The mammary pheromone of the rabbit: from where does it come?

Newborn rabbits, Oryctolagus cuniculus, are directed to their mother's nipples by specialized odour cues. Previous investigations have suggested that these cues are released from the doe's abdominal surface from structures located around the nipple. We tested pups with samples of various cutaneous tissues or fluids collected from lactating females to determine the location of the source of the odour cues. After finding that the nipples from lactating does were more attractive than those of virgin females, we conducted three experiments using skin samples collected at increasing distance from the nipples, dermal and mammary tissues taken below the nipples, and milk collected at different lev…

Molecular cloning and expression of Tenebrio molitor ultraspiracle during metamorphosis and in vivo induction of its phosphorylation by 20-hydroxyecdysone.

Using a RT-PCR approach, the Tenebrio molitor homologue of Drosophila Ultraspiracle (TmUSP) was characterized. Its DNA binding domain shows a degree of identity with those of the other insect USPs. However, the ligand binding domain is closer to those of retinoid X receptors. Using an antibody raised against DmUSP, Western blot analysis of proteins from epidermis and other tissues revealed five immunoreactive bands, corresponding to different phosphorylated forms of a unique polypeptide, as shown by lambda-phosphatase treatment. The nuclear form of TmUSP seems unphosphorylated. An in vivo 20-hydroxyecdysone treatment increases considerably and rapidly the phosphorylated forms of TmUSP. This…

Identification, sequence and mRNA expression pattern during metamorphosis of a cDNA encoding a glycine-rich cuticular protein in Tenebrio molitor

The study of insect cuticular proteins and their sequences is of interest because they are involved in protein-protein and protein-chitin interactions which confer the mechanical properties and fine architecture of the cuticle. Moreover, in the coleopteran Tenebrio molitor there is a dramatic change in cuticular architecture between pre- and postecdysial secretion. We report the isolation, by differential screening, and the sequence characterization of a cDNA clone encoding a cuticular protein of T. molitor, ACP17. After insertion in the expression vector pEX1, the recognition of the fusion protein by an anti-cuticular monoclonal antibody confirmed the cuticular nature of ACP17. Northern hy…

The Drosophila ACP65A cuticle gene: deletion scanning analysis of cis-regulatory sequences and regulation by DHR38.

The regulatory sequences of the Drosophila ACP65A cuticle gene were analyzed in vivo in transgenic flies, using both fusion genes constructs and transposase-mediated deletions within a P element containing ACP65A regulatory sequences fused to the lacZ gene (deletion scanning). The sequences located between −594 and +161 are sufficient to confer both temporal and spatial expression specificities, indicating the presence of tissue-specific enhancers and response elements to hormone-induced factors. In addition, timing of expression and tissue-specificity appear to be controlled by distinct cis-regulatory elements, which suggests the existence of independent hormonal and tissue-specific signal…

Morphogenesis of the wing Anlagen in the mealworm beetle tenebrio molitor during the last larval instar.

The wing Anlagen of Tenebrio develop from epidermal cells located on the lateral margins of meso- and metathoraces. Three to four days after larval ecdysis, these cells start to proliferate slowly, continuing to do so until day 13 which corresponds to the period of the pupal commitment of the remaining epidermis. The wing Anlagen cells then proliferate rapidly until day 18.5. Three days before pupal ecdysis, the mitotic index falls suddenly while 40% of the Anlagen cells disappear owing to cell degeneration. The sudden changes observed in the mitotic index are correlated with two hemolymphatic peaks in ecdysteroid levels. Anlagen of the forewings and hindwings show similar development excep…

Molecular characterization of a new adult male putative calycin specific to tergal aphrodisiac secretion in the cockroach Leucophaea maderae

