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. 2003 Jan;163(1):227–237. doi: 10.1093/genetics/163.1.227

amontillado, the Drosophila homolog of the prohormone processing protease PC2, is required during embryogenesis and early larval development.

Lowell Y M Rayburn 1, Holly C Gooding 1, Semil P Choksi 1, Dhea Maloney 1, Ambrose R Kidd 3rd 1, Daria E Siekhaus 1, Michael Bender 1
PMCID: PMC1462398  PMID: 12586710

Abstract

Biosynthesis of most peptide hormones and neuropeptides requires proteolytic excision of the active peptide from inactive proprotein precursors, an activity carried out by subtilisin-like proprotein convertases (SPCs) in constitutive or regulated secretory pathways. The Drosophila amontillado (amon) gene encodes a homolog of the mammalian PC2 protein, an SPC that functions in the regulated secretory pathway in neuroendocrine tissues. We have identified amon mutants by isolating ethylmethanesulfonate (EMS)-induced lethal and visible mutations that define two complementation groups in the amon interval at 97D1 of the third chromosome. DNA sequencing identified the amon complementation group and the DNA sequence change for each of the nine amon alleles isolated. amon mutants display partial embryonic lethality, are defective in larval growth, and arrest during the first to second instar larval molt. Mutant larvae can be rescued by heat-shock-induced expression of the amon protein. Rescued larvae arrest at the subsequent larval molt, suggesting that amon is also required for the second to third instar larval molt. Our data indicate that the amon proprotein convertase is required during embryogenesis and larval development in Drosophila and support the hypothesis that AMON acts to proteolytically process peptide hormones that regulate hatching, larval growth, and larval ecdysis.

