Growth-blocking peptide expressed in the insect nervous system--cloning and functional characterization.

Abstract

Growth-blocking peptide (GBP) is an insect biogenic peptide that retards larval growth and, consequently, delays the onset of pupation of lepidopteran larvae. Transcription of the GBP gene has been proven to be regulated in a tissue-dependent manner by an approximately 0.8-kb mRNA transcript in fat body and 2.5-kb transcript in the brain central nervous system (CNS) [Hayakawa, Y., Ohnishi, A., Yamanaka, A., Izumi, S. & Tomino, S. (1995) FEBS Lett. 367, 185-189]. The GBP cDNA in the brain CNS was isolated, sequenced and found to be almost identical with that in the fat body, except that it encodes a long 5'-untranslated region (approximately 1.1 kb), thus indicating that the brain CNS is synthesized as a polyprotein precursor and released through posttranslational processing in the same manner as that predicted in the fat body. Sequence analyses of the genomic GBP gene fragments demonstrated that the 5'-untranslated region contains an intron encoding a sequence element, TGATAA, which has been reported to be important for expression in the insect larval fat body, suggesting that this element contributes to a tissue-specific expression of the shorter GBP mRNA in fat body. Ribonuclease protection assays showed that parasitization by a parasitic wasp elevated the expression of GBP mRNA in brain CNS. The parasitization-induced increase of GBP precursor and GBP concentrations was also demonstrated in brain CNS. Brain CNS dopamine was increased eightfold by incubating the tissue with GBP for 8 h through the elevation of dopa decarboxylase activity by a transcriptional enhancement perhaps due to GBP-induced increase of cytoplasmic Ca2+ concentration. Dopamine elevation reduced in vitro incorporation of methionine into proteins synthesized in brain CNS.