Marine Biotechnology

, Volume 13, Issue 1, pp 22–31 | Cite as

Cloning, Tissue Expression Analysis, and Functional Characterization of Two Δ6-Desaturase Variants of Sea Bass (Dicentrarchus labrax L.)

  • Ester Santigosa
  • Florian Geay
  • Thierry Tonon
  • Herve Le Delliou
  • Heiner Kuhl
  • Richard Reinhardt
  • Laurent Corcos
  • Chantal Cahu
  • José Luis Zambonino-Infante
  • David MazuraisEmail author
Original Article


Fish are the main source of the n-3 highly unsaturated fatty acids, which are crucial for human health. Their synthesis from C18 precursors is mediated by desaturases and elongases, but the activity of these enzymes has not been conclusively established in marine fish species. This study reports the cloning, tissue expression, and functional characterization of a sea bass (Dicentrarchus labrax L.) Δ6-desaturase and one of its splicing variants. Two cDNAs with open reading frames of 1,346 and 1,354 bp were cloned and named D6D and D6D-V, respectively. Both deduced protein sequences (445 and 387 amino acids, respectively) contained two transmembrane regions and the N-terminal cytochrome b5 domain with the HPGG motif characteristic of microsomal desaturases. D6D presents three histidine-rich regions, whereas in D6D-V, an insertion of eight nucleotides in the boundaries of exons 10 and 11 modified the third histidine-rich domain and led to insertion of a premature STOP codon, resulting in a shorter predicted protein. Quantitative real-time polymerase chain reaction assay of gene expression showed that D6D was highly expressed in the brain and intestine, and to a lesser extent, in muscle and liver; meanwhile, D6D-V was expressed in all tissues tested, but at level at least 200-fold lower than D6D. Functional analysis in yeast showed that sea bass D6D encodes a fully functional Δ6-desaturase with no residual Δ5-desaturase activity. This desaturase does not exhibit a clear preference for n-3 versus n-6 C18 substrates. Interestingly, D6D-V is a nonfunctional protein, suggesting that the C-terminal end is indispensable for protein activity.


Sea bass (Dicentrarchus labraxDesaturase HUFA biosynthesis Fish EPA 



This work was partly supported by Marine Genomique Europe, by Axe1 “Génomique et Chimie Bleue” Europôle Mer, and by IFR 148 ScInBioS. The authors thank Erick Desmarais for his assistance in gene sequence treatment. E. Santigosa was supported by a postdoctoral fellowship from the Fundación Alfonso Martin Escudero (Spain).

Supplementary material

10126_2010_9264_MOESM1_ESM.doc (40 kb)
Supplementary Table 1 (DOC 40 kb)
10126_2010_9264_MOESM2_ESM.doc (39 kb)
Supplementary Table 2 (DOC 39 kb)
10126_2010_9264_MOESM3_ESM.doc (104 kb)
Supplementary Fig. 1 (DOC 104 kb)


