Isolation and primary structure of the eclosion hormone of the tobacco hornworm, Manduca sexta

Kataoka, Hiroshi; Troetschler, Ruth G.; Kramer, Steven J.; Cesarin, B J; Schooley, David A.

doi:10.1016/0006-291x(87)90592-4

Cited by 84 publications

(31 citation statements)

References 11 publications

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“…Although it appears to occur throughout the insects, biochemical and molecular analyses of this peptide have thus far been confined to large moths [7][8][9]11,13]. Our studies have now identified the EH gene from the dipteran, Drosophila melanogaster, and shown that this gene is expressed in a single pair of brain neurons which we presume are the homologues to the two pairs of EH-producing cells found in Lepidoptera.…”

Section: Discussionmentioning

confidence: 72%

Isolation, characterization and expression of the eclosion hormone gene of Drosophila melanogaster

Horodyski

Ewer

Riddiford

et al. 1993

European Journal of Biochemistry

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Eclosion hormone (EH) is a neuropeptide that triggers the performance of ecdysis behaviors at the end of a molt. We have isolated the EH gene from Drosophila melanogasfer, and localized the gene to the right arm of chromosome 3 at band position 90B1-2. The 97-amino-acid translation product contains a signal peptide followed by a 73-amino-acid prohormone. The N-terminus of the prohormone has diverged from lepidopteran EH both in its length and amino acid composition, and contains a potential endoproteolytic cleavage site. The deduced sequence of Drosophila EH is 58% identical (36 of 62 amino acids) to that of Manduca EH. The EH gene is expressed as a 0.8-kb transcript in a single pair of brain neurons which extend their processes the entire length of the central nervous system and also to the corpora cardiaca portion of the ring gland. These cells show massive depletion of immunoreactive EH at ecdysis.Over the last 10 years, work in invertebrates in general and insects in particular has led to identification of a large number of neuropeptides, many of which are expressed in a small number of cells within the central nervous system (CNS) [l, 21. These findings raise a number of important questions such as the role of these peptides in the normal physiology and development of the organism and the mechanisms that restrict peptide expression to a select set of cells within the CNS. To answer both questions, Drosophila melunogaster provides a number of distinct advantages. Mutants in a desired gene can be generated with relative ease and P-element transformation technology allows examination of regulatory regions that control peptide expression [3, 41. Consequently, we have turned to Drosophila to further our analysis of the neuropeptide eclosion hormone (EH), that triggers ecdysis of insects [5].Eclosion hormone was first found in moths [6] and later shown to be a 62-amino-acid peptide [7 -91 produced by two pairs of ventromedial neurosecretory cells in the brain [lO-131. In moths it has marked behavioral and developmental actions. It is known to act directly on the CNS to release the stereotyped motor programs that bring about the shedding of the old cuticle at the end of each molt [14]. Circulating peptide also acts on peripheral tissues to cause a diverse set of effects such as an increase in plasticity of wing cuticle [15], discharge of secretion by dermal glands [16], and the programmed degeneration of ecdysial muscles [17]. Yet the full range of EH actions are not known because it is not possible to block selectively either release or action of EH in moths. It appeared feasible to study EH in Drosophila because CNS extracts of both Calliphora [5] and Drosophila (Kimura and Truman, unpublished data) show EH activity in moth bioassays. Moreover, blood from ecdysing fleshflies, Sarcophaga bullata, stimulates premature ecdysis behavior in pharate (before ecdysis) flies [18]. The recent cloning of the EH gene from two species of moths, Munduca sexfa [ll] and Bombyx mori [13], has provided an avenue to search for t...

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Section: Discussionmentioning

confidence: 72%

Isolation, characterization and expression of the eclosion hormone gene of Drosophila melanogaster

Horodyski

Ewer

Riddiford

et al. 1993

European Journal of Biochemistry

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“…To isolate the gene that encodes Manduca EH, we designed a 72-nt-long probe based on the sequence of EH from the NH2 terminus to amino acid 24 (8,9) and codon bias data available for Drosophila genes (29,30) (Fig. 1).…”

Section: Resultsmentioning

confidence: 99%

“…Immediately thereafter is the NH2 terminus of the known sequence of EH. The translated sequence of the remainder of the precursor protein is identical to the complete sequence of EH (8,9). Therefore, the EH precursor consists of a 26-amino acid signal sequence followed by a single copy of the 62-amino acid neuropeptide.…”

Section: Resultsmentioning

confidence: 99%

“…The primary target of this peptide is the central nervous system, although other nonneural targets are known. Purification and sequencing of EH from two different moths, Manduca sexta (8,9) and Bombyx mori (10), has revealed a 62-amino acid peptide that shows 80% identity between the two species. This neuropeptide, however, has no similarity with any other known neuropeptide from either vertebrates or invertebrates (8).…”

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confidence: 99%

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Isolation and expression of the eclosion hormone gene from the tobacco hornworm, Manduca sexta.

Horodyski

Riddiford²,

Truman³

1989

Proc. Natl. Acad. Sci. U.S.A.

