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. 2003 Apr 1;371(Pt 1):125–130. doi: 10.1042/BJ20021343

Granular gland transcriptomes in stimulated amphibian skin secretions.

Tianbao Chen 1, Susan Farragher 1, Anthony J Bjourson 1, David F Orr 1, Pingfan Rao 1, Chris Shaw 1
PMCID: PMC1223249  PMID: 12413397

Abstract

Amphibian defensive skin secretions are complex, species-specific cocktails of biologically active molecules, including many uncharacterized peptides. The study of such secretions for novel peptide discovery is time-limited, as amphibians are in rapid global decline. While secretion proteome analysis is non-lethal, transcriptome analysis has until now required killing of specimens prior to skin dissection for cDNA library construction. Here we present the discovery that polyadenylated mRNAs encoding dermal granular gland peptides are present in defensive skin secretions, stabilized by endogenous nucleic acid-binding amphipathic peptides. Thus parallel secretory proteome and transcriptome analyses can be performed without killing the specimen in this model amphibian system--a finding that has important implications in conservation of biodiversity within this threatened vertebrate taxon and whose mechanistics may have broader implications in biomolecular science.

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Selected References

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  1. Berger L., Speare R., Daszak P., Green D. E., Cunningham A. A., Goggin C. L., Slocombe R., Ragan M. A., Hyatt A. D., McDonald K. R. Chytridiomycosis causes amphibian mortality associated with population declines in the rain forests of Australia and Central America. Proc Natl Acad Sci U S A. 1998 Jul 21;95(15):9031–9036. doi: 10.1073/pnas.95.15.9031. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Blaustein A. R., Hoffman P. D., Hokit D. G., Kiesecker J. M., Walls S. C., Hays J. B. UV repair and resistance to solar UV-B in amphibian eggs: a link to population declines? Proc Natl Acad Sci U S A. 1994 Mar 1;91(5):1791–1795. doi: 10.1073/pnas.91.5.1791. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Cottingham I. R., Millar A., Emslie E., Colman A., Schnieke A. E., McKee C. A method for the amidation of recombinant peptides expressed as intein fusion proteins in Escherichia coli. Nat Biotechnol. 2001 Oct;19(10):974–977. doi: 10.1038/nbt1001-974. [DOI] [PubMed] [Google Scholar]
  4. Cruciani R. A., Barker J. L., Zasloff M., Chen H. C., Colamonici O. Antibiotic magainins exert cytolytic activity against transformed cell lines through channel formation. Proc Natl Acad Sci U S A. 1991 May 1;88(9):3792–3796. doi: 10.1073/pnas.88.9.3792. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Daly J. W. Thirty years of discovering arthropod alkaloids in amphibian skin. J Nat Prod. 1998 Jan;61(1):162–172. doi: 10.1021/np970460e. [DOI] [PubMed] [Google Scholar]
  6. De Perez G., Hindelang C. Ultrastructure of venom glands in the frog (Rana esculenta). Arch Anat Microsc Morphol Exp. 1985;74(3):216–227. [PubMed] [Google Scholar]
  7. Delfino G. Le ghiandole granulose cutanee di Alytes cisternasii Boscà e Discoglossus pictus Otth (Anfibi, Anuri, Discoglossidi): struttura, ultrastruttura e alcuni dati citochimici. Arch Ital Anat Embriol. 1979 Jan-Mar;84(1):81–106. [PubMed] [Google Scholar]
  8. Erspamer V., Melchiorri P., Falconieri Erspamer G., Montecucchi P. C., de Castiglione R. Phyllomedusa skin: a huge factory and store-house of a variety of active peptides. Peptides. 1985;6 (Suppl 3):7–12. doi: 10.1016/0196-9781(85)90343-2. [DOI] [PubMed] [Google Scholar]
  9. Goraya J., Wang Y., Li Z., O'Flaherty M., Knoop F. C., Platz J. E., Conlon J. M. Peptides with antimicrobial activity from four different families isolated from the skins of the North American frogs Rana luteiventris, Rana berlandieri and Rana pipiens. Eur J Biochem. 2000 Feb;267(3):894–900. doi: 10.1046/j.1432-1327.2000.01074.x. [DOI] [PubMed] [Google Scholar]
  10. Houlahan J. E., Findlay C. S., Schmidt B. R., Meyer A. H., Kuzmin S. L. Quantitative evidence for global amphibian population declines. Nature. 2000 Apr 13;404(6779):752–755. doi: 10.1038/35008052. [DOI] [PubMed] [Google Scholar]
  11. Jilek A., Engel E., Beier D., Lepperdinger G. Murine Bv8 gene maps near a synteny breakpoint of mouse chromosome 6 and human 3p21. Gene. 2000 Oct 3;256(1-2):189–195. doi: 10.1016/s0378-1119(00)00355-3. [DOI] [PubMed] [Google Scholar]