  1. Aki T, Shimada Y, Inagaki K, Higashimoto H, Kawamoto S, Shiget S, Ono K, Suzuki O (1999) Molecular cloning and functional characterisation of rat Δ6 fatty acid desaturase. Biochem Biophys Res Commun 255:575–579PubMedCrossRefGoogle Scholar
  2. Bell JG, Waagbø R (2008) Safe and nutritious aquaculture produce: benefits and risks of alternative sustainable aquafeeds. In: Holmer M, Black KD, Duarte CM, Marba N, Karakassis I (eds) Aquaculture in the ecosystem. Springer Verlag, Netherlands, pp 185–225CrossRefGoogle Scholar
  3. Brunner EJ, Jones PJ, Friel S, Bartley M (2009) Fish, human health and marine ecosystem health: policies in collision. Int J Epidemiol 38:93–100PubMedCrossRefGoogle Scholar
  4. Cho HP, Nakamura MT, Clarke SD (1999) Cloning, expression and nutritional regulation of the human Δ6-desaturase. J Biol Chem 274:471–477PubMedCrossRefGoogle Scholar
  5. Cook HW (1996) Fatty acid desaturation and chain elongation in eukaryote. In: Vance DE, Vance JE (eds) Biochemistry of lipids, lipoproteins and membranes, vol 129. Elsevier, Amsterdam, p 152Google Scholar
  6. Folch J, Lees M, Sloane-Stanley GH (1957) A simple method for the isolation and purification of total lipids from animal tissues. J Biol Chem 226:497–509PubMedGoogle Scholar
  7. Food and Agricultural Organisation (FAO) (2006). State of world aquaculture 2008. Fao Fisheries Technical Paper No. 500. FAO, Rome, p 134Google Scholar
  8. Ghioni C, Tocher DR, Bell MV, Dick JR, Sargent JR (1999) Low C18 to C20 fatty acid elongase activity and limited conversion of stearidonic acid, 18:4n-3, to eicosapentaenoic acid, 20:5n-3, in a cell line from the turbot, Scophthalmus maximus. Biochim Biophys Acta 1437:170–181PubMedGoogle Scholar
  9. Hastings N, Agaba M, Tocher DR, Leaver MJ, Dick JR, Sargent JR, Teale AJ (2001) A vertebrate fatty acid desaturase with Δ5 and Δ6 activities. Proc Natl Acad Sci USA 98:14304–14309PubMedCrossRefGoogle Scholar
  10. Hastings N, Agaba MK, Tocher DR, Zheng X, Dickson CA, Dick JR, Teale AJ (2005) Molecular cloning and functional characterization of fatty acyl desaturase and elongase cDNAs involved in the production of eicosapentaenoic and docosahexaenoic acids from α-linolenic acid in Atlantic salmon (Salmo salar). Mar Biotechnol 6:463–474CrossRefGoogle Scholar
  11. Hoffmann M, Wagner M, Abbadi A, Fulda M, Feussner I (2008) Metabolic engineering of ω3-very long chain polyunsaturated fatty acid production by an exclusively acyl-CoA-dependent pathway. J Biol Chem 283:22352–22362PubMedCrossRefGoogle Scholar
  12. Kaewsuwan S, Cahoon EB, Perroud P-F, Wiwat C, Panvisavas N, Quatrano RS, Cove DJ, Bunyapraphatsara N (2006) Identification and functional characterization of the moss Physcomitrella patens Δ5-desaturase gene involved in arachidonic and eicosapentaenoic acid biosynthesis. J Biol Chem 281:21988–21997PubMedCrossRefGoogle Scholar
  13. Kajikawa M, Yamato KT, Kohzu Y, Nojiri M, Sakuradani E, Shimizu S, Sakai Y, Fukuzawa H, Ohyama K (2004) Isolation and characterization of D6-desaturase, an ELO-like enzyme and D5-desaturase from the Liverwort Marchantia polymorpha and production of arachidonic and eicosapentaenoic acids in the methylotrophic yeast Pichia pastoris. Plant Mol Biol 54:335–352PubMedCrossRefGoogle Scholar
  14. Mourente G, Bell JG (2006) Partial replacement of dietary fish oil with blends of vegetable oils (rapeseed, linseed and palm oils) in diets for European sea bass (Dicentrarchus labrax L.) over a long term growth study: effects on muscle and liver fatty acid composition and effectiveness of a fish oil finishing diet. Comp Biochem Physiol B 145:389–399PubMedCrossRefGoogle Scholar
  15. Mourente G, Dick JR (2002) Influence of partial substitution of dietary fish oil by vegetable oils on the metabolism of [1-C14]18:3n-3 in isolated hepatocytes of European sea bass (Dicentrarchus labrax L.). Fish Physiol Biochem 26:297–308CrossRefGoogle Scholar
  16. Mourente G, Dick JR, Bell JG, Tocher DR (2005a) Effect of partial substitution of dietary fish oil by vegetable oils on desaturation and beta-oxidation of [1-C-14]18:3n-3 (LNA) and [1-C-14]20:5n-3 (EPA) in hepatocytes and enterocytes of European sea bass (Dicentrarchus labrax L.). Aquaculture 248:173–186CrossRefGoogle Scholar
