Abstract
Antisera to orcokinin B, CCHamide 1, and CCHamide 2 recognize enteroendocrine cells in the midgut of the fruitfly Drosophila melanogaster and its larvae. Although the antisera to CCHamide 1 and 2 are mutually cross-reactive, polyclonal mouse antisera raised to the C-terminals of their respective precursors allowed the identification of the two different peptides. In both larva and adult, CCHamide 2 immunoreactive endocrine cells are large and abundant in the anterior midgut and are also present in the anterior part of the posterior midgut. The CCHamide 2 immunoreactive endocrine cells in the posterior midgut are also immunoreactive with antiserum to allatostatin C. CCHamide 1 immunoreactivity is localized in endocrine cells in different regions of the midgut; those in the caudal part of the posterior midgut are identical with the allatostatin A cells. In the larva, CCHamide 1 enteroendocrine cells are also present in the endocrine junction and in the anterior part of the posterior midgut. Like in other insect species, the Drosophila orcokinin gene produces two different transcripts, A and B. Antiserum to the predicted biologically active peptide from the B-transcript recognizes enteroendocrine cells in both larva and adult. These are the same cells as those expressing β-galactosidase in transgenic flies in which the promoter of the orcokinin gene drives expression of this enzyme. In the larva, a variable number of orcokinin-expressing enteroendocrine cells are found at the end of the middle midgut, while in the adult, those cells are most abundant in the middle midgut, while smaller numbers are present in the anterior midgut. In both larva and adult, these cells also express allatostatin C. We also made a specific polyclonal antiserum to the NPF precursor in order to determine more precisely the expression of this peptide in the midgut. Using this antiserum, we find expression in the midgut to be the same as described previously using transgenic flies, while in the adult, midgut expression appears to be concentrated in the middle midgut, thus suggesting that in the anterior midgut only minor quantities of NPF are produced.
Similar content being viewed by others
References
Adkins AL, Moran AV (2007) Non-invasive serial blood collection in guinea pigs (Cavia porcellus). USAMRICD-07-02, U.S. Army Medical Research Institute
Brown MR, Crim JW, Arata RC, Cai HN, Chun C, Shen P (1999) Identification of a Drosophila brain-gut peptide related to the neuropeptide Y family. Peptides 20:1035–1042
Buchon N, Osman D, David FP, Fang HY, Boquete JP, Deplancke B, Lemaitre B (2013) Morphological and molecular characterization of adult midgut compartmentalization in Drosophila. Cell Rep 3:1725–1738
Bungart D, Kegel G, Burdzik S, Keller R (1995) Structure-activity relationships of the crustacean myotropic neuropeptide orcokinin. Peptides 16:199–204
Celniker SE, Dillon LA, Gerstein MB, Gunsalus KC, Henikoff S, Karpen GH, Kellis M, Lai EC, Lieb JD, MacAlpine DM, Micklem G, Piano F, Snyder M, Stein L, White KP, Waterston RH (2009) Unlocking the secrets of the genome. Nature 459:927–930
Chintapalli VR, Wang J, Dow JAT (2007) Using FlyAtlas to identify better Drosophila melanogaster models of human disease. Nat Genet 39:715–720
Dircksen H, Burdzik S, Sauter A, Keller R (2000) Two orcokinins and the novel octapeptide orcomyotropin in the hindgut of the crayfish Orconectes limosus: identified myostimulatory neuropeptides originating together in neurones of the terminal abdominal ganglion. J Exp Biol 203:2807–2818
