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Walter E. Bollenbacher - One of the best experts on this subject based on the ideXlab platform.
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The prothoracicotropic hormone (PTTH) of the commercial silkmoth, Bombyx mori, in the CNS of the tobacco hornworm, Manduca sexta.
Peptides, 1994Co-Authors: Rosemary S. Gray, David P. Muehleisen, Eva J. Katahira, Walter E. BollenbacherAbstract:Immunocytochemistry revealed that a Bombyx mori prothoracicotropic hormone (PTTH)-like peptide is expressed by the Manduca sexta big PTTH-producing neurons, the lateral neurosecretory cell group III (L-NSC III). Independent PCR of genomic DNA and a L-NSC III cDNA library yielded products with 99% sequence similarity to the cDNA encoding Bombyx PTTH. This similarity necessitated evaluation of the relationship between Manduca big PTTH and Bombyx PTTH by 1) bioassay of IEF separated Manduca PTTH and 2) direct assessment of Bombyx PTTH biological activity with Manduca prothoracic glands. Together, these studies indicate that Bombyx PTTH and Manduca PTTH are different peptides expressed by the L-NSC III. The possible physiological significance of a Bombyx PTTH-like peptide in Manduca and its coexpression with Manduca big PTTH by the L-NSC III are discussed.
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ultrastructure of prothoracic glands during larval pupal development of the tobacco hornworm Manduca sexta a reappraisal
Journal of Morphology, 1993Co-Authors: W K Hanton, R D Watson, Walter E. BollenbacherAbstract:The structure of Manduca sexta prothoracic glands was investigated using a protocol that preserves membranes. During the last larval stadium, prothoracic gland cells increase in diameter, volume, protein content, and perhaps number, enhancing their capacity to produce ecdysteroids. The glands' strand-of-cells morphology, their in situ location, the presence of gap junctions between cells, and junctional foot-like structures within cells support previous findings that prothoracicotropic hormone stimulates ecdysteroidogenesis via Ca2+-induced Ca2+ release. A different method of tissue fixation from that previously used to investigate the ultrastructure of Manduca sexta prothoracic glands has revealed a significantly different ultrastructure. These new findings begin to define roles for endoplasmic reticulum and mitochondria in ecdysteroid synthesis and support the hypothesis that the glands secrete the steroid hormone via exocytosis. The structural dynamics of the glands are discussed in the context of the glands' function during Manduca sexta larvalpupal development. © 1993 Wiley-Liss, Inc.
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Immunoaffinity purification of the neuropeptide prothoracicotropic hormone from Manduca sexta.
Peptides, 1993Co-Authors: David P. Muehleisen, Rosemary S. Gray, Eva J. Katahira, Mark K. Thomas, Walter E. BollenbacherAbstract:The prothoracicotropic hormones (PTTH) are cerebral peptides that control insect postembryonic development by stimulating the prothoracic glands to synthesize ecdysteroids. Using immunoaffinity chromatography and SDS-PAGE, a 25.5 kDa big PTTH has been purified from Manduca sexta. Based upon SDS-PAGE and Western blot analysis, the native form of big PTTH appears to be a dimer with monomers of 16.5 kDa. Four HPLC-separated fragments of this acidic peptide were sequenced and exhibited no sequence similarity with Bombyx mori PTTH. In agreement with this finding, the basic Bombyx PTTH had no PTTH bioactivity in Manduca. One sequenced fragment of the Manduca PTTH is approximately 70% similar to the vertebrate cellular retinoid binding proteins, suggesting these binding proteins may be present in insects.
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Developmental expression of the prothoracicotropic hormone in the CNS of the tobacco hornworm Manduca sexta.
