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Marine Biotechnology

, Volume 16, Issue 3, pp 256–264 | Cite as

Conserved Mechanisms for Germ Cell-Specific Localization of nanos3 Transcripts in Teleost Species with Aquaculture Significance

  • Adrijana Škugor
  • Krasimir Slanchev
  • Jacob Seilø Torgersen
  • Helge Tveiten
  • Øivind AndersenEmail author
Original Article

Abstract

The importance of the aquaculture production is increasing with the declining global fish stocks, but early sexual maturation in several farmed species reduces muscle growth and quality, and escapees could have a negative impact on wild populations. A possible solution to these problems is the production of sterile fish by ablation of the embryonic primordial germ cells (PGCs), a technique developed in zebrafish. Cell-specific regulation of mRNA stability is crucial for proper specification of the germ cell lineage and commonly involves microRNA (miRNA)-mediated degradation of targeted mRNAs in somatic cells. This study reports on the functional roles of conserved motifs in the 3′ untranslated region (UTR) of the miRNA target gene nanos3 identified in Atlantic cod, Atlantic salmon, and zebrafish. The 3′UTR of cod nanos3 was sufficient for targeting the expression of green fluorescent protein (GFP) to the presumptive PGCs in injected embryos of the three phylogenetically distant species. 3′UTR elements of importance for PGC-specific expression were further examined by fusing truncated 3′UTR variants of cod nanos3 to GFP followed by injections in zebrafish embryos. The expression patterns of the GFP constructs in PGCs and somatic cells suggested that the proximal U-rich region is responsible for the PGC-specific stabilization of the endogenous nanos3 mRNA. Morpholino-mediated downregulation of the RNA-binding protein Dead end (DnD), a PGC-specific inhibitor of miRNA action, abolished the fluorescence of the PGCs in cod and zebrafish embryos, suggesting a conserved DnD-dependent mechanism for germ cell survival and migration.

Keywords

Primordial germ cells Dead end Nanos3 miR-430 Gadus morhua 

Notes

Acknowledgments

We thank Anne Grethe Hestnes for excellent assistance with microscopy and Hanne Johnsen for preparing cod embryos for WISH analysis. Salmon and cod miR-430 sequences were kindly provided by Julian Hamfjord and Steinar Johansen. The study was financially supported by the Norwegian Research Council (project number 190371).