Fusé M, Bendena WG, Donly BC, Tobe SS, Orchard I (1998) In situ hybridization analysis of leucomyosuppressin mRNA expression in the cockroach, Diploptera punctata. J Comp Neurol 395:328–341
Grimmelikhuijzen CJP, Graff D (1986) Isolation of pyroGlu-Gly-Arg-Phe-NH2 (Antho-RFamide), a neuropeptide from sea anemones. Proc Natl Acad Sci USA 83:9817–9821
Hansen KK, Hauser F, Williamson M, Weber SB, Grimmelikhuijzen CJP (2011) The Drosophila genes CG14593 and CG30106 code for G-protein-coupled receptors specifically activated by the neuropeptides CCH-amide-1 and CCH-amide-2. Biochem Biophys Res Commun 404:184–189
Hauser F, Cazzamali G, Williamson M, Park Y, Li B, Tanaka Y, Predel R, Neupert S, Schachtner J, Verleyen P, Grimmelikhuijzen CJP (2008) A genome-wide inventory of neurohormone GPCRs in the red flour beetle Tribolium castaneum. Front Neuroendocrinol 29:142–165
Hauser F, Neupert S, Williamson M, Predel R, Tanaka Y, Grimmelikhuijzen CJP (2010) Genomics and peptidomics of neuropeptides and protein hormones present in the parasitic wasp Nasonia vitripennis. J Proteome Res 9:5296–5310
Hofer S, Dircksen H, Tollbäck P, Homberg U (2005) Novel insect orcokinins: characterization and neuronal distribution in the brains of selected dicondylian insects. J Comp Neurol 490:57–71
Hummon AB, Richmond TA, Verleyen P, Baggerman G, Huybrechts J, Ewing MA, Vierstraete E, Rodriguez-Zas SL, Schoofs L, Robinson GE, Sweedler JV (2006) From the genome to the proteome: uncovering peptides in the Apis brain. Science 314:647–649
Huybrechts J, Bonhomme J, Minoli S, Prunier-Leterme N, Dombrovsky A, Abdel-Latief M, Robichon A, Veenstra JA, Tagu D (2010) Neuropeptide and neurohormone precursors in the pea aphid, Acyrthosiphon pisum. Insect Mol Biol 19(Suppl 2):87–95
Ida T, Takahashi T, Tominaga H, Sato T, Kume K, Ozaki M, Hiraguchi T, Maeda T, Shiotani H, Terajima S, Sano H, Mori K, Yoshida M, Miyazato M, Kato J, Murakami N, Kangawa K, Kojima M (2011a) Identification of the novel bioactive peptides dRYamide-1 and dRYamide-2, ligands for a neuropeptide Y-like receptor in Drosophila. Biochem Biophys Res Commun 410:872–877
Ida T, Takahashi T, Tominaga H, Sato T, Kume K, Yoshizawa-Kumagaye K, Nishio H, Kato J, Murakami N, Miyazato M, Kangawa K, Kojima M (2011b) Identification of the endogenous cysteine-rich peptide trissin, a ligand for an orphan G protein-coupled receptor in Drosophila. Biochem Biophys Res Commun 414:44–48
Ida T, Takahashi T, Tominaga H, Sato T, Sano H, Kume K, Ozaki M, Hiraguchi T, Shiotani H, Terajima S, Nakamura Y, Mori K, Yoshida M, Kato J, Murakami N, Miyazato M, Kangawa K, Kojima M (2012) Isolation of the bioactive peptides CCHamide-1 and CCHamide-2 from Drosophila and their putative role in appetite regulation as ligands for G protein-coupled receptors. Front Endocrinol (Lausanne) 3:177
Isabel G, Martin JR, Chidami S, Veenstra JA, Rosay P (2005) AKH-producing neuroendocrine cell ablation decreases trehalose and induces behavioral changes in Drosophila. Am J Physiol Regul Integr Comp Physiol 288(2):R531–R538
Johard HAD, Enell LE, Gustafsson E, Trifilieff P, Veenstra JA, Nässel DR (2008) Intrinsic neurons of Drosophila mushroom bodies express short neuropeptide F: Relations to extrinsic neurons expressing different neurotransmitters. J Comp Neurol 507:1479–1496