The Journal of comparative neurology, 1993Co-Authors: Anne L. Westbrook, Sheila A. Regan, Walter E. BollenbacherAbstract:The prothoracicotropic hormone is an insect neuropeptide released into the hemolymph to signal molting and metamorphosis through its stimulation of steroidogenesis. The only known source of the prothoracicotropic hormone in the tobacco hornworm, Manduca sexta, has been a group of lateral cerebral neurosecretory cells, the L-NSC III. In this study, the developmental and spatial distribution of the prothoracicotropic hormone was examined throughout the life cycle of Manduca. In common with many vertebrates and invertebrates in which neuropeptides are located in several regions within the central nervous system (CNS), the prothoracicotropic hormone phenotype in Manduca is expressed by CNS neurons in addition to the L-NSC III. These neurons are located in the brain, frontal ganglion, and subesophageal ganglion. One cerebral neurosecretory cell group, the ventromedial neurons, expresses the prothoracicotropic hormone phenotype and the behavioral neurohormone, eclosion hormone. Whereas the L-NSC III and the ventromedial neurons express the peptide phenotype throughout the life cycle, the other neurons express the peptide only during the embryonic and larval stages. This precise spatial and temporal expression of the prothoracicotropic hormone by different groups of neurosecretory cells raises the possibility that in Manduca the peptide may, in addition to its known neuroendocrine function, play other physiological roles in different ways at different stages of the life cycle.
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Ultrastructure of prothoracic glands during larval-pupal development of the tobacco hornworm, Manduca sexta: a reappraisal
Journal of morphology, 1993Co-Authors: Walter E. BollenbacherAbstract:The structure of Manduca sexta prothoracic glands was investigated using a protocol that preserves membranes. During the last larval stadium, prothoracic gland cells increase in diameter, volume, protein content, and perhaps number, enhancing their capacity to produce ecdysteroids. The glands' strand-of-cells morphology, their in situ location, the presence of gap junctions between cells, and junctional foot-like structures within cells support previous findings that prothoracicotropic hormone stimulates ecdysteroidogenesis via Ca(2+)-induced Ca2+ release. A different method of tissue fixation from that previously used to investigate the ultrastructure of Manduca sexta prothoracic glands has revealed a significantly different ultrastructure. These new findings begin to define roles for endoplasmic reticulum and mitochondria in ecdysteroid synthesis and support the hypothesis that the glands secrete the steroid hormone via exocytosis. The structural dynamics of the glands are discussed in the context of the glands' function during Manduca sexta larval-pupal development.
Lawrence I Gilbert - One of the best experts on this subject based on the ideXlab platform.
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Manduca sexta prothoracicotropic hormone: evidence for a role beyond steroidogenesis
Archives of insect biochemistry and physiology, 2009Co-Authors: Robert Rybczynski, Chelsea A. Snyder, John Hartmann, Sho Sakurai, Lawrence I GilbertAbstract:Prothoracicotropic hormone (PTTH) is a homodimeric brain peptide hormone that positively regulates the production of ecdysteroids by the prothoracic gland of Lepidoptera and probably other insects. PTTH was first purified from heads of adult domestic silkworms, Bombyx mori. Prothoracic glands of Bombyx and Manduca sexta undergo apoptosis well before the adult stage is reached, raising the recurring question of PTTH function at these later stages. Because Bombyx has been domesticated for thousands of years, the possibility exists that the presence of PTTH in adult animals is an accidental result of domestication for silk production. In contrast, Manduca has been raised in the laboratory for only five or six decades. The present study found that Manduca brains contain PTTH at all stages examined post-prothoracic gland apoptosis, i.e., pharate adult and adult life, and that PTTH-dependent changes in protein phosphorylation and protein synthesis were observed in several reproductive and reproduction-associated organs. The data indicate that PTTH indeed plays a role in non-steroidogenic tissues and suggest possible future avenues for determining which cellular processes are being so regulated.
