Expression of odorant-binding proteins and chemosensory proteins in some Hymenoptera

https://doi.org/10.1016/j.ibmb.2005.01.002Get rights and content

Abstract

The expression of chemosensory proteins (CSPs) and odorant-binding proteins (OBPs) in individuals of different castes and ages have been monitored in three species of social hymenopterans, Polistes dominulus (Hymenoptera, Vespidae), Vespa crabro (Hymenoptera, Vespidae) and Apis mellifera (Hymenoptera, Apidae), using PCR with specific primers and polyclonal antibodies. In the paper wasp P. dominulus, OBP is equally expressed in antennae, wings and legs of all castes and ages, while CSP is often specifically present in antennae and in some cases also in legs. In the vespine species V. crabro CSP is antennal specific, while OBP is also expressed in legs and wings. The three CSPs and the five OBPs of A. mellifera show a complex pattern of expression, where both classes of proteins include members specifically expressed in antennae and others present in other parts of the body. These data indicate that at least in some hymenopteran species CSPs are specifically expressed in antennae and could perform roles in chemosensory perception so far assigned only to OBPs.

Introduction

Odorant-binding proteins (OBPs) and chemosensory proteins (CSPs) are two classes of small soluble proteins, that mediate, with mechanisms not completely clarified, chemical communication at the peripheral level (Vogt and Riddiford, 1981; Pelosi, 1998; Angeli et al., 1999; Sandler et al., 2000; Lartigue et al., 2002; Tegoni et al., 2004). These proteins are extremely concentrated in the lymph filling chemosensilla of antennae, tarsi and other chemoreception structures (Steinbrecht, 1998; Shanbhag et al., 2001). CSPs are also expressed in other parts of the body, such as specialised glands or secretory cells, probably involved in some cases in the delivery of semiochemicals (Jacquin-Joly et al., 2001).

Other functions for these proteins cannot be excluded and at least one member of the CSP family has been reported to be associated with regenerating limbs in the cockroach (Kitabayashi et al., 1998).

A great number of OBPs and CSPs have been identified and characterised at the protein level in species belonging to several orders of insects (McKenna et al., 1994; Pikielny et al., 1994; Tuccini et al., 1996; Maleszka and Stange, 1997; Wojtasek et al., 1998, Wojtasek et al., 1999; Picimbon and Leal, 1999; Vogt et al., 1999; Nagnan-Le Meillour et al., 2000; Marchese et al., 2000; Picimbon et al., 2000, Picimbon et al., 2001; Jacquin-Joly et al., 2001; Ishida et al., 2002a; Riviere et al., 2003; Ban et al., 2003a, Ban et al., 2003b).

In addition, hundreds of sequences have been obtained from cDNA or directly from genomes (Hekmat-Scafe et al., 2002; Xu et al., 2003; Zhou et al., 2004a; Wanner et al., 2004). Therefore, the presence in the genome of sequences encoding members of the OBP and CSP classes (Hekmat-Scafe et al., 2002; Xu et al., 2003; Zhou et al., 2004a) is not enough to conclude that their expression products could be involved in chemical communication.

The three-dimensional folding of four OBPs and one CSP have been resolved (Sandler et al., 2000; Horst et al., 2001; Lee et al., 2002; Lartigue et al., 2002; Campanacci et al., 2003; Mosbah et al., 2003; Kruse et al., 2003; Lartigue et al., 2004).

In social insects chemical communication is rather complex and can also mediate recognition between different castes or individuals of different nests. Such complexity is documented by a rich pheromonal language, making use of diverse chemical compounds, including long-chain hydrocarbons (Vander Meer et al., 1998).

It is still unkown whether a similar complexity also exists at the level of proteins (soluble polypeptides, such as OBPs and CSPs and membrane-bound receptors) involved in chemoreception. The genomes of Apis mellifera and Bombyx mori have recently been completed, thus allowing interesting comparisons between social and non-social species. However, a study at the protein level is still necessary to complement the genomic information.