0014-5793 (Print) Journal Article Research Support, Non-U.S. Gov't; Lma-p18 is an epicuticular surface protein specific to the tergal gland aphrodisiac secretion of Leucophaea maderae adult males. Native Lma-p18 was purified and the complete cDNA sequence was determined by RT-PCR using primers based on Edman degradation fragments. Northern blot and in situ hybridization analyses showed that Lma-p18 is expressed exclusively in the anterior part of male tergal gland, which is exposed only during sexual behavior. Sequence analysis indicated that Lma-p18 belongs to the calycin superfamily and is very similar to Lma-p22, the first known male-specific tergal protein in L. maderae. Lma-p18 and Lma…

Development of the wing discs of Zophobas atratus under natural and experimental conditions: occurrence of a gradual larval-pupal commitment in the epidermis of tenebrionid beetles.

Using light and electron microscopy, we studied the development of the wing discs in the large beetle Zophobas atratus, under natural and experimental conditions. A reversible differentiation of the wing discs is usually observed during supernumerary instars of crowded larvae. Juvenile hormone analog (JHA) application during the wandering period or compelled experimental crowding during the larval-pupal switchover – or commitment – inhibits the onset of metamorphosis. Isolation, followed by recrowding, also induces the disc cells to secrete unusual cuticular material. Recrowding is able to trigger the reversal of metamorphosis during the 4-day period when larval-pupal commitment is taking p…

The precocious commitment of wing Anlagen in Tenebrio molitor revealed by the addition of 20-hydroxyecdysone.

Abstract An injection of 20-hydroxyecdysone (10 μg per animal) 6–13 days after the moult of the last larval instar of Tenebrio molitor induces the development of prothetelic larvae and larval-pupal intermediates. The state of larval-pupal switchover, or commitment, is only disclosed at the time of injection of the moulting hormone. Prothetelic A and B larvae, with small and medium sized wing Anlagen, undergo another larval or pupal instar. Prothetelic C larvae with bigger Anlagen are unable to moult, but the adult programme is expressed. Ecdysed larval-pupal intermediates give more or less perfect adults, while unecdysed mealworms, imprisoned in their larval cuticle, also expressed the adul…

Molecular characterization of Lma-p54, a new epicuticular surface protein in the cockroach Leucophaea maderae (Dictyoptera, oxyhaloinae)

0965-1748 (Print) Journal Article Research Support, Non-U.S. Gov't; The epicuticular surface protein Lma-p54 is imbedded in the "cuticular waxes" which cover the abdominal surface of the adult Leucophaea maderae. Natural Lma-p54 was purified and the complete cDNA sequence was determined by RT-PCR using primers based on Edman degradation fragments. Northern blot and in situ hybridization analyses showed that Lma-p54 was expressed in the adult abdominal epidermis and in the chemical sense organs of both sexes. Sequence alignment indicates that Lma-p54 is closely related to aspartic proteases (EC 3.4.23). However, there are critical amino acid substitutions at the level of the active site and,…

cDNA cloning and deduced amino acid sequence of a major, glycine-rich cuticular protein from the coleopteran Tenebrio molitor. Temporal and spatial distribution of the transcript during metamorphosis

0014-2956 (Print) Journal Article Research Support, Non-U.S. Gov't; In Coleoptera, the elytra (forewings), with a very hard and thick cuticle, protect the membranous and delicate hindwings against mechanical stress. We have isolated and characterized a cDNA encoding a major cuticle protein in Tenebrio molitor, named ACP-20. The deduced amino acid sequence is roughly tripartite, with two terminal glycine-rich domains and a central region showing pronounced similarities with some other hard cuticle proteins. Northern blot and in situ hybridization analyses reveal that ACP-20 gene expression is developmentally regulated since transcript accumulation occurs only in epidermal regions synthesizin…

A Novel Chitin-binding Protein from the Vestimentiferan Riftia pachyptila Interacts Specifically with β-Chitin