References

  1. Bailey TL, Elkan C (1994) Fitting a mixture model by expectation maximization to discover motifs in biopolymers. Proc Int Conf Intell Syst Mol Biol 2:28–36PubMedGoogle Scholar
  2. Bizuayehu TT, Babiak J, Norberg B, Fernandes JM, Johansen SD, Babiak I (2012) Sex-biased miRNA expression in Atlantic halibut (Hippoglossus hippoglossus) brain and gonads. Sex Dev 6:257–266CrossRefPubMedGoogle Scholar
  3. Blaser H, Reichman-Fried M, Castanon I, Dumstrei K, Marlow FL, Kawakami K, Solnica-Krezel L, Heisenberg CP, Raz E (2006) Migration of zebrafish primordial germ cells: a role for myosin contraction and cytoplasmic flow. Dev Cell 11(5):613–627CrossRefPubMedGoogle Scholar
  4. Curtis D, Treiber DK, Tao F, Zamore PD, Williamson JR, Lehmann R (1997) A CCHC metal-binding domain in Nanos is essential for translational regulation. EMBO J 16:834–843PubMedCentralCrossRefPubMedGoogle Scholar
  5. D'Agostino I, Merritt C, Chen PL, Seydoux G, Subramaniam K (2006) Translational repression restricts expression of the C. elegans Nanos homolog NOS-2 to the embryonic germline. Dev Biol 292:244–252CrossRefPubMedGoogle Scholar
  6. Drivenes Ø, Taranger GL, Edvardsen RB (2012) Gene expression profiling of Atlantic cod (Gadus morhua) embryogenesis using microarray. Mar Biotechnol (NY) 14:167–176CrossRefGoogle Scholar
  7. Felsenstein J (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39:783–791CrossRefGoogle Scholar
  8. Fujimoto T, Nishimura T, Goto-Kazeto R, Kawakami Y, Yamaha E, Arai K (2010) Sexual dimorphism of gonadal structure and gene expression in germ cell-deficient loach, a teleost fish. Proc Natl Acad Sci U S A 107:17211–17216PubMedCentralCrossRefPubMedGoogle Scholar
  9. Gavis ER, Lehmann R (1994) Translational regulation of nanos by RNA localization. Nature 369:315–318CrossRefPubMedGoogle Scholar
  10. Giraldez AJ, Mishima Y, Rihel J, Grocock RJ, Van Dongen S, Inoue K, Enright AJ, Schier AF (2006) Zebrafish MiR-430 promotes deadenylation and clearance of maternal mRNAs. Sci 312:75–79CrossRefGoogle Scholar
  11. Hashimoto H, Kawaguchi S, Hara K, Nakamura K, Shimizu T, Tamaru Y, Sato M (2009) Purification, crystallization and initial X-ray diffraction study of the zinc-finger domain of zebrafish Nanos. Acta Crystallogr Sect F Struct Biol Cryst Commun 65:959–961PubMedCentralCrossRefPubMedGoogle Scholar
  12. Higaki S, Eto Y, Kawakami Y, Yamaha E, Kagawa N, Kuwayama M, Nagano M, Katagiri S, Takahashi Y (2010) Production of fertile zebrafish (Danio rerio) possessing germ cells (gametes) originated from primordial germ cells recovered from vitrified embryos. Reproduction 139:733–740CrossRefPubMedGoogle Scholar
  13. Johansen SD, Karlsen BO, Furmanek T, Andreassen M, Jørgensen TE, Bizuayehu TT, Breines R, Emblem A, Kettunen P, Luukko K, Edvardsen RB, Nordeide JT, Coucheron DH, Moum T (2011) RNA deep sequencing of the Atlantic cod transcriptome. Comp Biochem Physiol D Genom Proteomics 6:18–22CrossRefGoogle Scholar
  14. Kedde M, Strasser MJ, Boldajipour B, Oude Vrielink JA, Slanchev K, le Sage C, Nagel R, Voorhoeve PM, van Duijse J, Ørom UA, Lund AH, Perrakis A, Raz E, Agami R (2007) RNA-binding protein Dnd1 inhibits microRNA access to target mRNA. Cell 131:1273–1286CrossRefPubMedGoogle Scholar
  15. Knaut H, Steinbeisser H, Schwarz H, Nüsslein-Volhard C (2002) An evolutionary conserved region in the vasa 3'UTR targets RNA translation to the germ cells in the zebrafish. Curr Biol 12:454–466CrossRefPubMedGoogle Scholar
  16. Köprunner M, Thisse C, Thisse B, Raz E (2001) A zebrafish nanos-related gene is essential for the development of primordial germ cells. Genes Dev 15:2877–2885PubMedCentralPubMedGoogle Scholar
  17. Lin F, Liu Q, Li M, Li Z, Hong N, Li J, Hong Y (2012) Transient and stable GFP expression in germ cells by the vasa regulatory sequences from the red seabream (Pagrus major). Int J Biol Sci 8:882–890PubMedCentralCrossRefPubMedGoogle Scholar
  18. Mickoleit M, Banisch TU, Raz E (2011) Regulation of hub mRNA stability and translation by miR430 and the dead end protein promotes preferential expression in zebrafish primordial germ cells. Dev Dyn 240:695–703CrossRefPubMedGoogle Scholar
  19. Mishima Y, Giraldez AJ, Takeda Y, Fujiwara T, Sakamoto H, Schier AF, Inoue K (2006) Differential regulation of germline mRNAs in soma and germ cells by zebrafish miR-430. Curr Biol 16:2135–2142PubMedCentralCrossRefPubMedGoogle Scholar
  20. Nagasawa K, Fernandes JM, Yoshizaki G, Miwa M, Babiak I (2013) Identification and migration of primordial germ cells in Atlantic salmon, Salmo salar: characterization of vasa, dead end, and lymphocyte antigen 75 genes. Mol Reprod Dev 80:118–131PubMedCentralCrossRefPubMedGoogle Scholar
