A Proteomic Investigation of Soluble Olfactory Proteins in Anopheles gambiae

Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) are small soluble polypeptides that bind semiochemicals in the lymph of insect chemosensilla. In the genome of Anopheles gambiae, 66 genes encode OBPs and 8 encode CSPs. Here we monitored their expression through classical proteomics (2D gel-MS analysis) and a shotgun approach. The latter method proved much more sensitive and therefore more suitable for tiny biological samples as mosquitoes antennae and eggs. Females express a larger number and higher quantities of OBPs in their antennae than males (24 vs 19). OBP9 is the most abundant in the antennae of both sexes, as well as in larvae, pupae and eggs. Of the 8 CSPs, 4 were detected in antennae, while SAP3 was the only one expressed in larvae. Our proteomic results are in fairly good agreement with data of RNA expression reported in the literature, except for OBP4 and OBP5, that we could not identify in our analysis, nor could we detect in Western Blot experiments. The relatively limited number of soluble olfactory proteins expressed at relatively high levels in mosquitoes makes further studies on the coding of chemical messages at the OBP level more accessible, providing for few specific targets. Identification of such proteins in Anopheles gambiae might facilitate future studies on host finding behavior in this important disease vector.     

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.     

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-…     

Construction and analysis of cDNA libraries from the antennae of male and female cotton bollworms Helicoverpa armigera (Hübner) and expression analysis of putative odorant-binding protein genes

Two high-quality cDNA libraries were constructed from female and male antennae of the cotton bollworm Helicoverpa armigera (Hübner). The titers were approximately 2.0 × 10⁶ pfu/ml for females and 2.3 × 10⁶ pfu/ml for males, and this complies with the test requirement. From the libraries, 1750 male ESTs and 1640 female ESTs were sequenced and further analyzed. We identified 15 olfactory genes (12 are new), and 14 of them have the characteristic six conserved cysteine residues. With the exception of OBP9, all the genes were classified as classical OBP genes. By alignment and cluster analysis, the 14 classical OBPs were divided into pheromone binding protein (PBP) genes, odorant binding protein (OBP) genes, general odorant binding protein 1 (GOBP1) genes, general odorant binding protein 2 (GOBP2) genes and antennae binding protein (ABP) genes. Among these genes, we obtained three PBP genes (PBP1–PBP3) including two new PBP genes, one new ABP gene, nine new OBP genes (OBP1–OBP9), one known GOBP1 gene and one known GOBP2 gene. Furthermore, the expression patterns of these 14 classical OBP genes were investigated in various tissues by real-time quantitative polymerase chain reaction (qPCR). The results indicated that some OBP genes are expressed differently in different sexes and tissues, but most of them are highly expressed in antennae.     

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.