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identification characterization and developmental expression of halloween genes encoding p450 enzymes mediating ecdysone biosynthesis in the tobacco hornworm Manduca sexta
Insect Biochemistry and Molecular Biology, 2006Co-Authors: Kim F Rewitz, Robert Rybczynski, James T Warren, Lawrence I GilbertAbstract:The insect molting hormone 20-hydroxyecdysone (20E) plays a central role in regulating gene expression during development and metamorphosis. In many Lepidoptera, the pro-hormone 3-dehydroecdysone (3DE), synthesized from cholesterol in the prothoracic gland, is rapidly converted to ecdysone (E) by a hemolymph reductase, and E is subsequently converted to 20E in various peripheral target tissues. Recently, four Drosophila melanogaster P450 enzymes, encoded by specific Halloween genes, were cloned and functionally characterized as mediating the last hydroxylation steps leading to 20E. We extended this work to the tobacco hornworm Manduca sexta, an established model for endocrinological and developmental studies. cDNA clones were obtained for three Manduca orthologs of CYP306A1 (phantom; phm, the 25-hydroxylase), CYP302A1 (disembodied; dib, the 22-hydroxylase) and CYP315A1 (shadow; sad, the 2-hydroxylase), expressed predominantly in the prothoracic gland during the fifth (final) larval instar and during pupal-adult development, with fifth instar mRNA levels closely paralleling the hemolymph ecdysteroid titer. The data indicate that transcriptional regulation of phm, dib and sad plays a role in the developmentally varying steroidogenic capacities of the prothoracic glands during the fifth instar. The consistent expression of the Halloween genes confirms the importance of the prothoracic glands in pupal-adult development. These studies establish Manduca as an excellent model for examining the regulation of the Halloween genes.
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Developmental expression of Manduca shade, the P450 mediating the final step in molting hormone synthesis.
Molecular and cellular endocrinology, 2006Co-Authors: Kim F Rewitz, Robert Rybczynski, James T Warren, Lawrence I GilbertAbstract:The ecdysone 20-monooxygenase (E20MO; 20-hydroxylase) is the enzyme that mediates the conversion of ecdysone (E) to the active insect molting hormone, 20-hydroxyecdysone (20E), which coordinates developmental progression. We report the identification and developmental expression of the Halloween gene shade (shd; CYP314A1) that encodes the E20MO in the tobacco hornworm, Manduca sexta. Manduca Shd (MsShd) mediates the conversion of E to 20E when expressed in Drosophila S2 cells. In accord with the central dogma, the data show that Msshd is expressed mainly in the midgut, Malpighian tubules, fat body and epidermis with very low expression in the prothoracic gland and nervous system. Developmental variations in E20MO enzymatic activity are almost perfectly correlated with comparable changes in the gene expression of Msshd in the fat body and midgut during the fifth instar and the beginning of pupal-adult development. The results indicate three successive and overlapping peaks of expression in the fat body, midgut and Malpighian tubules, respectively, during the fifth larval instar. The data suggest that precise tissue-specific transcriptional regulation controls the levels, and thereby the activity, of the Manduca E20MO.
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Dynamic regulation of prothoracic gland ecdysteroidogenesis: Manduca sexta recombinant prothoracicotropic hormone and brain extracts have identical effects.
Insect biochemistry and molecular biology, 2000Co-Authors: Lawrence I Gilbert, Akira Mizoguchi, Qisheng Song, Wendy A. Smith, Shinji Nagata, Robert Rybczynski, R Morreale, H Matubayashi, M Shionoya, Hiroshi KataokaAbstract:Multiple assays were conducted in order to determine if the recently available recombinant prothoracicotropic hormone (rPTTH) from Manduca sexta is identical, or similar, to the natural hormone and if results from its use in a variety of assays confirm, or are inconsistent with, previous studies over the past 20years on PTTH action using brain extract. Brain extracts and rPTTH showed similar, if not identical, effects on the cell biology of Manduca prothoracic gland cells with the following results: increased levels of cAMP (adenosine 3':5' cyclic monophosphate) synthesis; requirement for extracellular Ca(2+) in in vitro studies; ecdysteroidogenesis stimulation in vitro; stimulation of general and specific protein synthesis; immunocytochemical identification of the two lateral cells in each brain hemisphere as the source of PTTH (the prothoracicotropes); the ability of antibodies to rPTTH to inhibit ecdysteroidogenesis stimulation in vitro; and the multiple phosphorylation of the ribosomal protein S6. The data revealed that brain extract and rPTTH show equivalent effects in all of the assays, indicating that this rPTTH is the natural PTTH of Manduca and that the data generated with brain extracts over the past two decades are indeed relevant.