Abstract A cDNA from Riftia pachyptila was cloned. It encodes a novel 21.3-kDa protein from the worm protective tube, named RCBP (for Riftia chitin-binding protein). On the basis of partial tube-peptide sequences previously obtained, experiments using reverse transcriptase-mediated polymerase chain reaction and rapid amplification of cDNA ends led to the complete cDNA sequence. Analysis of its deduced amino acid sequence shows the presence of two chitin-binding domains. These domains are closely related to type 2 chitin-binding domains that are restricted to the animal kingdom. We showed by affinity assay and immunogold labeling that RCBP is the first protein so far known that binds specifi…

Characterization of a cDNA clone encoding a glycine-rich cuticular protein of Tenebrio molitor: developmental expression and effect of a juvenile hormone analogue

0962-1075 (Print) Journal Article; The complete sequence of a cDNA clone, isolated from epidermal mRNA of Tenebrio molitor using a monoclonal antibody raised against an adult-specific cuticular antigen only present in the hard cuticle, was obtained after primer extension at the 5' end. From this cDNA sequence, the deduced protein encompasses 199 amino acids (including a signal peptide) with a total molecular weight of 20.7 kDa. The protein exhibits a bipartite structure: glycine-rich region located in its NH2-terminal part and a carboxy-terminal domain sharing homologies with other cuticular proteins of Orthoptera, Diptera and Lepidoptera. In-situ hybridization analysis shows that the corre…

Peripheral olfactory function in rats after chemotherapeutic treatment with the anticancer drug docetaxel

International audience; Clinical studies have documented that chemotherapeutic cancer treatment in humans is often associated with weight loss and decreased enjoyment of food. Beside taste, olfaction plays a role in the food intake regulation. We assessed whether hemotherapeutic cancer treatment compromises olfactory function in rats treated with docetaxel (Taxotere, TAX), an antineoplastic drug which disrupts the structures necessary for cell survival and division. Electroolfactogram responses (EOG) can indicate morpho-pathological changes in olfactory epithelium. Male rats received either a single, two or three intravenous injections (one per week) of an estimated 10% lethal dose (LD10) o…

Characterization of an olfactory binding protein Xtro OBP in xenopus tropicalis in the aerial cavity

Cloning and Sequencing of a cDNA Encoding a Larval-Pupal-Specific Cuticular Protein in Tenebrio Molitor (Insecta, Coleoptera). Developmental Expression and Effect of a Juvenile Hormone Analogue

A cDNA clone encoding a larval-pupal cuticular protein, named TMLPCP-22, has been isolated by screening a library in expression vector with a monoclonal antibody made against pupal cuticular proteins of Tenebrio molitor. Northern-blot and in situ hybridization analyses showed that the expression of TMLPCP-22 is regulated in a stage-specific and tissue-specific manner; the transcript was present during the secretion of preecdysial larval and pupal cuticles and was restricted to epidermal cells. No expression was observed during adult cuticle deposition. In supernumerary pupae obtained after application of a juvenile hormone analogue, which is known to inhibit the adult programme, TMLPCP-22 m…

Specific expression of olfactory binding protein in the aerial olfactory cavity of adult and developing Xenopus

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Orchestin, a calcium-binding phosphoprotein, is a matrix component of two successive transitory calcified biomineralizations cyclically elaborated by a terrestrial crustacean.

Orchestia cavimana is a crustacean that cyclically replaces its calcified cuticle during molting cycles in order to grow. Its terrestrial way of life requires storage of calcium during each premolt period, as calcareous concretions, in tubular diverticula of the midgut. During the postmolt period the stored calcium is reabsorbed and is translocated through the storage organ epithelium as calcified small spherules. In a previous study, we sequenced and characterized a remarkable component of the organic matrix of the premolt storage structures, Orchestin, which is a calcium-binding phosphoprotein. In this paper, we analyzed the spatiotemporal expression of the orchestin gene by Northern blot…

Developmental profiles of epidermal mRNAs during the pupal-adult molt of Tenebrio molitor and isolation of a cDNA clone encoding an adult cuticular protein: effects of a juvenile hormone analogue

