PRECURSOR POLYPROTEIN FOR MULTIPLE NEUROPEPTIDES SECRETED FROM THE SUBESOPHAGEAL GANGLION OF THE SILKWORM BOMBYX-MORI - CHARACTERIZATION OF THE CDNA-ENCODING THE DIAPAUSE HORMONE PRECURSOR AND IDENTIFICATION OF ADDITIONAL PEPTIDES

Peptidergic neurons, which serve as source of various endocrine neuropeptides, were identified in the suboesophageal ganglion (SG) and brain of insects. In the silkworm Bombyx mori, SG is known to secrete two neuropeptides, diapause hormone (DH) responsible for induction of embryonic diapause and pheromone biosynthesis-activating neuropeptide, which share a pentapeptide amide, Phe-Xaa-Pro-Arg-Leu-NH2 (Xaa = Gly or Ser), at the C terminus. We have isolated cDNA clones for DH from the cDNA library of SG by using oligonucleotide probes. The molecular characterization of the cDNA reveals that the mRNA encodes an open reading frame consisting of 192 aa residues in which the 24-aa DH peptide is localized at the N-terminal region just after the signal peptide. A homology search proposed that the cDNA encodes pheromone biosynthesis-activating neuropeptide and three other neuropeptides [alpha-, beta-, and gamma-SG neuropeptide (SGNP)] in the region following DH, all of which are flanked by possible tryptic cleavage sites and share the Phe-Xaa-Pro-Arg-Leu-Gly sequence at the C terminus. Northern hybridization analysis clearly showed that the gene expression was limited to SG. We chemically synthesized alpha-, beta-, and gamma-SGNP and used them to identify components in extracts of SG and to examine biological functions. Alpha- and gamma-SGNP were identified in extracts of SG, and the synthetic beta- and gamma-SGNP expressed weak DH activity. These results indicate that DH, along with four other neuropeptides, is generated from a common precursor polyprotein that is encoded by a single mRNA transcribed in neurosecretory cells of SG.