So far, only few proteins of both classes have been reported in social insects. The most extensive study has been performed in the honeybee A. mellifera. Five genes encoding members of the OBP family (ASP1: Danty et al., 1999; ASP2: Briand et al., 2001; ASP4: AAL60417; ASP5: AAL60422; ASP6: AAL60421) and six encoding CSPs have been reported (ASP3c: Briand et al., 2002; W-AP1: Kamikouchi et al., 2004; BB17: BI510373; AmelCG4, AmelCG5, AmelCG6: Wanner et al., 2004). In the paper wasp Polistes dominulus we have identified one OBP and one CSP, but failed to detect sequences orthologous to those of the honeybee (Calvello et al., 2003). Several OBPs have also been reported in some ant species of the genus Solenopsis (Krieger and Ross, 2002) and a CSP in the argentine ant Linepithema humile (Ishida et al., 2002b). Interestingly, this last protein, as the CSP of P. dominulus, was found to be specific of antennae, whereas all previous studies in other insect species had shown that OBPs were antennal specific, while CSPs had been detected in different organs.

Despite such wide information at the structural level, functional data are still scarce and not conclusive. Ligand-binding experiments so far have failed in detecting clear-cut specificity of OBPs or CSPs towards semiochemicals of the species being examined, although better affinities have been reported in some cases with chemicals that are or could be active in chemical communication (Briand et al., 2002; Riviere et al., 2003; Calvello et al., 2003). Altered behaviour in insects lacking a specific OBP have been observed in two cases. In the first report, Drosophila mutants in which the gene expressing LUSH, a member of the OBP family, was deleted exhibited an altered behaviour towards ethanol (Kim et al., 1998). However, an alternative interpretation of such results has been proposed (Zhou et al., 2004b). The second case regards colonies of fire ants (Solenopsis invicta), where the lack of an OBP is apparently related with anomalous production of several queens in the colony (Krieger and Ross, 2002). For this OBP, that is synthesised in the thorax, a function in the correct release rather than in the perception of chemical signals seems more likely.

The distribution of OBPs and CSPs in different chemosensory structures and other parts of the body, as well as their expression during the insect's life cycle and, as far as social insects are concerned, in different castes and ranks, could provide useful information to the understanding of the physiological role of these soluble proteins in chemical communication.

Here we report the identification and cloning of a CSP in the wasp Vespa crabro and monitor the expression of both proteins in different organs and different castes of this species and of the paper wasp P. dominulus, using Western blot experiments. We also performed a similar investigation in the honeybee A. mellifera, using the five OBPs and the three CSPs described in the literature.

In Polistes wasps female castes are usually divided according to the appearance of the individuals in the various parts of the colony cycle. Workers are typically the first females which emerge from the newly founded nests and work on the colony without reproducing; foundresses are those females which emerge in an advanced colony phase, mate and hibernate and found a new nest the following year. No sharp differences exist in morphology between the two castes so that it is still unsure if a true pre-imaginal caste differentiation occurs in these insects. This is certain, on the other hand, in V. crabro and A. mellifera where the caste of an individual is determined at the larval level.

Section snippets

Insects

Individuals of P. dominulus, V. crabro and A. mellifera were selected according to caste and age. We collected and analysed “hibernating” females and active foundresses, later differentiated into active dominant and subordinate foundresses, as well as workers and males of P. dominulus; queens, males and workers of V. crabro; newly emerged workers, foragers, virgin queens, mated queens and drones of A. mellifera. All the specimens of P. dominulus and V. crabro came from different wild colonies.