0012-1606 (Print) Journal Article; Changes in translatable mRNAs from the wing epidermis of the Coleoptera Tenebrio molitor have been investigated during metamorphosis by analysis of in vitro translated products. Striking differences between the patterns obtained from mRNAs extracted during pupal and adult cuticle secretion indicated that a drastic change in gene expression occurs during the pupal-adult transition. In addition to these stage-specific modifications, the mRNA patterns changed within each cuticular synthesis program (pupal or adult), especially at ecdysis. After tritiated leucine incorporation, some of the major radiolabeled cuticular proteins showed similar changes suggesting…

Structure, organization and expression of two clustered cuticle protein genes during the metamorphosis of an insect, Tenebrio molitor.

A 4-kb DNA segment of Tenebrio molitor (Insecta, Coleoptera) genomic DNA containing two larval-pupal cuticular genes has been cloned and sequenced. These genes, transcribed in opposite directions, are related in DNA sequence and the proteins encoded are very similar. Each of them contains a single intron located inside the sequence encoding the signal peptide, and a conserved sequence at -200 bp from the mRNA start position. These similarities in sequence suggest that these genes have evolved by duplication followed by diversification and that they are members of a family of genes with a common ancestry. They are the first example of clustered genes in Tenebrio molitor.

Characterization of two new cuticular genes specifically expressed during the post-ecdysial molting period in Tenebrio molitor

Abstract In a previous study, we have isolated a cDNA, TM-ACP17 , coding for a post-ecdysial adult protein of Tenebrio molitor . After screening of a genomic library with TM-ACP17 , we report isolation and sequencing of TM-ACP17 gene and a new gene, TM-LPCP29 , coding for a larval–pupal protein. These two genes exhibit a common sequence of 15 nucleotides and a characteristic of most cuticular protein genes so far described: an intron interrupting the signal peptide. The deduced aa sequence of TM-LPCP29 exhibits a high percentage of Ala (26.5%) and Val (17.5%) and is highly hydrophobic. In the N-terminal part, the motif VAAPV is repeated ten times. Numerous histidine residues are present in …

Variability of ecdysteroid-induced cell cycle alterations in Drosophila Kc sublines.

. The cell cycle of two lines isolated from Drosophila Kc cells was followed by flow cytofluorometry and cell counting. The first line is the 8-9K clone which grew in a medium supplemented with 5% serum; the second, named subline Kc0, grew in a serum-free medium. The stationary phase is characterized by a G2 cell accumulation: 73% in the 8-9K clone and 50% in the Kc0 subline. When the medium was supplemented with the steroid moulting hormone 20-hydroxyecdysone, more than 90% of 8-9K cells and 65% of Kc0 cells were progressively arrested in G2. In the continuous presence of 20-hydroxyecdysone, most of the 8-9K cells remain G2-arrested; no massive G2 release into M was observed and only a few…

Nucleotide sequence of an adult-specific cuticular protein gene from the beetle Tenebrio molitor: effects of 20-hydroxyecdysone on mRNA accumulation.

0962-1075 (Print) Journal Article Research Support, Non-U.S. Gov't; The accumulation of transcripts from two adult-specific cuticular genes (ACP-20 and ACP-22) is shown to be modified after addition of exogenous 20-hydroxyecdysone. In the continuous presence of high levels of the hormone, the expression of ACP-20 gene is significantly weaker than that of untreated controls, while ACP-22 expression is 2.5-fold increased. During active synthesis of the ACP messages, a 0.5 microg 20-hydroxyecdysone injection causes a rapid 2-fold increase in ACP-22 mRNA and is not able to repress ACP-20 mRNA accumulation. We conclude that these genes whose transcripts appear in an almost coordinated manner in …

Characterization of a cDNA encoding RP43, a CUB-domain-containing protein from the tube of Riftia pachyptila (Vestimentifera), and distribution of its transcript