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Molecular Cloning, Developmental Expression, and Phosphorylation of Ribosomal Protein S6 in the Endocrine Gland Responsible for Insect Molting
The Journal of biological chemistry, 1997Co-Authors: Qisheng Song, Lawrence I GilbertAbstract:Phosphorylation of ribosomal protein S6 is requisite for prothoracicotropic hormone (PTTH)-stimulated specific protein synthesis and subsequent ecdysteroidogenesis in the prothoracic glands of the tobacco hornworm, Manduca sexta. To better understand the role of S6 in regulating ecdysteroidogenesis, S6 cDNA was isolated from a Manduca prothoracic gland cDNA library and sequenced. The deduced protein is comprised of 253 amino acids, has a molecular weight of 29,038, and contains four copies of a 10-amino acid motif defining potential DNA-binding sites. This Manduca S6 possesses a consensus recognition sequence for the p70(s6k) binding domain as well as six seryl residues at the carboxyl-terminal sequence of 17 amino acids. Phosphoamino acid analysis revealed that the phosphorylation of Manduca prothoracic gland S6 is limited exclusively to serine residues. Although alterations in the quantity of S6 mRNA throughout the last larval instar and early pupal-adult development were not well correlated with the hemolymph ecdysteroid titer, developmental expression and phosphorylation of S6 were temporally correlated with PTTH release and the hemolymph ecdysteroid titer. These data provide additional evidence that S6 phosphorylation is a critical element in the transduction pathway leading to PTTH-stimulated ecdysteroidogenesis.
Nathan J. Tublitz - One of the best experts on this subject based on the ideXlab platform.
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Sequence and expression of the CAPA/CAP2b gene in the tobacco hawkmoth, Manduca sexta.
Journal of Experimental Biology, 2004Co-Authors: Poh Kheng Loi, Nathan J. TublitzAbstract:The gene coding for cardioacceleratory peptide 2b (CAP2b; pELYAFPRV) has been isolated and sequenced from the moth Manduca sexta (GenBank accession #AY649544). Because of its significant homology to the CAPA gene in Drosophila melanogaster, this gene is called the Manduca CAPA gene. The Manduca CAPA gene is 958 nucleotides long with 29 untranslated nucleotides from the beginning of the sequence to the putative start initiation site. The CAPA gene has a single open reading frame, 441 nucleotides long, that codes for a predicted precursor protein of 147 amino acids. The predicted prepropeptide encodes a single copy of each of three deduced propeptides, a CAP2b propeptide, with a Q substituted for an E at the N-terminus (QLYAFPRVa), and two novel CAP2b-related propeptides (DGVLNLYPFPRVa and TEGPGMWFGPRLa). To reduce confusion and to adopt a more standardized nomenclature, we rename pELYAFPRVa as Mas-CAPA-1 and assign the names of Mas-CAPA-2 to DGVLNLYPFPRVa and Mas-PK-1 (Pyrokinin-1) to TEGPGMWFGPRLa. The spatial and temporal expression pattern of the CAPA gene in the Manduca central nervous system (CNS) was determined in all major post-embryonic stages using in situ hybridization techniques. The CAPA gene is expressed in a total of 27 pairs of neurons in the post-embryonic Manduca CNS. A total of 16 pairs of cells is observed in the brain, two pairs in the sub-esophageal ganglion (SEG), one pair in the third thoracic ganglion (T3), one pair in each unfused abdominal ganglion (A1-A6) and two pairs in the fused terminal ganglion. The mRNA from the CAPA gene is present in nearly every ganglion in each post-embryonic stage. The number of cells expressing the CAPA gene varies during post-embryonic life, starting at 54 cells in first-instar larvae and declining to a minimum of 14 cells midway through adult development.