  17. Mourente G, Good JE, Bell JG (2005b) Partial substitution of fish oil with rapeseed, linseed and olive oils in diets for European sea bass (Dicentrarchus labrax L.): effects on flesh fatty acid composition, plasma prostaglandins E2 and F, immune function and effectiveness of a fish oil finishing diet. Aquac Nutr 11:25–40CrossRefGoogle Scholar
  18. Napier JA, Michaelson LV, Sayanova O (2003) The role of cytochrome b5 fusion desaturases in the synthesis of polyunsaturated fatty acids. Prost Leukot Essent Fatty Acids 68:135–143CrossRefGoogle Scholar
  19. Pauly D, Watson R, Alder J (2005) Global trends in world fisheries: impacts on marine ecosystems and food security. Philos Trans R Soc Lond B Biol Sci 360:5–12PubMedCrossRefGoogle Scholar
  20. Saitou N, Nei M (1987) The neighbor-joining method. A new method for reconstructing phylogenetic trees. Mol Biol Evol 4:406–425PubMedGoogle Scholar
  21. Sargent JR, Bell JR, Bell MV, Henderson RJ, Tocher DR (1995) Requirement criteria for essential fatty acids. J Appl Ichthyol 11:183–198CrossRefGoogle Scholar
  22. Sargent JR, Bell JG, McEvoy L, Tocher DR, Estevez A (1999) Recent developments in the essential fatty acid nutrition of fish. Aquaculture 177:191–199CrossRefGoogle Scholar
  23. Sargent JR, Tocher DR, Bell JG (2002) The lipids. In: Halver JE, Hardy RW (eds) Fish nutrition, 3rd edn. Academic, USA, pp 181–257Google Scholar
  24. Sayanova OV, Beaudoin F, Michaelson LV, Shewry PR, Napier JA (2003) Identification of Primula fatty acid Δ6-desaturases with n-3 substrate preferences. FEBS Lett 542:100–104PubMedCrossRefGoogle Scholar
  25. Seiliez I, Panserat S, Corraze G, Kaushik S, Bergot P (2003) Cloning and nutritional regulation of a Δ6-desaturase-like enzyme in the marine teleost gilthead sea bream (Sparus aurata). Comp Biochem Physiol B 135:449–460PubMedCrossRefGoogle Scholar
  26. Shanklin J, Whittle E, Fox BG (1994) Eight histidine residues are catalytically essential in a membrane-associated iron enzyme, stearoyl-CoA desaturase, and are conserved in alkane hydroxylase and xylene monooxygenase. Biochemistry 33:12787–12794PubMedCrossRefGoogle Scholar
  27. Tidwell JH, Allan GL (2001) Fish as food: aquaculture's contribution. Ecological and economic impacts and contributions of fish farming and capture fisheries. Embo Reports 2:958–963PubMedCrossRefGoogle Scholar
  28. Tocher DR, Ghioni C (1999) Fatty acid metabolism in marine fish: low activity of Δ5 desaturation in gilthead sea bream (Sparus aurata) cells. Lipids 34:433–440PubMedCrossRefGoogle Scholar
  29. Tocher D, Zheng X, Schlechtriem C, Hastings N, Dick J, Teale A (2006) Highly unsaturated fatty acid synthesis in marine fish: cloning, functional characterization, and nutritional regulation of fatty acyl Δ6-desaturase of Atlantic cod (Gadus morhua L.). Lipids 41:1003–1016PubMedCrossRefGoogle Scholar
  30. Zheng X, Seiliez I, Hastings N, Tocher DR, Panserat S, Dickson CA, Bergot P, Teale AJ (2004) Characterization and comparison of fatty acyl Δ6-desaturase cDNAs from freshwater and marine teleost fish species. Comp Biochem Physiol B 139:269–279PubMedCrossRefGoogle Scholar
  31. Zheng X, Tocher DR, Dickson CA, Dick JR, Bell JG, Teale AJ (2005) Highly unsaturated fatty acid synthesis in vertebrates: new insights with the cloning and characterization of a Δ6-desaturase of Atlantic salmon. Lipids 40:13–24PubMedCrossRefGoogle Scholar
  32. Zheng X, King Z, Xu Y, Monroig O, Morais S, Tocher DR (2009) Physiological roles of fatty acyl desaturases and elongases in marine fish: characterisation of cDNAs of fatty acyl Δ6-desaturase and elovl5 elongase of cobia (Rachycentron canadum). Aquaculture 290:122–131CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Ester Santigosa
    • 1
  • Florian Geay
    • 1
  • Thierry Tonon
    • 2
    • 3
  • Herve Le Delliou
    • 1
  • Heiner Kuhl
    • 4
  • Richard Reinhardt
    • 4
  • Laurent Corcos
    • 5
    • 6
  • Chantal Cahu
    • 1
  • José Luis Zambonino-Infante
    • 1
  • David Mazurais
    • 1
    Email author
  1. 1.Ifremer Marine Fish Nutrition Team, Nutrition Aquaculture and Genomics Research UnitUMR 1067, Ifremer, Technopole Brest-IroisePlouzanéFrance
  2. 2.UPMC Univ Paris 6, UMR 7139 Végétaux marins et biomolécules, Station BiologiqueRoscoffFrance
  3. 3.CNRS, UMR 7139 Végétaux marins et biomolécules, Station BiologiqueRoscoffFrance
  4. 4.Max-Planck-Institute Molecular GeneticsBerlin-DahlemGermany
  5. 5.Université de Brest, INSERM, U613, ECLABrestFrance
  6. 6.Faculté de MédecineUniversité Européenne de BretagneBrestFrance

Personalised recommendations