Kirkness EF, Haas BJ, Sun W, Braig HR, Perotti MA, Clark JM, Lee SH, Robertson HM, Kennedy RC, Elhaik E, Gerlach D, Kriventseva EV, Elsik CG, Graur D, Hill CA, Veenstra JA, Walenz B, Tubío JM, Ribeiro JM, Rozas J, Johnston JS, Reese JT, Popadic A, Tojo M, Raoult D, Reed DL, Tomoyasu Y, Kraus E, Mittapalli O, Margam VM, Li HM, Meyer JM, Johnson RM, Romero-Severson J, Vanzee JP, Alvarez-Ponce D, Vieira FG, Aguadé M, Guirao-Rico S, Anzola JM, Yoon KS, Strycharz JP, Unger MF, Christley S, Lobo NF, Seufferheld MJ, Wang N, Dasch GA, Struchiner CJ, Madey G, Hannick LI, Bidwell S, Joardar V, Caler E, Shao R, Barker SC, Cameron S, Bruggner RV, Regier A, Johnson J, Viswanathan L, Utterback TR, Sutton GG, Lawson D, Waterhouse RM, Venter JC, Strausberg RL, Berenbaum MR, Collins FH, Zdobnov EM, Pittendrigh BR (2010) Genome sequences of the human body louse and its primary endosymbiont provide insights into the permanent parasitic lifestyle. Proc Natl Acad Sci USA 107:12168–12173
LaJeunesse DR, Johnson B, Presnell JS, Catignas KK, Zapotoczny G (2010) Peristalsis in the junction region of the Drosophila larval midgut is modulated by DH31 expressing enteroendocrine cells. BMC Physiol 10:14
Li S, Torre-Muruzabal T, Søgaard KC, Ren GR, Hauser F, Engelsen SM, Pødenphanth MD, Desjardins A (2013) Grimmelikhuijzen CJP (2013) Expression patterns of the Drosophila neuropeptide CCHamide-2 and its receptor may suggest hormonal signaling from the gut to the brain. PLoS ONE 8:e76131. doi:10.1371/journal.pone.0076131
Liu F, Baggerman G, D’Hertog W, Verleyen P, Schoofs L, Wets G (2006) In silico identification of new secretory peptide genes in Drosophila melanogaster. Mol Cell Proteomics 5:510–522
Lu D, Lee KY, Horodyski FM, Witten JL (2002) Molecular characterization and cell-specific expression of a Manduca sexta FLRFamide gene. J Comp Neurol 446:377–396
Marianes A, Spradling AC (2013) Physiological and stem cell compartmentalization within the Drosophila midgut. Elife 27:e00886. doi:10.7554/eLife.00886
McQuilton P, St Pierre SE, Thurmond J, FlyBase Consortium (2012) FlyBase 101--the basics of navigating FlyBase. Nucleic Acids Res 40:D706–D714
O’Brien LE, Soliman SS, Li X, Bilder D (2011) Altered modes of stem cell division drive adaptive intestinal growth. Cell 147:603–614
Ons S, Richter F, Urlaub H, Pomar RR (2009) The neuropeptidome of Rhodnius prolixus brain. Proteomics 9:788–792
Park D, Veenstra JA, Park JH, Taghert PH (2008) Mapping peptidergic cells in Drosophila: where DIMM fits in. PLoS ONE 3(3):e1896. doi:10.1371/journal.pone.0001896
Pascual N, Castresana J, Valero ML, Andreu D, Bellés X (2004) Orcokinins in insects and other invertebrates. Insect Biochem Mol Biol 34:1141–1146
Radford JC, Davies SA, Dow JAT (2002) Systematic G-protein coupled receptor analysis in Drosophila melanogaster identifies a leucokinin receptor with novel roles. J Biol Chem 277:38810–38817
Reichwald K, Unnithan GC, Davis NT, Agricola H, Feyereisen R (1994) Expression of the allatostatin gene in endocrine cells of the cockroach midgut. Proc Natl Acad Sci USA 91:11894–11898
Reiher W, Shirras C, Kahnt J, Baumeister S, Isaac RE, Wegener C (2011) Peptidomics and peptide hormone processing in the Drosophila midgut. J Proteome Res 10:1881–1892
Roller L, Yamanaka N, Watanabe K, Daubnerová I, Žitňan D, Kataoka H, Tanaka Y (2008) The unique evolution of neuropeptide genes in the silkworm Bombyx mori. Insect Biochem Mol Biol 38:1147–1157
