Metabolic engineering of omega-3 long-chain polyunsaturated fatty acids in plants using an acyl-CoA Delta6-desaturase with omega3-preference from the marine microalga Micromonas pusilla.

Long-chain (> or = C20) polyunsaturated fatty acids (LC-PUFA) EPA and DHA (20:5(Delta5,8,11,14,17) and 22:6(Delta4,7,10,13,16,19)) have well-documented health benefits against coronary heart disease, rheumatoid arthritis and other disorders. Currently, the predominant sources of these fatty acids are marine fish and algal oils, but research is being conducted to ensure that a sustainable, land-based production system can be developed. We here describe the metabolic engineering of an artificial pathway that produces 26% EPA in leaf triacylglycerol using a newly-identified Delta6-desaturase from the marine microalga Micromonas pusilla. We also demonstrate that this enzyme appears to function as an acyl-CoA desaturase that has preference for omega3 substrates both in planta and in yeast. Phylogenetic analysis indicates that this desaturase shares highly conserved motifs with previously described acyl-CoA Delta6-desaturases.

[1]  H. Schaller,et al.  Expression in yeast and tobacco of plant cDNAs encoding acyl CoA:diacylglycerol acyltransferase. , 2000, European journal of biochemistry.

[2]  S. Clough,et al.  Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. , 1998, The Plant journal : for cell and molecular biology.

[3]  A. Bimbo Current and future sources of raw materials for the long‐chain omega‐3 fatty acid market , 2007 .

[4]  Johnathan A Napier,et al.  The production of unusual fatty acids in transgenic plants. , 2007, Annual review of plant biology.

[5]  D. Baulcombe,et al.  An enhanced transient expression system in plants based on suppression of gene silencing by the p19 protein of tomato bushy stunt virus. , 2003, The Plant journal : for cell and molecular biology.

[6]  S. Robert Production of Eicosapentaenoic and Docosahexaenoic Acid-Containing Oils in Transgenic Land Plants for Human and Aquaculture Nutrition , 2005, Marine Biotechnology.

[7]  P. Waterhouse,et al.  Intron-mediated improvement of a selectable marker gene for plant transformation using Agrobacterium tumefaciens , 1997 .

[8]  J. Lake,et al.  Omega-3 fatty acids: evidence basis for treatment and future research in psychiatry. , 2006, The Journal of clinical psychiatry.

[9]  G. Nagel,et al.  The influence of the dietary intake of fatty acids and antioxidants on hay fever in adults , 2003, Allergy.

[10]  A. Green,et al.  Isolation and characterization of genes from the marine microalga Pavlova salina encoding three front-end desaturases involved in docosahexaenoic acid biosynthesis. , 2007, Phytochemistry.

[11]  B. Worm,et al.  Rapid worldwide depletion of predatory fish communities , 2003, Nature.

[12]  Ian J. Brown,et al.  Food Omega-3 Fatty Acid Intake of Individuals (Total, Linolenic Acid, Long-Chain) and Their Blood Pressure: INTERMAP Study , 2007, Hypertension.

[13]  T. Zank,et al.  Acyl Carriers Used as Substrates by the Desaturases and Elongases Involved in Very Long-chain Polyunsaturated Fatty Acids Biosynthesis Reconstituted in Yeast* , 2003, Journal of Biological Chemistry.

[14]  C. Schacky A Review of Omega-3 Ethyl Esters for Cardiovascular Prevention and Treatment of Increased Blood Triglyceride Levels , 2006 .

[15]  Rhoda Au,et al.  Plasma phosphatidylcholine docosahexaenoic acid content and risk of dementia and Alzheimer disease: the Framingham Heart Study. , 2006, Archives of neurology.

[16]  M. Truksa,et al.  Metabolic engineering of plants for polyunsaturated fatty acid production , 2008, Molecular Breeding.

[17]  Xiao Qiu,et al.  Stepwise engineering to produce high yields of very long-chain polyunsaturated fatty acids in plants , 2005, Nature Biotechnology.

[18]  K. Yamato,et al.  Isolation and Characterization of Δ6-Desaturase, an ELO-Like Enzyme and Δ5-Desaturase from the Liverwort Marchantia Polymorpha and Production of Arachidonic and Eicosapentaenoic Acids in the Methylotrophic Yeast Pichia Pastoris , 2004, Plant Molecular Biology.

[19]  H. Moreau,et al.  In vivo characterization of the first acyl-CoA Delta6-desaturase from a member of the plant kingdom, the microalga Ostreococcus tauri. , 2005, The Biochemical journal.

[20]  D. Tocher Issues surrounding fish as a source of omega‐3 long‐chain polyunsaturated fatty acids , 2009 .

[21]  Peter J S Jones,et al.  Fish, human health and marine ecosystem health: policies in collision. , 2009, International journal of epidemiology.

[22]  Peer Bork,et al.  Interactive Tree Of Life (iTOL): an online tool for phylogenetic tree display and annotation , 2007, Bioinform..

[23]  J. Kremer,et al.  Effects of high-dose fish oil on rheumatoid arthritis after stopping nonsteroidal antiinflammatory drugs. Clinical and immune correlates. , 1995, Arthritis and rheumatism.

[24]  B. Meyer,et al.  Dietary intakes and food sources of omega-6 and omega-3 polyunsaturated fatty acids , 2003, Lipids.

[25]  A. Simopoulos Omega-3 Fatty Acids in Inflammation and Autoimmune Diseases , 2002, Journal of the American College of Nutrition.

[26]  A. Green,et al.  Heterologous production of GLA and SDA by expression of an Echium plantagineum Δ6-desaturase gene , 2006 .

[27]  W. J. Dyer,et al.  A rapid method of total lipid extraction and purification. , 1959, Canadian journal of biochemistry and physiology.

[28]  I. Feussner,et al.  Metabolic Engineering of ω3-Very Long Chain Polyunsaturated Fatty Acid Production by an Exclusively Acyl-CoA-dependent Pathway* , 2008, Journal of Biological Chemistry.

[29]  A. Green,et al.  A leaf-based assay using interchangeable design principles to rapidly assemble multistep recombinant pathways. , 2009, Plant biotechnology journal.

[30]  Omega-3 fatty acids and mood disorders. , 2006, The American journal of psychiatry.