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Primary structure of a cardioactive neuropeptide from the tobacco hawkmoth, Manduca sexta
FEBS letters, 1992Co-Authors: Clement C. Cheung, Terry D. Lee, Poh Kheng Loi, Andy W. Sylwester, Nathan J. TublitzAbstract:The amino acid sequence of the first of a family of insect cardioregulatory peptides from the tobacco hawkmoth, Manduca sexta, has been determined using a combination of Edman degradation microsequencing and mass spectroscopy. This peptide contains 9 amino acid residues and an observed mass for the monoisotopic protonated molecule of 956.4 Da. There are two cysteines at positions 3 and 9 forming a disulfide bridge and the carboxyl-terminus is amidated. The structure of this peptide, Pro-Phe-Cys-Asn-Ala-Phe-Thr-Gly-Cys-NH2, is identical to a peptide recently isolated from crabs called crustacean cardioactive peptide (CCAP) and we propose that this peptide be named Manduca CCAP.
Lynn M. Riddiford - One of the best experts on this subject based on the ideXlab platform.
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insulin igf signaling regulates the change in commitment in imaginal discs and primordia by overriding the effect of juvenile hormone
Developmental Biology, 2008Co-Authors: Takashi Koyama, Lynn M. Riddiford, Maria O SyropyatovaAbstract:At the beginning of the final larval (fifth) instar of Manduca sexta, imaginal precursors including wing discs and eye primordia initiate metamorphic changes, such as pupal commitment, patterning and cell proliferation. Juvenile hormone (JH) prevents these changes in earlier instars and in starved final instar larvae, but nutrient intake overcomes this effect of JH in the latter. In this study, we show that a molecular marker of pupal commitment, broad, is up-regulated in the wing discs by feeding on sucrose or by bovine insulin or Manduca bombyxin in starved final instar larvae. This effect of insulin could not be prevented by JH. In vitro insulin had no effect on broad expression but relieved the suppression of broad expression by JH. This effect of insulin was directly on the disc as shown by its reduction in the presence of insulin receptor dsRNA. In starved penultimate fourth instar larvae, broad expression in the wing disc was not up-regulated by insulin. The discs became responsive to this action of insulin during the molt to the fifth instar together with the ability to become pupally committed in response to 20-hydroxyecdysone. Thus, the Manduca bombyxin acts as a metamorphosis-initiating factor in the imaginal precursors.
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Evolution of the Drosophila broad locus: the Manduca sexta broad Z4 isoform has biological activity in Drosophila.
Development genes and evolution, 2003Co-Authors: Cynthia A. Bayer, Lynn M. Riddiford, Xiaofeng Zhou, Baohua Zhou, Laurence Von KalmAbstract:The Drosophila melanogaster broad locus is essential for normal metamorphic development. Broad encodes three genetically distinct functions (rbp, br, and 2Bc) and a family of four zinc-finger DNA-binding proteins (Z1-Z4). The Z1, Z2, and Z3 protein isoforms are primarily associated with the rbp, br, and 2Bc genetic functions respectively. The Z4 protein isoform also provides some rbp genetic function, however an essential function for the Z4 isoform in metamorphosis has not been identified. To determine the degree of conservation of Z4 function between the tobacco hornworm Manduca sexta and Drosophila we generated transgenic Drosophila expressing the Manduca broad Z4 isoform and used this transgene to rescue rbp mutant lethality during Drosophila metamorphosis. We find that the Manduca Z4 protein has significant biological activity in Drosophila with respect to rescue of rbp-associated lethality. There was also some overlap in effects on cuticle gene expression between the Manduca Z4 and Drosophila Z1 isoforms that was not shared with the Drosophila Z4 isoform. Our findings show that Z4 function has been conserved over the 260-million-year period since the divergence of Diptera and Lepidoptera, and are consistent with the hypothesis that the Drosophila Z4 and Manduca Z4 isoforms have essential roles in metamorphosis.