Sakai T, Satake H, Minakata H, Takeda M (2004) Characterization of crustacean cardioactive peptide as a novel insect midgut factor: isolation, localization, and stimulation of alpha-amylase activity and gut contraction. Endocrinology 145:5671–5678
Stangier J, Hilbich C, Burdzik S, Keller R (1992) Orcokinin: a novel myotropic peptide from the nervous system of the crayfish, Orconectes limosus. Peptides 13:859–864
Sterkel M, Oliveira PL, Urlaub H, Hernandez-Martinez S, Rivera-Pomar R, Sheila Ons S (2012) OKB, a novel family of brain-gut neuropeptides from insects. Insect Biochem Mol Biol 42:466–473
Tager HS (1976) Coupling of peptides to albumin with difluorodinitrobenzene. Anal Biochem 71:367–375
Veenstra JA (2000) Mono- and dibasic proteolytic cleavage sites in insect neuroendocrine peptide precursors. Arch Insect Biochem Physiol 43:49–63
Veenstra JA (2009) Peptidergic paracrine and endocrine cells in the midgut of the fruit fly maggot. Cell Tissue Res 336:309–323
Veenstra JA, Schooneveld H (1984) Immunocytochemical localization of neurons in the nervous system of the Colorado potato beetle with antisera against FMRFamide and bovine pancreatic polypeptide. Cell Tissue Res 235:303–308
Veenstra JA, Agricola HJ, Sellami A (2008) Regulatory peptides in fruit fly midgut. Cell Tissue Res 334:499–516
Wu Q, Wen T, Lee G, Park JH, Cai HN, Shen P (2003) Developmental control of foraging and social behavior by the Drosophila neurpeptide Y-like system. Neuron 39:147–161
Yamanaka N, Roller L, Zitnan D, Satake H, Mizoguchi A, Kataoka H, Tanaka Y (2011) Bombyx Orcokinins are brain-gut peptides involved in the neuronal regulation of ecdysteroidogenesis. J Comp Neurol 519:238–246
Žďárek J, Nachman RJ, Denlinger DL (2000) Parturition hormone in the tsetse Glossina morsitans: activity in reproductive tissues from other species and response of tsetse to identified neuropeptides and other neuroactive compounds. J Insect Physiol 46:213–219
Zhang Z, Schwartz S, Wagner L, Miller W (2000) A greedy algorithm for aligning DNA sequences. J Comput Biol 7:203–214
Zoephel J, Reiher W, Rexer KH, Kahnt J, Wegener C (2012) Peptidomics of the agriculturally damaging larval stage of the cabbage root fly Delia radicum (Diptera: Anthomyiidae). PLoS ONE 7(7):e41543. doi:10.1371/journal.pone.0041543
Acknowledgments
We thank Drs Yoshiaki Tanaka and Akira Mizoguchi for the generous gift of their mouse antiserum to Bombyx orcokinin A, Prof Cok Grimmelikhuijzen for the guinea pig FMRFamide antiserum, Dr Hans Smit for recovering it after 30 years, and Rafael Pineau for some of the mice that were used for making antisera. J.A.V. thanks an anonymous reviewer of a different manuscript for pointing him to the publication by Liu et al. The Drosophila community at large, as well as Institutional funding from the CNRS and the supply of different flies strains from the Bloomington Stock Center, is also gratefully acknowledged.
Author information
Authors and Affiliations
Corresponding author
Electronic supplementary material
Below is the link to the electronic supplementary material.
Fig. S1
(JPEG 5391 kb)
Rights and permissions
About this article
Cite this article
Veenstra, J.A., Ida, T. More Drosophila enteroendocrine peptides: Orcokinin B and the CCHamides 1 and 2. Cell Tissue Res 357, 607–621 (2014). https://doi.org/10.1007/s00441-014-1880-2
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00441-014-1880-2