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Eclosion hormone (EH) is a 62-amino acid neuropeptide that initiates the ecdysis behavior of insects. The EH-encoding gene of the tobacco hornworm, Manduca sexta, was isolated by using a designed 72-mer oligonucleotide probe. Sequence analysis of this gene and its corresponding cDNA showed that the EH gene is 7.8 kilobases and consists of three exons. Exon I is totally nontranslated; exon II contains a 26-amino acid signal peptide and amino acids 1-4 of the EH peptide, and exon III encodes the remainder of the peptide. The EH gene is present in a single copy per haploid genome and transcribes an 0.8-kb mRNA that is expressed in larval, diapausing pupal, and developing adult brains but not in the ventral nerve cord or in nonneural tissues. In situ hybridization showed that the EH gene is expressed in two pairs of ventromedial neurosecretory cells in brains of both larvae and developing adults.As in vertebrates, insect neuropeptides are a diverse class of regulatory molecules that affect a number of physiological processes relating to growth, metamorphosis, reproduction, and homeostasis (1, 2). Despite their importance, the structures of relatively few of these neuropeptides are known; even less is known about the genes that code for them. Recent advances have been made in the molecular characterization and cloning of genes for the insect neuropeptides PheMet-Arg-Phe-NH2 (FMRFamide) (3, 4), bombyxin (5), and adipokinetic hormone (6).Growth and metamorphosis in insects is characterized by a series of molts when a new cuticle is produced. The shedding of the old cuticle at ecdysis is caused by the eclosion hormone (EH), a neuropeptide that triggers this behavior and directs associated physiological changes (7). The primary target of this peptide is the central nervous system, although other nonneural targets are known. Purification and sequencing of EH from two different moths, Manduca sexta (8, 9) and Bombyx mori (10), has revealed a 62-amino acid peptide that shows 80% identity between the two species. This neuropeptide, however, has no similarity with any other known neuropeptide from either vertebrates or invertebrates (8).Using an oligonucleotide based on this sequence, we now report the isolation and characterization of the EH genet.This gene proves to code only for the pre-EH molecule and to be expressed primarily in a set of four ventromedial cells in the brain of larvae and of developing adults.EXPERIMENTAL PROCEDURES Animals. Larvae of the tobacco hornworm, M. sexta, were reared individually on an artificial diet at 26°C under a 17-hr light:7-hr dark photoperiod as described (11). At the start of the wandering stage, the larvae were transferred to wooden blocks in which they pupated. Developing adults were transferred to a photoperiod of 12-hr light: 12-hr dark. Staging of developing adults was by days after pupal ecdysis, except for the final 5 days, which were determined by reference to a defined set of cuticular markers (12).Oligonucleotide Probe Construction and Labeling. To construct the 72 nu...

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“…Bactrocera dorsalis ͉ ecdysis ͉ oriental fruit fly ͉ cyclic GMP ͉ ecdysis triggering hormone E closion hormone (EH), an insect neuropeptide involved in shedding of old cuticle during ecdysis, was first described in the giant silk moth (1), and subsequently identified in Manduca sexta, Bombyx mori, and Drosophila melanogaster (2)(3)(4)(5). In Manduca, EH is produced by ventromedial (VM) cells in the brain and released into the hemolymph (6) in response to circulating ecdysis-triggering hormone (ETH) from Inka cells in epitracheal glands (7,8).…”

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confidence: 99%

Receptor guanylyl cyclases in Inka cells targeted by eclosion hormone

Chang

Yang

Adams

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

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A signature of eclosion hormone (EH) action in insect ecdysis is elevation of cGMP in Inka cells, leading to massive release of ecdysis triggering hormone (ETH) and ecdysis initiation. Although this aspect of EH-induced signal transduction is well known, the receptor mediating this process has not been identified. Here, we describe a receptor guanylyl cyclase BdmGC-1 and its isoform BdmGC-1B in the Oriental fruit fly Bactrocera dorsalis that are activated by EH. The B form exhibits the conserved domains and putative N-glycosylation sites found in BdmGC-1, but possesses an additional 46-amino acid insertion in the extracellular domain and lacks the C-terminal tail of BdmGC-1. Combined immunolabeling and in situ hybridization reveal that BdmGC-1 is expressed in Inka cells. Heterologous expression of BdmGC-1 in HEK cells leads to robust increases in cGMP following exposure to low picomolar concentrations of EH. The B-isoform responds only to higher EH concentrations, suggesting different physiological roles of these cyclases. We propose that BdmGC-1 and BdmGC-1〉 are high-and low-affinity EH receptors, respectively.

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Isolation and primary structure of the eclosion hormone of the tobacco hornworm, Manduca sexta

Cited by 84 publications

References 11 publications

Isolation, characterization and expression of the eclosion hormone gene of Drosophila melanogaster

Isolation, characterization and expression of the eclosion hormone gene of Drosophila melanogaster

Isolation and expression of the eclosion hormone gene from the tobacco hornworm, Manduca sexta.

Receptor guanylyl cyclases in Inka cells targeted by eclosion hormone

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