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Molecular cloning of the prothoracicotropic hormone from the tobacco hornworm, Manduca sexta.
Insect biochemistry and molecular biology, 2003Co-Authors: M Shionoya, Lynn M. Riddiford, Shinji Nagata, Hideki Matsubayashi, Masako Asahina, Hisato Kuniyoshi, Hiroshi KataokaAbstract:A cDNA encoding a putative precursor of prothoracicotropic hormone (PTTH) from the tobacco hornworm, Manduca sexta, was isolated and sequenced. This clone contains an open reading frame encoding a 226-amino acid prepropeptide hormone. The deduced amino acid sequence is composed of a signal sequence, a precursor domain and a mature hormone and shows similarities to the other PTTHs that have been cloned from closely related lepidopteran species, Bombyx mori, Samia cynthia ricini, Antheraea peryni, and Hyalophora cecropia. Although these cDNAs showed slightly less similarities in predicted amino acid sequences, seven cysteine residues and the hydrophobic regions within those mature peptides were conserved. In situ hybridization using a cDNA probe encoding the Manduca PTTH showed that PTTH mRNA was in two pairs of neurosecretory cells in the Manduca brain. The recombinant putative ManducaPTTH produced in E. coli was biologically active, both causing a larval molt in neck-ligated Manduca 4th instar larvae (ED50 = 50 pM) and the adult molt of diapausing Manduca pupae (ED50 = 79 pM), but was unable to stimulate molting of debrained Bombyx pupae. 2003 Elsevier Ltd. All rights reserved.
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Isolation and embryonic expression of an abdominal-A-like gene from the lepidopteran, Manduca sexta.
Development (Cambridge England), 1991Co-Authors: Lisa M. Nagy, Ronald Booker, Lynn M. RiddifordAbstract:Using sequence homology to the Drosophila Antennapedia gene, we isolated a homeobox-containing gene from the lepidopteran, Manduca sexta. Sequence analysis and in situ hybridizations to tissue sections suggest that the Manduca gene encodes a lepidopteran homologue of the Drosophila Bithorax complex gene abdominal-A. The predicted amino acid sequence of a 76 amino acid region that includes the homeobox and the regions immediately flanking it are identical between the Manduca and Drosophila genes. Northern blots reveal that the Manduca abd-A gene is expressed first in the early embryo and continues to be expressed throughout later embryonic and larval stages. In situ hybridizations show that the posterior half of the first abdominal segment marks the anterior border of the Manduca abd-A expression. This expression pattern demonstrates the conservation of parasegments as domains of gene activity in the lepidopteran embryo. The Manduca abd-A expression extends from the posterior half of the first abdominal segment through the tenth abdominal segment, a domain that is greater than that of the Drosophila abd-A expression, and reflects the difference in visible segment number between the two insects.
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Juvenile hormone and “retinoic acid” receptors in Manduca epidermis
Insect Biochemistry, 1991Co-Authors: Subba Reddy Palli, Lynn M. Riddiford, Kiyoshi HirumaAbstract:Abstract Previous studies reviewed here have indicated that juvenile hormone (JH) specifically binds to a 29 kDa protein in epidermal nuclei from Manduca sexta larvae. Also, a 29 kDa nuclear protein that showed the same developmental pattern bound specifically to a larval cuticle gene LCP14. These results indicate that the 29 kDa nuclear protein is likely a JH receptor. Two retinoids (SRI 5942-64 and Ro 13-6298) were found to be weak JH mimics with ED 50 s 60–100 times higher than that of JH III in the black Manduca larval bioassay. A genomic clone ( Manduca “RAR”) then was isolated using the human retinoic acid receptor (hRAR) cDNA and the homologous region was sequenced. Thirteen out of 14 amino acids constituting the C-terminal half of the second zinc finger were identical in Manduca “RAR” and hRAR. Manduca “RAR” selected two mRNAs (3.8 and 4.5 kb) that are expressed at the peaks of the ecdysteroid titer during both the larval and the pupal molts, but not during the intermolt periods. When pieces of integument from day two 4th instar larvae were cultured with 4 × 10 −6 M 20-hydroxyecdysone (20HE), Manduca “RAR” mRNA increased 13–15-fold by 6 h, then decreased after 12 h in the continuous presence of 20HE. The presence of 3 × 10 −6 M JH slowed the rate of induction by 20HE. Thus, the “RAR” gene product is likely not the 29 kDa JH receptor but rather a transcriptional regulatory factor whose presence during a molt is modulated by JH.
