| Budget Amount *help |
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2006: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 2005: ¥2,300,000 (Direct Cost: ¥2,300,000)
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| Research Abstract |
Virgin females of the Japanese giant looper (Ascotis selenaria cretacea, Assc) in the family of Geometridae secrete C_19 3,4-epoxy-6,9-diene to attract males. Previous experiments have suggested that the epoxyalkenyl pheromone is biosynthesized from the corresponding 3,6,9-triene produced outside of a pheromone gland and transported to it via hemolymph after association with lipophorin. In order to clarify this transport, high-density lipophorin (HDLp) in the female moths showing two bands (apoLp I with ca.250 kDa and apoLp II with ca.80 kDa) on an SDS-PAGE was purified by KBr equilibrium density-gradient ultracentrifugation, and the association of the triene was confirmed by GC-MS analysis of a solvent extract from the isolated protein. Next, the role of HDLp was revealed by a topical application of the deuterated trienyl precursor to the abdomens of the females. The trienyl precursor was associated with HDLp. In their pheromone glands, the triene and the deuterated epoxy pheromone we
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re detected, indicating movement of the triene via the hemolymph. Experiments with male moths of A.s.cretacea and female moths of Bombyx moil showed the same association of HDLp with the triene topically applied. This result suggested that the adult females of A.s.cretacea did not develop HDLp specialized in the triene transport.
To regulate its biosynthesis in the pheromone gland, Assc females produce a pheromone biosynthesis-activating neuropeptide (PBAN) in the suboesophageal ganglion (SG), as do females in many lepidopteran species. We have isolated Assc-PBAN cDNA, which encodes 181 amino acids, including a PBAN homologue and four other putative peptides : a diapause hormone homologue, α-SG neuropeptide (SGNP), β-SGNP, and γ-SGNP, all of which shared an FXPR(K)L motif on their C-termini. Although PBANs with 30-35 amino acids have been characterized from 15 other species, the Assc-PBAN homologue consisted of 28 amino acids and showed low homology (< 46 %) compared with the others. Assc-β-SGNP with 8 amino acids was also shorter than the other β-SGNPs (16-22 amino acids). Furthermore, all of the known PBAN cDNAs have a GRR sequence between β-SGNP and PBAN as a cleavage site, but the Assc-PBAN cDNA showed an unusual GR sequence at the corresponding position, indicating the possibility of non-cleavage between the β-SGNP and PBAN. The chromatographic behavior of the natural pheromonotoropic peptide revealed the unique structure of Assc-PBAN including βSGNP, i.e., SVDFTPRLGRQLVDDVPQRQQIEEDRLGSRTRFFSPRL-NH_2, as the first determination of PBAN from the insects producing an epoxyalkenyl sex pheromone. Less
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