Expression of OBP and CSP in P. dominulus

Previous investigation, using MALDI-TOF to analyse low molecular weight proteins in males and workers of the paper wasp, had indicated the specific expression of CSP in the antennae and the wider presence of OBP in antennae, wings and legs (Calvello et al., 2003). To further investigate this aspect, we have used antisera against recombinant P. dominulus OBP and CSP in Western blot experiments to monitor the expression of the two proteins in different parts of the body of individuals of

Discussion

The demonstration that olfactory receptors can respond to odorants and pheromones when expressed in foreign species (Wetzel et al., 2001; Dobritsa et al., 2003; Hallem et al., 2004) seems to leave no space for a role of soluble binding proteins (OBPs and CSPs) in insect chemoreception. However, olfactory receptors, when expressed in heterologous systems, exhibit kinetics both in the response and in the recovery phases, different from those measured in their natural environment. Therefore,

References (59)

  • J.F. Picimbon et al.

    Olfactory soluble proteins of cockroaches

    Insect. Biochem. Mol. Biol.

    (1999)
  • J.F. Picimbon et al.

    Chemosensory proteins of Locusta migratoria (Orthoptera: Acrididae)

    Insect Biochem. Mol. Biol.

    (2000)
  • J.F. Picimbon et al.

    Insect Biochem. Mol. Biol.

    (2001)
  • C.W. Pikielny et al.

    Members of a family of Drosophila putative odorant-binding proteins are expressed in different subsets of olfactory hairs

    Neuron

    (1994)
  • B.H. Sandler et al.

    Sexual attraction in the silkworm moth: structure of the pheromone-binding-protein–bombykol complex

    Chem. Biol.

    (2000)
  • M. Tegoni et al.

    Structural aspects of sexual attraction and chemical communication in insects

    Trends Biochem. Sci.

    (2004)
  • A. Tuccini et al.

    Putative odorant-binding protein in antennae and legs of Carausius morosus (Insecta, Phasmatodea)

    Insect Biochem. Mol. Biol.

    (1996)
  • H. Wojtasek et al.

    Attracted or repelled?—a matter of two neurons, one pheromone binding protein, and a chiral center

    Biochem. Biophys. Res. Commun.

    (1998)
  • H. Wojtasek et al.

    Identification and cloning of odorant binding proteins from the scarab beetle Phyllopertha diversa

    Biochem. Biophys. Res. Commun.

    (1999)
  • J.J. Zhou et al.

    “Plus-C” odorant-binding protein genes in two Drosophila species and the malaria mosquito Anopheles gambiae

    Gene

    (2004)
  • J.J. Zhou et al.

    Revisiting the odorant-binding protein LUSH of Drosophila melanogaster: evidence for odour recognition and discrimination

    FEBS Lett

    (2004)
  • S. Angeli et al.

    Purification, structural characterization, cloning and immunocytochemical localization of chemoreception proteins from Schistocerca gregaria

    Eur. J. Biochem.

    (1999)
  • L.P. Ban et al.

    Chemosensory proteins of Locusta migratoria

    Insect Mol. Biol.

    (2003)
  • L.P. Ban et al.

    Biochemical characterisation and bacterial expression of an odorant-binding protein from Locusta migratoria

    Cell. Mol. Life Sci.

    (2003)
  • L. Briand et al.

    Ligand binding and physico-chemical properties of ASP2, a recombinant odorant-binding protein from honeybee (Apis mellifera L.)

    Eur. J. Biochem.

    (2001)
  • L. Briand et al.

    Characterization of a chemosensory protein (ASP3c) from honeybee (Apis mellifera L.) as a brood pheromone carrier

    Eur. J. Biochem.

    (2002)
  • M. Calvello et al.

    Soluble proteins of chemical communication in the social wasp Polistes dominulus

    Cell. Mol. Life. Sci.

    (2003)
  • V. Campanacci et al.

    Moth chemosensory protein exhibits drastic conformational changes and cooperativity on ligand binding

    Proc. Natl. Acad. Sci. USA

    (2003)
  • F.R. Dani et al.

    Can cuticular lipids provide sufficient information for within-colony nepotism in wasps?

    Proc. R. Soc. London B

    (2003)
  • Cited by (0)

    View full text