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Selected References

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  1. Adams M. D., Celniker S. E., Holt R. A., Evans C. A., Gocayne J. D., Amanatides P. G., Scherer S. E., Li P. W., Hoskins R. A., Galle R. F. The genome sequence of Drosophila melanogaster. Science. 2000 Mar 24;287(5461):2185–2195. doi: 10.1126/science.287.5461.2185. [DOI] [PubMed] [Google Scholar]
  2. Anderson K. V., Jürgens G., Nüsslein-Volhard C. Establishment of dorsal-ventral polarity in the Drosophila embryo: genetic studies on the role of the Toll gene product. Cell. 1985 Oct;42(3):779–789. doi: 10.1016/0092-8674(85)90274-0. [DOI] [PubMed] [Google Scholar]
  3. Benjannet S., Rondeau N., Day R., Chrétien M., Seidah N. G. PC1 and PC2 are proprotein convertases capable of cleaving proopiomelanocortin at distinct pairs of basic residues. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3564–3568. doi: 10.1073/pnas.88.9.3564. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bergeron F., Leduc R., Day R. Subtilase-like pro-protein convertases: from molecular specificity to therapeutic applications. J Mol Endocrinol. 2000 Feb;24(1):1–22. doi: 10.1677/jme.0.0240001. [DOI] [PubMed] [Google Scholar]
  5. Brar B. K., Lowry P. J. The differential processing of proenkephalin A in mouse and human breast tumour cell lines. J Endocrinol. 1999 Jun;161(3):475–484. doi: 10.1677/joe.0.1610475. [DOI] [PubMed] [Google Scholar]
  6. Bryan P., Pantoliano M. W., Quill S. G., Hsiao H. Y., Poulos T. Site-directed mutagenesis and the role of the oxyanion hole in subtilisin. Proc Natl Acad Sci U S A. 1986 Jun;83(11):3743–3745. doi: 10.1073/pnas.83.11.3743. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Creemers J. W., Jackson R. S., Hutton J. C. Molecular and cellular regulation of prohormone processing. Semin Cell Dev Biol. 1998 Feb;9(1):3–10. doi: 10.1006/scdb.1997.0195. [DOI] [PubMed] [Google Scholar]
  8. Ewer J., Gammie S. C., Truman J. W. Control of insect ecdysis by a positive-feedback endocrine system: roles of eclosion hormone and ecdysis triggering hormone. J Exp Biol. 1997 Mar;200(Pt 5):869–881. doi: 10.1242/jeb.200.5.869. [DOI] [PubMed] [Google Scholar]
  9. Furuta M., Carroll R., Martin S., Swift H. H., Ravazzola M., Orci L., Steiner D. F. Incomplete processing of proinsulin to insulin accompanied by elevation of Des-31,32 proinsulin intermediates in islets of mice lacking active PC2. J Biol Chem. 1998 Feb 6;273(6):3431–3437. doi: 10.1074/jbc.273.6.3431. [DOI] [PubMed] [Google Scholar]
  10. Furuta M., Yano H., Zhou A., Rouillé Y., Holst J. J., Carroll R., Ravazzola M., Orci L., Furuta H., Steiner D. F. Defective prohormone processing and altered pancreatic islet morphology in mice lacking active SPC2. Proc Natl Acad Sci U S A. 1997 Jun 24;94(13):6646–6651. doi: 10.1073/pnas.94.13.6646. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gammie S. C., Truman J. W. Neuropeptide hierarchies and the activation of sequential motor behaviors in the hawkmoth, Manduca sexta. J Neurosci. 1997 Jun 1;17(11):4389–4397. doi: 10.1523/JNEUROSCI.17-11-04389.1997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Garofalo Robert S. Genetic analysis of insulin signaling in Drosophila. Trends Endocrinol Metab. 2002 May-Jun;13(4):156–162. doi: 10.1016/s1043-2760(01)00548-3. [DOI] [PubMed] [Google Scholar]
  13. Hwang J. R., Siekhaus D. E., Fuller R. S., Taghert P. H., Lindberg I. Interaction of Drosophila melanogaster prohormone convertase 2 and 7B2. Insect cell-specific processing and secretion. J Biol Chem. 2000 Jun 9;275(23):17886–17893. doi: 10.1074/jbc.M000032200. [DOI] [PubMed] [Google Scholar]
  14. Jiang N., Kolhekar A. S., Jacobs P. S., Mains R. E., Eipper B. A., Taghert P. H. PHM is required for normal developmental transitions and for biosynthesis of secretory peptides in Drosophila. Dev Biol. 2000 Oct 1;226(1):118–136. doi: 10.1006/dbio.2000.9832. [DOI] [PubMed] [Google Scholar]
  15. Li T., Bender M. A conditional rescue system reveals essential functions for the ecdysone receptor (EcR) gene during molting and metamorphosis in Drosophila. Development. 2000 Jul;127(13):2897–2905. doi: 10.1242/dev.127.13.2897. [DOI] [PubMed] [Google Scholar]
  16. Loi P. K., Emmal S. A., Park Y., Tublitz N. J. Identification, sequence and expression of a crustacean cardioactive peptide (CCAP) gene in the moth Manduca sexta. J Exp Biol. 2001 Aug;204(Pt 16):2803–2816. doi: 10.1242/jeb.204.16.2803. [DOI] [PubMed] [Google Scholar]
  17. McNabb S. L., Baker J. D., Agapite J., Steller H., Riddiford L. M., Truman J. W. Disruption of a behavioral sequence by targeted death of peptidergic neurons in Drosophila. Neuron. 1997 Oct;19(4):813–823. doi: 10.1016/s0896-6273(00)80963-0. [DOI] [PubMed] [Google Scholar]
