify all OBP genes expressed in the chemosensory tissues in D. antiqua, RNA-seq analysis was carried out. In addition to the seven OBP genes previously identified, we found eight novel OBPs. Comparisons with Drosophila melanogaster Meigen OBP genes revealed that these 15 D. antiqua OBPs cover the structural variety observed in D. melanogaster OBPs, including Plus C and Minus C OBPs. These results suggest that a relatively large repertoire of chemosensory genes is maintained even in a specialist feeder.     

Molecular evolution of odorant-binding protein genes OS-E and OS-F in Drosophila

The Drosophila olfactory genes OS-E and OS-F are members of a family of genes that encode insect odorant-binding proteins (OBPs). OBPs are believed to transport hydrophobic odorants through the aqueous fluid within olfactory sensilla to the underlying receptor proteins. The recent discovery of a large family of olfactory receptor genes in Drosophila raises new questions about the function, diversity, regulation, and evolution of the OBP family. We have investigated the OS-E and OS-F genes in a variety of Drosophila species. These studies highlight potential regions of functional significance in the OS-E and OS-F proteins, which may include a region required for interaction with receptor proteins. Our results suggest that the two genes arose by an ancient gene duplication, and that in some lineages, one or the other gene has been lost. In D. virilis, the OS-F gene shows a different spatial pattern of expression than in D. melanogaster. One of the OS-F introns shows a striking degree of conservation between the two species, and we identify a putative regulatory sequence within this intron. Finally, a phylogenetic analysis places both OS-E and OS-F within a large family of insect OBPs and OBP-like proteins.     

Unique Features of Odorant-Binding Proteins of the Parasitoid Wasp Nasonia vitripennis Revealed by Genome Annotation and Comparative Analyses

Insects are the most diverse group of animals on the planet, comprising over 90% of all metazoan life forms, and have adapted to a wide diversity of ecosystems in nearly all environments. They have evolved highly sensitive chemical senses that are central to their interaction with their environment and to communication between individuals. Understanding the molecular bases of insect olfaction is therefore of great importance from both a basic and applied perspective. Odorant binding proteins (OBPs) are some of most abundant proteins found in insect olfactory organs, where they are the first component of the olfactory transduction cascade, carrying odorant molecules to the olfactory receptors. We carried out a search for OBPs in the genome of the parasitoid wasp Nasonia vitripennis and identified 90 sequences encoding putative OBPs. This is the largest OBP family so far reported in insects. We report unique features of the N. vitripennis OBPs, including the presence and evolutionary origin of a new subfamily of double-domain OBPs (consisting of two concatenated OBP domains), the loss of conserved cysteine residues and the expression of pseudogenes. This study also demonstrates the extremely dynamic evolution of the insect OBP family: (i) the number of different OBPs can vary greatly between species; (ii) the sequences are highly diverse, sometimes as a result of positive selection pressure with even the canonical cysteines being lost; (iii) new lineage specific domain arrangements can arise, such as the double domain OBP subfamily of wasps and mosquitoes.     

冈比亚按蚊嗅觉结合蛋白候选基因cDNA的克隆、鉴定及其表达型分析

疟蚊主要依靠嗅觉发现寄主。非洲疟蚊冈比亚按蚊Anopheles gambiae是一种嗜吸人血的疟疾传播媒介昆虫。该文作者基于其全基因组序列,采用RT-PCR和标准分子克隆技术获得2个嗅觉结合蛋白候选基因agLZ3788和agLZ9988。测序分析结果表明,它们具有嗅觉结合蛋白的标志性结构域。进一步采用半定量RT-PCR技术研究了它们的空间表达型,结果发现它们不但在雌蚊触角中表达,也在其他部位(尤其是蚊虫足部)有强的表达。这一发现说明疟蚊嗅觉结合蛋白可能具有更广的功能,也为进一步重组表达和功能研究提供了重要依据。     

A large family of divergent Drosophila odorant-binding proteins expressed in gustatory and olfactory sensilla.

We identified a large family of putative odorant-binding protein (OBP) genes in the genome of Drosophila melanogaster. Some of these genes are present in large clusters in the genome. Most members are expressed in various taste organs, including gustatory sensilla in the labellum, the pharyngeal labral sense organ, dorsal and ventral cibarial organs, as well as taste bristles located on the wings and tarsi. Some of the gustatory OBPs are expressed exclusively in taste organs, but most are expressed in both olfactory and gustatory sensilla. Multiple binding proteins can be coexpressed in the same gustatory sensillum. Cells in the tarsi that express OBPs are required for normal chemosensation mediated through the leg, as ablation of these cells dramatically reduces the sensitivity of the proboscis extension reflex to sucrose. Finally, we show that OBP genes expressed in the pharyngeal taste sensilla are still expressed in the poxneuro genetic background while OBPs expressed in the labellum are not. These findings support a broad role for members of the OBP family in gustation and olfaction and suggest that poxneuro is required for cell fate determination of labellar but not pharyngeal taste organs.     

