Characteristics common to a cytokine family spanning five orders of insects.

Growth-blocking peptide (GBP) is a member of an insect cytokine family with diverse functions including growth and immunity controls. Members of this cytokine family have been reported in 15 species of Lepidoptera, and we have recently identified GBP-like peptides in Diptera such as Lucilia cuprina and Drosophila melanogaster, indicating that this peptide family is not specific to Lepidoptera. In order to extend our knowledge of this peptide family, we purified the same family peptide from one of the tenebrionids, Zophobas atratus,(1) isolated its cDNA, and sequenced it. The Z. atratus GBP sequence together with reported sequence data of peptides from the same family enabled us to perform BLAST searches against EST and genome databases of several insect species including Coleoptera, Diptera, Hymenoptera, and Hemiptera and identify homologous peptide genes. Here we report conserved structural features in these sequence data. They consist of 19-30 amino acid residues encoded at the C terminus of a 73-152 amino acid precursor and contain the motif C-x(2)-G-x(4,6)-G-x(1,2)-C-[KR], which shares a certain similarity with the motif in the mammalian EGF peptide family. These data indicate that these small cytokines belonging to one family are present in at least five insect orders.

Conserved repertoire of orthologous vomeronasal type 1 receptor genes in ruminant species.

BACKGROUND: In mammals, pheromones play an important role in social and innate reproductive behavior within species. In rodents, vomeronasal receptor type 1 (V1R), which is specifically expressed in the vomeronasal organ, is thought to detect pheromones. The V1R gene repertoire differs dramatically between mammalian species, and the presence of species-specific V1R subfamilies in mouse and rat suggests that V1R plays a profound role in species-specific recognition of pheromones. In ruminants, however, the molecular mechanism(s) for pheromone perception is not well understood. Interestingly, goat male pheromone, which can induce out-of-season ovulation in anestrous females, causes the same pheromone response in sheep, and vice versa, suggesting that there may be mechanisms for detecting "inter-species" pheromones among ruminant species. RESULTS: We isolated 23 goat and 21 sheep intact V1R genes based on sequence similarity with 32 cow V1R genes in the cow genome database. We found that all of the goat and sheep V1R genes have orthologs in their cross-species counterparts among these three ruminant species and that the sequence identity of V1R orthologous pairs among these ruminants is much higher than that of mouse-rat V1R orthologous pairs. Furthermore, all goat V1Rs examined thus far are expressed not only in the vomeronasal organ but also in the main olfactory epithelium. CONCLUSION: Our results suggest that, compared with rodents, the repertoire of orthologous V1R genes is remarkably conserved among the ruminants cow, sheep and goat. We predict that these orthologous V1Rs can detect the same or closely related chemical compound(s) within each orthologous set/pair. Furthermore, all identified goat V1Rs are expressed in the vomeronasal organ and the main olfactory epithelium, suggesting that V1R-mediated ligand information can be detected and processed by both the main and accessory olfactory systems. The fact that ruminant and rodent V1Rs have distinct features suggests that ruminant and rodent V1Rs have evolved distinct functions.

Xenopus V1R vomeronasal receptor family is expressed in the main olfactory system.

To date, over 100 vomeronasal receptor type 1 (V1R) genes have been identified in rodents. V1R is specifically expressed in the rodent vomeronasal organ (VNO) and is thought to be responsible for pheromone reception. Recently, 21 putatively functional V1R genes were identified in the genome database of the amphibian Xenopus tropicalis. Amphibians are the first vertebrates to possess a VNO. In order to determine at which point during evolution the vertebrate V1R genes began to function in the vomeronasal system, we analyzed the expression of all putatively functional V1R genes in Xenopus olfactory organs. We found that V1R expression was not detected in the VNO but was specifically detected in the main olfactory epithelium (MOE). We also observed that V1R-expressing cells in the MOE coexpressed Gi2, thus suggesting that the V1R-Gi2-mediated signal transduction pathway, which is considered to play an important role in pheromone reception in the rodent VNO, exists in the amphibian MOE. These results suggest that V1R-mediated signal transduction pathway functions in Xenopus main olfactory system.

Localization of the candidate genes ELOVL5 and SCD1 for 'male effect' pheromone synthesis in goats (Capra hircus).

The 'male effect' is a well-known phenomenon in female sheep and goats, whereby pheromone-induced activation of reproductive function occurs. In a previous study, we showed that the genes for elongation of long-chain fatty acids family member 5 (ELOVL5) and stearoyl-CoA desaturase 1 (SCD1) increased their expression significantly, concomitant with induction of pheromone synthesis. Therefore, these genes were considered to be prime candidate genes for pheromone synthesis. In the present study, we performed in situ hybridization to investigate where these two genes are expressed in goat skin. Strong positive signals were detected for both genes in the head skin of the male goat, which is the main site of pheromone production, and were mainly in the basal layer of the sebaceous gland cells, with the remaining cells showing negligible signals. None of the cells in the rump skin of the male goat or the head skin of the orchidectomized goat, neither of which produce pheromone, exhibited strong positive signals. The present study demonstrates that expression of these two candidate genes for pheromone synthesis is primarily localized in the sebaceous glands of the pheromone-producing skin region.

Stable knock-down of vomeronasal receptor genes in transgenic Xenopus tadpoles.

Xenopus V2R (xV2R), a family of G-protein-coupled receptors with seven transmembrane domains, is expressed in the Xenopus vomeronasal organ (VNO). There are six subgroups of xV2R, one of which, xV2RE, is predominantly expressed in the VNO. To understand the function of xV2R during VNO development, we developed a new method to achieve stable siRNA-suppression of the V2RE genes by introducing siRNA expression transgenes into the genomes of unfertilized eggs. We found that some of the derived transgenic tadpoles lacked VNOs and that their olfactory epithelium was fused. With the exception of one tadpole, expression of xV2RE was not detected in morphologically abnormal mutant tadpoles, although the olfactory marker protein and the olfactory receptors were expressed. These results suggest that we successfully produced transgenic tadpoles in which xV2RE expression was stably suppressed by siRNA, and that xV2RE plays a role in the morphogenesis of olfactory organs.

Rapid evolution of the male-specific antibacterial protein andropin gene in Drosophila.

Andropin, which encodes an antibacterial protein, is closely linked to the Cecropin gene cluster of D. melanogaster. Andropin and Cecropins are considered to have originated from one common ancestor. However, the expression pattern of Andropin is distinct from that of Cecropins, being restricted to the adult male ejaculatory duct. To elucidate the evolutionary process of Andropin, we have sequenced Andropin genes from D. melanogaster and its closely related species. In D. melanogaster, the nucleotide diversity of Andropin is remarkably low compared to that of Cecropin. In contrast, nonsynonymous substitutions of Andropin are conspicuously frequent between species. From genomic Southern analysis, Andropin-like genes are present in at least the melanogaster species subgroup. The series of present results suggests that Andropin was born in the course of constructing the Drosophila Cecropin gene family and then started to evolve rapidly, in contrast to Cecropins.