Poh Kheng Loi - One of the best experts on this subject based on the ideXlab platform.
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Sequence and expression of the CAPA/CAP2b gene in the tobacco hawkmoth, Manduca sexta.
Journal of Experimental Biology, 2004Co-Authors: Poh Kheng Loi, Nathan J. TublitzAbstract:The gene coding for cardioacceleratory peptide 2b (CAP2b; pELYAFPRV) has been isolated and sequenced from the moth Manduca sexta (GenBank accession #AY649544). Because of its significant homology to the CAPA gene in Drosophila melanogaster, this gene is called the Manduca CAPA gene. The Manduca CAPA gene is 958 nucleotides long with 29 untranslated nucleotides from the beginning of the sequence to the putative start initiation site. The CAPA gene has a single open reading frame, 441 nucleotides long, that codes for a predicted precursor protein of 147 amino acids. The predicted prepropeptide encodes a single copy of each of three deduced propeptides, a CAP2b propeptide, with a Q substituted for an E at the N-terminus (QLYAFPRVa), and two novel CAP2b-related propeptides (DGVLNLYPFPRVa and TEGPGMWFGPRLa). To reduce confusion and to adopt a more standardized nomenclature, we rename pELYAFPRVa as Mas-CAPA-1 and assign the names of Mas-CAPA-2 to DGVLNLYPFPRVa and Mas-PK-1 (Pyrokinin-1) to TEGPGMWFGPRLa. The spatial and temporal expression pattern of the CAPA gene in the Manduca central nervous system (CNS) was determined in all major post-embryonic stages using in situ hybridization techniques. The CAPA gene is expressed in a total of 27 pairs of neurons in the post-embryonic Manduca CNS. A total of 16 pairs of cells is observed in the brain, two pairs in the sub-esophageal ganglion (SEG), one pair in the third thoracic ganglion (T3), one pair in each unfused abdominal ganglion (A1-A6) and two pairs in the fused terminal ganglion. The mRNA from the CAPA gene is present in nearly every ganglion in each post-embryonic stage. The number of cells expressing the CAPA gene varies during post-embryonic life, starting at 54 cells in first-instar larvae and declining to a minimum of 14 cells midway through adult development.
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Primary structure of a cardioactive neuropeptide from the tobacco hawkmoth, Manduca sexta
FEBS letters, 1992Co-Authors: Clement C. Cheung, Terry D. Lee, Poh Kheng Loi, Andy W. Sylwester, Nathan J. TublitzAbstract:The amino acid sequence of the first of a family of insect cardioregulatory peptides from the tobacco hawkmoth, Manduca sexta, has been determined using a combination of Edman degradation microsequencing and mass spectroscopy. This peptide contains 9 amino acid residues and an observed mass for the monoisotopic protonated molecule of 956.4 Da. There are two cysteines at positions 3 and 9 forming a disulfide bridge and the carboxyl-terminus is amidated. The structure of this peptide, Pro-Phe-Cys-Asn-Ala-Phe-Thr-Gly-Cys-NH2, is identical to a peptide recently isolated from crabs called crustacean cardioactive peptide (CCAP) and we propose that this peptide be named Manduca CCAP.