  12. Kiesecker J. M., Blaustein A. R., Belden L. K. Complex causes of amphibian population declines. Nature. 2001 Apr 5;410(6829):681–684. doi: 10.1038/35070552. [DOI] [PubMed] [Google Scholar]
  13. Kobayashi S., Hirakura Y., Matsuzaki K. Bacteria-selective synergism between the antimicrobial peptides alpha-helical magainin 2 and cyclic beta-sheet tachyplesin I: toward cocktail therapy. Biochemistry. 2001 Dec 4;40(48):14330–14335. doi: 10.1021/bi015626w. [DOI] [PubMed] [Google Scholar]
  14. Lai Ren, Liu Hen, Lee Wen Hui, Zhang Yun. A novel proline rich bombesin-related peptide (PR-bombesin) from toad Bombina maxima. Peptides. 2002 Mar;23(3):437–442. doi: 10.1016/s0196-9781(01)00642-8. [DOI] [PubMed] [Google Scholar]
  15. Lazarus L. H., Attila M. The toad, ugly and venomous, wears yet a precious jewel in his skin. Prog Neurobiol. 1993 Oct;41(4):473–507. doi: 10.1016/0301-0082(93)90027-p. [DOI] [PubMed] [Google Scholar]
  16. Melchiorri D., Bruno V., Besong G., Ngomba R. T., Cuomo L., De Blasi A., Copani A., Moschella C., Storto M., Nicoletti F. The mammalian homologue of the novel peptide Bv8 is expressed in the central nervous system and supports neuronal survival by activating the MAP kinase/PI-3-kinase pathways. Eur J Neurosci. 2001 May;13(9):1694–1702. doi: 10.1046/j.1460-9568.2001.01549.x. [DOI] [PubMed] [Google Scholar]
  17. Mollay C., Wechselberger C., Mignogna G., Negri L., Melchiorri P., Barra D., Kreil G. Bv8, a small protein from frog skin and its homologue from snake venom induce hyperalgesia in rats. Eur J Pharmacol. 1999 Jun 18;374(2):189–196. doi: 10.1016/s0014-2999(99)00229-0. [DOI] [PubMed] [Google Scholar]
  18. Moore K. S., Bevins C. L., Brasseur M. M., Tomassini N., Turner K., Eck H., Zasloff M. Antimicrobial peptides in the stomach of Xenopus laevis. J Biol Chem. 1991 Oct 15;266(29):19851–19857. [PubMed] [Google Scholar]
  19. Roseghini M., Falconieri Erspamer G., Severini C., Simmaco M. Biogenic amines and active peptides in extracts of the skin of thirty-two European amphibian species. Comp Biochem Physiol C. 1989;94(2):455–460. doi: 10.1016/0742-8413(89)90097-2. [DOI] [PubMed] [Google Scholar]
  20. Russo D. A., Petryk A., August C. S. Telomerase activity and phenotypic characterization in harvested bone marrow from a child with a germline cell cancer. Transplant Proc. 1997 Jun;29(4):2002–2002. doi: 10.1016/s0041-1345(97)00204-2. [DOI] [PubMed] [Google Scholar]
  21. Salmon A. L., Johnsen A. H., Bienert M., McMurray G., Nandha K. A., Bloom S. R., Shaw C. Isolation, structural characterization, and bioactivity of a novel neuromedin U analog from the defensive skin secretion of the Australasian tree frog, Litoria caerulea. J Biol Chem. 2000 Feb 18;275(7):4549–4554. doi: 10.1074/jbc.275.7.4549. [DOI] [PubMed] [Google Scholar]
  22. Seon A. A., Pierre T. N., Redeker V., Lacombe C., Delfour A., Nicolas P., Amiche M. Isolation, structure, synthesis, and activity of a new member of the calcitonin gene-related peptide family from frog skin and molecular cloning of its precursor. J Biol Chem. 2000 Feb 25;275(8):5934–5940. doi: 10.1074/jbc.275.8.5934. [DOI] [PubMed] [Google Scholar]
  23. Tyler M. J., Stone D. J., Bowie J. H. A novel method for the release and collection of dermal, glandular secretions from the skin of frogs. J Pharmacol Toxicol Methods. 1992 Dec;28(4):199–200. doi: 10.1016/1056-8719(92)90004-k. [DOI] [PubMed] [Google Scholar]
  24. Wachinger M., Kleinschmidt A., Winder D., von Pechmann N., Ludvigsen A., Neumann M., Holle R., Salmons B., Erfle V., Brack-Werner R. Antimicrobial peptides melittin and cecropin inhibit replication of human immunodeficiency virus 1 by suppressing viral gene expression. J Gen Virol. 1998 Apr;79(Pt 4):731–740. doi: 10.1099/0022-1317-79-4-731. [DOI] [PubMed] [Google Scholar]
  25. Wyman T. B., Nicol F., Zelphati O., Scaria P. V., Plank C., Szoka F. C., Jr Design, synthesis, and characterization of a cationic peptide that binds to nucleic acids and permeabilizes bilayers. Biochemistry. 1997 Mar 11;36(10):3008–3017. doi: 10.1021/bi9618474. [DOI] [PubMed] [Google Scholar]
  26. Zasloff M. Magainins, a class of antimicrobial peptides from Xenopus skin: isolation, characterization of two active forms, and partial cDNA sequence of a precursor. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5449–5453. doi: 10.1073/pnas.84.15.5449. [DOI] [PMC free article] [PubMed] [Google Scholar]

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