  21. Nei M, Kumar S (2000) Molecular evolution and phylogenetics. Oxford University Press, New YorkGoogle Scholar
  22. Okutsu T, Yano A, Nagasawa K, Shikina S, Kobayashi T, Takeuchi Y, Yoshizaki GJ (2006) Manipulation of fish germ cell: visualization, cryopreservation and transplantation. Reprod Dev 52:685–693CrossRefGoogle Scholar
  23. Presslauer C, Nagasawa K, Fernandes JM, Babiak I (2012) Expression of vasa and nanos3 during primordial germ cell formation and migration in Atlantic cod (Gadus morhua L.). Theriogenology 78:1262–1277CrossRefPubMedGoogle Scholar
  24. Saito T, Fujimoto T, Maegawa S, Inoue K, Tanaka M, Arai K, Yamaha E (2006) Visualization of primordial germ cells in vivo using GFP-nos1 3′UTR mRNA. Int J Dev Biol 50:691–699CrossRefPubMedGoogle Scholar
  25. Saito T, Goto-Kazeto R, Fujimoto T, Kawakami Y, Arai K, Yamaha E (2010) Inter-species transplantation and migration of primordial germ cells in cyprinid fish. Int J Dev Biol 54:1481–1486PubMedGoogle Scholar
  26. Saitou N, Nei M (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425PubMedGoogle Scholar
  27. Slanchev K, Stebler J, de la Cueva-Méndez G, Raz E (2005) Development without germ cells: the role of the germ line in zebrafish sex differentiation. Proc Natl Acad Sci U S A 102:4074–4079PubMedCentralCrossRefPubMedGoogle Scholar
  28. Slanchev K, Stebler J, Goudarzi M, Cojocaru V, Weidinger G, Raz E (2009) Control of Dead end localization and activity—implications for the function of the protein in antagonizing miRNA function. Mech Dev 126:270–277CrossRefPubMedGoogle Scholar
  29. Suzuki H, Saba R, Sada A, Saga Y (2010) The Nanos3-3′UTR is required for germ cell specific NANOS3 expression in mouse embryos. PLoS One 5:e9300PubMedCentralCrossRefPubMedGoogle Scholar
  30. Takeda Y, Mishima Y, Fujiwara T, Sakamoto H, Inoue K (2009) DAZL relieves miRNA-mediated repression of germline mRNAs by controlling poly(A) tail length in zebrafish. PLoS One 4:e7513PubMedCentralCrossRefPubMedGoogle Scholar
  31. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739PubMedCentralCrossRefPubMedGoogle Scholar
  32. Taranger GL, Carrillo M, Schulz RW, Fontaine P, Zanuy S, Felip A, Weltzien FA, Dufour S, Karlsen O, Norberg B, Andersson E (2010) Control of puberty in farmed fish. Gen Comp Endocrinol 165:483–515CrossRefPubMedGoogle Scholar
  33. Thisse C, Thisse B (2008) High-resolution in situ hybridization to whole-mount zebrafish embryos. Nat Protoc 3:59–69CrossRefPubMedGoogle Scholar
  34. Weidinger G, Stebler J, Slanchev K, Dumstrei K, Wise C, Lovell-Badge R, Thisse C, Thisse B, Raz E (2003) Dead end, a novel vertebrate germ plasm component, is required for zebrafish primordial germ cell migration and survival. Curr Biol 13:1429–1434CrossRefPubMedGoogle Scholar
  35. Westerfield M (1995) The zebrafish book: a guide for the laboratory use of zebrafish (Danio rerio), 3rd edn. University of Oregon Press, EugeneGoogle Scholar
  36. Wiszniak SE, Dredge BK, Jensen KB (2011) HuB (elavl2) mRNA is restricted to the germ cells by post-transcriptional mechanisms including stabilisation of the message by DAZL. PLoS One 6:e20773PubMedCentralCrossRefPubMedGoogle Scholar
  37. Wolke U, Weidinger G, Köprunner M, Raz E (2002) Multiple levels of posttranscriptional control lead to germ line-specific gene expression in the zebrafish. Curr Biol 12:289–294CrossRefPubMedGoogle Scholar
  38. Yoshizaki G, Tago Y, Takeuchi Y, Sawatari E, Kobayashi T, Takeuchi T (2005) Green fluorescent protein labeling of primordial germ cells using a nontransgenic method and its application for germ cell transplantation in Salmonidae. Biol Reprod 73:88–93CrossRefPubMedGoogle Scholar
  39. Yoshizaki G, Okutsu T, Morita T, Terasawa M, Yazawa R, Takeuchi Y (2012) Biological characteristics of fish germ cells and their application to developmental biotechnology. Reprod Domest Anim 47(Suppl 4):187–192CrossRefPubMedGoogle Scholar
  40. Youngren KK, Coveney D, Peng X, Bhattacharya C, Schmidt LS, Nickerson ML, Lamb BT, Deng JM, Behringer RR, Capel B, Rubin EM, Nadeau JH, Matin A (2005) The Ter mutation in the dead end gene causes germ cell loss and testicular germ cell tumours. Nature 435(7040):360–364PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  • Adrijana Škugor
    • 1
  • Krasimir Slanchev
    • 2
  • Jacob Seilø Torgersen
    • 1
  • Helge Tveiten
    • 3
  • Øivind Andersen
    • 1
    • 4
    Email author
  1. 1.NofimaAasNorway
  2. 2.Max-Planck Institute of NeurobiologyMartinsriedGermany
  3. 3.NofimaTromsøNorway
  4. 4.Department of Animal and Aquaculture SciencesNorwegian University of Life SciencesAasNorway

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