Evolution of noctuid pheromone binding proteins: identification of PBP in the black cutworm moth, Agrotis ipsilon.

Male black cutworm moths (Agrotis ipsilon, Lepidoptera, Noctuoidea, Noctuidae), which are attracted by a three-component pheromone blend ((Z)-7-dodecenyl acetate, Z7-12:Ac; (Z)-9-tetradecenyl acetate, Z9-14:Ac; (Z)-11-hexadecenyl acetate, Z11-16:Ac), express diverse antennal pheromone binding proteins (PBPs). Two PBP isoforms (Aips-1 and Aips-2) that show 46% identity were cloned from antennal cDNA of male A. ipsilon. The protein Aips-1 displays a high degree of identity (70-95%) with PBPs of other noctuiids, but shows only 42-65% identity with the PBPs of more phylogenetically distant species. The other protein, Aips-2, represents a distinct group of PBP that includes proteins from Sphingidae and Yponomeutidae. These differences observed suggest that each of the two PBPs may be tuned to a specific pheromone ligand.

Identity and expression pattern of chemosensory proteins in Heliothis virescens (Lepidoptera, Noctuidae).

Analyzing the chemosensory organs of the moth Heliothis virescens, three proteins belonging to the family of insect chemosensory proteins (CSPs) have been cloned; they are called HvirCSP1, HvirCSP2 and HvirCSP3. The HvirCSPs show about 50% identity between each other and 30-76% identity to CSPs from other species. Overall, they are rather hydrophilic proteins but include a conserved hydrophobic motif. Tissue distribution and temporal expression pattern during the last pupal stages were assessed by Northern blots. HvirCSP mRNAs were detected in various parts of the adult body with a particular high expression level in legs. The expression of HvirCSP1 in legs started early during adult development, in parallel with the appearance of the cuticle. HvirCSP1 mRNA was detectable five days before eclosion (day E-5), increased dramatically on day E-3 and remained at high level into adult life. The tissue distribution and the time course of appearance of HvirCSPs are in agreement with a possible role in contact chemosensation.

Purification and molecular cloning of chemosensory proteins from Bombyx mori.

Soluble low molecular weight acidic proteins are suspected to transport stimulus molecules to the sensory neurons within insect sensilla. From the antennae of Bombyx mori, we have purified and sequenced a protein (BmorCSP1) bearing sequence similarity to a class of soluble chemosensory proteins recently discovered in several orders of insects. Based on its N-terminal sequence, the cDNA encoding this protein has been amplified and cloned. Differential screening of a B. mori antennal cDNA library led to the identification of a second gene encoding a related protein (BmorCSP2), sharing 35-40% identity to BmorCSP1 and chemosensory proteins from other species. The predicted secondary structures of moth's, chemosensory proteins comprise alpha-helical foldings at conserved positions and a reduced hydrophobicity with respect to this novel family of chemosensory proteins.

Chemosensory proteins of Locusta migratoria (Orthoptera: Acrididae).

We describe a family of proteins abundantly expressed in the chemoreception sensory organs, the antennae and legs, of the desert locust, Locusta migratoria (Orthoptera, Acrididae). Using polymerase chain reaction-based approaches and homology screenings, "OS-D"-like proteins were identified in L. migratoria. The different sub-types (LmigOS-Ds) are very similar to each other and share about 50-70% identity with OS-Ds from Drosophila melanogaster and Periplaneta americana. A similar degree of identity was also observed with moth OS-Ds. Northern blot analysis revealed a strong expression of the LmigOS-Ds in the antennae and legs, suggesting their involvement in chemosensory processes. Despite the lack of direct evidence for their role in chemosensation, LmigOS-Ds and their homologs seem to constitute a large protein family, characterized by a striking abundance and diversity among insect chemosensory organs.

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

Wehave identified, cloned, and characterized two odorant binding proteins from the pale brown chafer, Phyllopertha diversa. One of the proteins (OBP1, 116 amino acids long) showed high amino acid identity (>90%) to two previously identified PBPs from scarab beetles. The second protein (OBP2) showed limited sequence similarity to lepidopteran and dipteran OBPs, but contained only 133 amino acids. Both proteins showed the occurrence of six highly conserved cysteines; electrospray mass spectral data suggested they are all bound in three disulfide bonds. During purification, OBP2 separated into several isoforms; N-terminal amino acid sequencing and electrospray ionization mass spectrometry demonstrated that they are different conformations of the same protein. In the native gel electrophoresis binding experiments, none of the OBPs bound 1, 3-dimethyl-2,4-(1H,3H)-quinazolinedione but different isoforms showed different binding affinities for (R)-japonilure, a pheromone from related scarab beetles, and bombykol, the pheromone from the silkworm moth, Bombyx mori. OBP1 also bound (R)-japonilure.