Identification of odorant-binding protein genes from antennal expressed sequence tags of the onion fly, <Emphasis Type="Italic">Delia antiqua</Emphasis>

Insect odorant-binding proteins (OBPs) are thought to play a crucial role in the chemosensation of hydrophobic molecules such as pheromones and host chemicals. The onion fly, Delia antiqua, is a specialist feeder of Allium plants, and utilizes a host odorant n-dipropyl disulfide as a cue for its oviposition. Because n-dipropyl disulfide is a highly hydrophobic compound, some OBPs might be indispensable for perception of it. However, no OBP gene has been identified in D. antiqua. Here, to obtain the DNA sequences of D. antiqua OBPs, we performed an analysis of antennal expressed sequence tags (ESTs). Among 288 EST clones, eight D. antiqua OBP genes were identified for the first time. Phylogenetic analysis revealed that each D. antiqua OBP gene is more closely related to its Drosophila orthologs than to the other D. antiqua OBP genes, suggesting that these OBP genes had emerged before the divergence of Delia and Drosophila species. All of the eight D. antiqua OBPs are expressed not only in the antennae but also in the legs, suggesting additional roles in the taste perception of non-volatile compounds. These findings serve as an important basis for understanding the molecular mechanisms underlying the host adaptations of D. antiqua.     

Functional study of mammalian Neph proteins in Drosophila melanogaster

Neph molecules are highly conserved immunoglobulin superfamily proteins (IgSF) which are essential for multiple morphogenetic processes, including glomerular development in mammals and neuronal as well as nephrocyte development in D. melanogaster. While D. melanogaster expresses two Neph-like proteins (Kirre and IrreC/Rst), three Neph proteins (Neph1-3) are expressed in the mammalian system. However, although these molecules are highly abundant, their molecular functions are still poorly understood. Here we report on a fly system in which we overexpress and replace endogenous Neph homologs with mammalian Neph1-3 proteins to identify functional Neph protein networks required for neuronal and nephrocyte development. Misexpression of Neph1, but neither Neph2 nor Neph3, phenocopies the overexpression of endogenous Neph molecules suggesting a functional diversity of mammalian Neph family proteins. Moreover, structure-function analysis identified a conserved and specific Neph1 protein motif that appears to be required for the functional replacement of Kirre. Hereby, we establish D. melanogaster as a genetic system to specifically model molecular Neph1 functions in vivo and identify a conserved amino acid motif linking Neph1 to Drosophila Kirre function.     

Wake up and smell the pheromones

Odorant binding proteins (OBPs) are abundant proteins of unknown function expressed at high levels in insect and vertebrate chemosensory organs. In this issue of Neuron, Xu et al. show that Drosophila OBP76a is necessary for fruit flies to respond to the aggregation pheromone 11-cis vaccenyl acetate. The results suggest a mechanism by which this OBP is intimately involved in pheromone signal transduction.     

Intriguing similarities between two novel odorant-binding proteins of locusts

Two novel odorant-binding proteins (OBPs) of locust, LmigOBP2 and LmigOBP3 are very different from each other and from the previously reported LmigOBP1 in their amino acid sequences. Moreover, OBP3 contains three additional cysteines, a fact not previously recorded in standard length OBPs. However, these two proteins exhibit remarkably similar binding affinities to a set of organic compounds. Such behaviour is supported by three-dimensional models, showing very similar folding for LmigOBP2 and LmigOBP3, but clearly different for LmigOBP1. Also several amino acid residues lining the binding pockets of the three proteins appear conserved in LmigOBP2 and LmigOBP3, but not in LmigOBP1. Western blot experiments revealed the presence of LmigOBP2 in antennae, mouth parts and cerci, but could not detected LmigOBP3 in any of these tissues. In immunocytochemistry, antibodies against LmigOBP2 strongly stained the outer lymph of sensilla chaetica of the antennae, in contrast with LmigOBP1, previously reported in sensilla basiconica.     

Identification and expression profiling of putative odorant-binding proteins in the malaria mosquitoes, Anopheles gambiae and A.arabiensis

~~Identification and expression profiling of putative odorant-binding proteins in the malaria mosquitoes, Anopheles gambiae and A. arabiensis1. Curtis, C. F., Introduction 1: An overview of mosquito biology, behaviour and importance, in Olfaction in Mosquito-Host Interactions (eds. Bock, G. R.. Cardew, G.), New York: Wiley, 1996, 3-7. 2. Nighom, A., Hildebrand. J. G.. Dissecting the molecular mechanisms of olfaction in a malaria-vector mosquito, PNAS, 2002, 99(3): 1113-…     

Discrimination of alarm pheromone (E)-β-farnesene by aphid odorant-binding proteins

OBPs have been recently demonstrated to be required for odour perception in insects and directly involved in odour discrimination. In aphids they might represent new interesting targets for the control of their population in agriculture. Based on sequence information available in the EST database, we have cloned four genes encoding odorant-binding proteins (OBP) in Acyrthosiphon pisum and homologous genes in other aphid species. Unlike OBPs from other orders of insects, that are greatly divergent, in aphids these proteins have been found to be highly conserved, with differences between species limited to only few amino acid substitutions. On the contrary, similarities between OBP sequences of the same species are poor with 31% or less of identical amino acids. Three selected OBPs (OBP1, OBP3 and OBP8) have been expressed in bacteria and purified. Ligand-binding experiments have shown similar behaviour of the three proteins towards several organic compounds, but also some significant selectivities. In particular, (E)-β-farnesene, the alarm pheromone and its related compound farnesol exhibited good affinity to OBP3, but did not bind the other two proteins. We suggest that OBP3 could mediate response of aphids to the alarm pheromone.