  18. Mesce Karen A., Fahrbach Susan E. Integration of endocrine signals that regulate insect ecdysis. Front Neuroendocrinol. 2002 Apr;23(2):179–199. doi: 10.1006/frne.2002.0228. [DOI] [PubMed] [Google Scholar]
  19. Muller L., Lindberg I. The cell biology of the prohormone convertases PC1 and PC2. Prog Nucleic Acid Res Mol Biol. 1999;63:69–108. doi: 10.1016/s0079-6603(08)60720-5. [DOI] [PubMed] [Google Scholar]
  20. Rockwell N. C., Wang G. T., Krafft G. A., Fuller R. S. Internally consistent libraries of fluorogenic substrates demonstrate that Kex2 protease specificity is generated by multiple mechanisms. Biochemistry. 1997 Feb 18;36(7):1912–1917. doi: 10.1021/bi961779l. [DOI] [PubMed] [Google Scholar]
  21. Rouillé Y., Duguay S. J., Lund K., Furuta M., Gong Q., Lipkind G., Oliva A. A., Jr, Chan S. J., Steiner D. F. Proteolytic processing mechanisms in the biosynthesis of neuroendocrine peptides: the subtilisin-like proprotein convertases. Front Neuroendocrinol. 1995 Oct;16(4):322–361. doi: 10.1006/frne.1995.1012. [DOI] [PubMed] [Google Scholar]
  22. Rouillé Y., Westermark G., Martin S. K., Steiner D. F. Proglucagon is processed to glucagon by prohormone convertase PC2 in alpha TC1-6 cells. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3242–3246. doi: 10.1073/pnas.91.8.3242. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rulifson Eric J., Kim Seung K., Nusse Roel. Ablation of insulin-producing neurons in flies: growth and diabetic phenotypes. Science. 2002 May 10;296(5570):1118–1120. doi: 10.1126/science.1070058. [DOI] [PubMed] [Google Scholar]
  24. Schubiger M., Wade A. A., Carney G. E., Truman J. W., Bender M. Drosophila EcR-B ecdysone receptor isoforms are required for larval molting and for neuron remodeling during metamorphosis. Development. 1998 Jun;125(11):2053–2062. doi: 10.1242/dev.125.11.2053. [DOI] [PubMed] [Google Scholar]
  25. Seidah N. G., Benjannet S., Hamelin J., Mamarbachi A. M., Basak A., Marcinkiewicz J., Mbikay M., Chrétien M., Marcinkiewicz M. The subtilisin/kexin family of precursor convertases. Emphasis on PC1, PC2/7B2, POMC and the novel enzyme SKI-1. Ann N Y Acad Sci. 1999 Oct 20;885:57–74. doi: 10.1111/j.1749-6632.1999.tb08665.x. [DOI] [PubMed] [Google Scholar]
  26. Shennan K. I., Smeekens S. P., Steiner D. F., Docherty K. Characterization of PC2, a mammalian Kex2 homologue, following expression of the cDNA in microinjected Xenopus oocytes. FEBS Lett. 1991 Jun 24;284(2):277–280. doi: 10.1016/0014-5793(91)80703-6. [DOI] [PubMed] [Google Scholar]
  27. Siezen R. J., Leunissen J. A. Subtilases: the superfamily of subtilisin-like serine proteases. Protein Sci. 1997 Mar;6(3):501–523. doi: 10.1002/pro.5560060301. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Smeekens S. P., Avruch A. S., LaMendola J., Chan S. J., Steiner D. F. Identification of a cDNA encoding a second putative prohormone convertase related to PC2 in AtT20 cells and islets of Langerhans. Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):340–344. doi: 10.1073/pnas.88.2.340. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Sossin W. S., Fisher J. M., Scheller R. H. Cellular and molecular biology of neuropeptide processing and packaging. Neuron. 1989 May;2(5):1407–1417. doi: 10.1016/0896-6273(89)90186-4. [DOI] [PubMed] [Google Scholar]
  30. Steiner D. F., Rouillé Y., Gong Q., Martin S., Carroll R., Chan S. J. The role of prohormone convertases in insulin biosynthesis: evidence for inherited defects in their action in man and experimental animals. Diabetes Metab. 1996 Apr;22(2):94–104. [PubMed] [Google Scholar]
  31. Yoshida H., Ohagi S., Sanke T., Furuta H., Furuta M., Nanjo K. Association of the prohormone convertase 2 gene (PCSK2) on chromosome 20 with NIDDM in Japanese subjects. Diabetes. 1995 Apr;44(4):389–393. doi: 10.2337/diab.44.4.389. [DOI] [PubMed] [Google Scholar]
  32. Zhou A., Martin S., Lipkind G., LaMendola J., Steiner D. F. Regulatory roles of the P domain of the subtilisin-like prohormone convertases. J Biol Chem. 1998 May 1;273(18):11107–11114. doi: 10.1074/jbc.273.18.11107. [DOI] [PubMed] [Google Scholar]
  33. Zhou A., Webb G., Zhu X., Steiner D. F. Proteolytic processing in the secretory pathway. J Biol Chem. 1999 Jul 23;274(30):20745–20748. doi: 10.1074/jbc.274.30.20745. [DOI] [PubMed] [Google Scholar]
  34. Zhu X. L., Ohta Y., Jordan F., Inouye M. Pro-sequence of subtilisin can guide the refolding of denatured subtilisin in an intermolecular process. Nature. 1989 Jun 8;339(6224):483–484. doi: 10.1038/339483a0. [DOI] [PubMed] [Google Scholar]
  35. Zitnan D., Ross L. S., Zitnanova I., Hermesman J. L., Gill S. S., Adams M. E. Steroid induction of a peptide hormone gene leads to orchestration of a defined behavioral sequence. Neuron. 1999 Jul;23(3):523–535. doi: 10.1016/s0896-6273(00)80805-3. [DOI] [PubMed] [Google Scholar]

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