A major 43kDa protein from the protective tube of Riftiapachyptila (Vestimentifera), named RP43, was partly microsequenced after isolation by SDS/PAGE from the protein fraction of tubes collected around the hydrothermal vents at the East Pacific Rise. On the basis of the partial peptide sequences obtained, experiments using reverse-transcriptase-mediated PCR and rapid amplification of cDNA ends led to the complete cDNA sequence. Analysis of deduced amino acid sequence of RP43 showed the presence of CUB domains (100–110-residue-spanning domains first reported in the complement subcomponents C1r/C1s, epidermal-growth-factor-related sea urchin protein and bone morphogenetic protein 1) that se…

Molecular characterization of a male-specific glycosyl hydrolase, Lma-p72, secreted on to the abdominal surface of the Madeira cockroach Leucophaea maderae (Blaberidae, Oxyhaloinae)

0264-6021 (Print) Journal Article Research Support, Non-U.S. Gov't; The epicuticular surface protein Lma-p72 is specific to the abdominal secretions of Leucophaea maderae (Madeira cockroach) adult males. Natural Lma-p72 was purified and the complete cDNA sequence determined by reverse-transcription PCR using primers based on Edman degradation fragments. Northern blot and in situ hybridization analyses showed that Lma-p72 was expressed in the tergal and sternal glands. Sequence alignment indicates that Lma-p72 is closely related to the family 1 glycosyl hydrolases (EC 3.2.1). Native Lma-p72 was proved to be active in the abdominal secretions and exhibit a beta-galactosidase-like activity. Ho…

Characterization of Urbain, a gene expressed during Bombyx mori imaginal wing discs morphogenesis

Cloning of Two Putative Ecdysteroid Receptor Isoforms from Tenebrio Molitor and their Developmental Expression in the Epidermis during Metamorphosis

Using the Drosophila EcR-B1 cDNA as a probe, we have cloned the putative ecdysteroid receptor from the mealworm Tenebrio molitor. We have isolated two cDNAs with different 5' termini that contain a complete open reading frame. These two cDNAs encode two proteins with distinct N-terminal regions corresponding to two isoforms. The coleopteran receptor is obviously related to the ecdysteroid receptor of other insects, but shares only 89% and 61% amino acid identities with the DNA-binding and ligand-binding domains of the Drosophila receptor, respectively. Its expression pattern has been examined in the epidermis during the last larval instar and pupal stage of T. molitor, in correlation with t…

Isolation and characterization of Urbain, a 20-hydroxyecdysone-inducible gene expressed during morphogenesis of Bombyx mori wing imaginal discs.

In insects, wing imaginal discs respond to the steroid hormone 20-hydroxyecdysone by initiating morphogenesis leading to the formation of the adult flight appendages. In this work we analyse the expression of a Bombyx gene, referred to as Urbain, whose cDNA had been previously isolated from wing discs (Chareyre et al. 1993). Accumulation of the 1.8 kb transcript occurs concomitantly with the increase of 20-hydroxyecdysone titer at every stage examined during post-embryonic development. In vitro, its accumulation is delayed 6–9 h after exposure to 20-hydroxyecdysone. Studies in the presence of cycloheximide have established that Urbain is a secondary response gene. The sequence of the mRNA c…

A pheromone-binding protein from the cockroach Leucophaea maderae: cloning, expression and pheromone binding

0264-6021 (Print) Journal Article; Odorant-binding proteins (OBPs) are thought to transport volatile compounds from air to their receptors through the sensillary lymph. In this protein family, the subgroup of pheromone-binding proteins (PBPs) is specifically tuned to the perception of the sexual pheromone. To date, the description of OBPs has been restricted to Endopterygota and Paraneoptera. Their expression in Orthopteroid has been hypothesized, but no evidence of OBP has been produced in this assemblage to date. In the present study, we describe the first OBP from a Dictyopteran insect that belongs to the cockroach Leucophaea maderae. The PBP of L. maderae (PBPLma) shares all the hallmar…