DOCSLIB.ORG

Identification of an Animal -3 Fatty Acid Desaturase by Heterologous Expression in Arabidopsis

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Identification of an Animal -3 Fatty Acid Desaturase by Heterologous Expression in Arabidopsis Proc. Natl. Acad. Sci. USA Vol. 94, pp. 1142–1147, February 1997 Biochemistry Identification of an animal v-3 fatty acid desaturase by heterologous expression in Arabidopsis (fatty acid desaturationyexpressed sequence tagyfat-1yarachidonic acidypolyunsaturated fatty acids) JAMES P. SPYCHALLA*†,ANTHONY J. KINNEY‡, AND JOHN BROWSE*§ *Institute of Biological Chemistry, Washington State University, Pullman, WA 99164-6340; and ‡Agricultural Products, Experiment Station 402, 4247 E.I. duPont de Nemours and Co., P.O. Box 80402, Wilmington, DE 19880-0402 Communicated by Christopher Somerville, Carnegie Institution of Washington, Stanford, CA, December 3, 1996 (received for review July 12, 1996) ABSTRACT In animals, fatty acid desaturases catalyze Biochemical studies of membrane-bound fatty acid desatu- key reactions in the synthesis of arachidonic acid and other rases in plants have proven equally difficult, and only one polyunsaturated fatty acids. A search of the Caenorhabditis enzyme has been purified to homogeneity (7). Higher plants elegans DNA databases, using the sequences of Arabidopsis produce many different unsaturated fatty acids (8), but in genes, identified several putative desaturases. Here we de- membrane lipids the major locations for double bonds are at scribe the characterization of the first of these genes, fat-1. The the D9, -12, and -15 (v-3) positions of 18-carbon acyl chains predicted protein encoded by a fat-1 cDNA showed 32–35% and the corresponding D7, -10, and -13 (v-3) positions of identity with both FAD2 and FAD3 of Arabidopsis. When 16-carbon chains (9). In Arabidopsis, the isolation of mutants expressed in transgenic plants, fat-1 resulted in a 90% increase with altered fatty acid compositions (10) has facilitated studies in the proportion of a-linolenic acid in root lipids. Wild-type of the biochemistry and function of the seven membrane Arabidopsis incorporated v-6 fatty acids (D8,11,14–20:3 and desaturases present in this plant (11). Furthermore, the ex- D5,8,11,14–20:4) into membrane lipids but did not desaturate tensive molecular genetic tools available in Arabidopsis al- them. By contrast, fat-1 transgenic plants efficiently desatu- lowed genes encoding desaturases to be cloned without the rated both of these fatty acids to the corresponding v-3 need to first solubilize and purify the enzymes themselves (12, products. These findings indicate that the C. elegans fat-1 gene 13). Interestingly, all the plant membrane-bound desaturases encodes the first animal representative of a class of glycero- use complex glycerolipids rather than acyl-CoAs as substrates. lipid desaturases that have previously been characterized in This finding and data indicating that the D5 desaturase from plants and cyanobacteria. The FAT-1 protein is an v-3 fatty rat liver acts on glycerolipids (14) suggest the possibility that acyl desaturase that recognizes a range of 18- and 20-carbon the majority of animal desaturases also catalyze glycerolipid- v-6 substrates. linked desaturation. However, the lack of progress in charac- terizing other animal desaturases has precluded further dis- Polyunsaturated fatty acids are important as structural com- cussion of this issue. ponents of membrane glycerolipids and as precursors of fam- We set out to use the Arabidopsis desaturase genes and the ilies of signaling molecules including prostaglandins, throm- extensive information now available from the Caenorhabditis boxanes, and leukotrienes (1, 2). The principal fatty acid elegans genome project (15) to identify and clone genes precursors of these signaling compounds are arachidonic acid encoding fatty acid desaturases in animals. C. elegans is a good (D5,8,11,14–20:4), providing an v-6 substrate that is respon- model in this respect because it elaborates a wide range of sible for the major synthesis of these compounds, and eicosa- polyunsaturated fatty acids including arachidonic and eicosa- pentaenoic acid (D5,8,11,14,17–20:5), an v-3 substrate that is pentaenoic acids, from very simple precursors available in its responsible for the parallel synthesis of many eicosanoids with diet (16, 17). In this communication, we report the isolation an additional double bond. An important class of enzymes and characterization of a cDNA from C. elegans that, when involved in the synthesis of polyunsaturated fatty acids are the expressed in Arabidopsis, encodes a novel fatty acid desaturase fatty acid desaturases that catalyze the introduction of double which can catalyze the introduction of an v-3 double bond into bonds into the hydrocarbon chain. In vertebrates, desaturases a range of 18- and 20-carbon fatty acids. are known to act at the D4, -5, -6, -8, and -9 positions (3). The 18:0-CoA D9 desaturase from rat liver has been characterized MATERIALS AND METHODS biochemically (4, 5), and the corresponding gene has been Database Searches. cloned (6). However, the remaining four enzymes have re- The National Center for Biotechnology mained recalcitrant to purification and genes that encode them Information’s (NCBI) peptide sequence database was searched using a BLAST server, with the peptide sequences of have not been isolated. Based on available information, and by the Arabidopsis thaliana FAD2, FAD6 and FAD7 fatty acid analogy to the 18:0-CoA desaturase, it is likely that these desaturases as queries. The GenBank accession numbers for additional enzymes are integral membrane proteins that re- the corresponding Arabidopsis cDNAs are L26296, U09503, quire other membrane components (cytochrome b and 5 and L22931, respectively. NADH:cytochrome b reductase) for activity (4), and it is 5 Cloning and Sequencing of a fat-1 cDNA. The partial cDNA these features that have limited progress in studying the identified by a database search, CEL10e11, was obtained from biochemistry and molecular genetics of these important syn- the C. elegans Genome Sequencing Center (Washington Uni- thetic reactions. versity School of Medicine, St. Louis). The cDNA insert was released by double digestion with HindIII and SacI, gel The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked ‘‘advertisement’’ in accordance with 18 U.S.C. §1734 solely to indicate this fact. Data deposition: The sequence reported in this paper has been deposited in the GenBank database (accession no. L41807). Copyright q 1997 by THE NATIONAL ACADEMY OF SCIENCES OF THE USA †Present address: Spychalla Farms, Inc., N3974 Highway 52, Antigo, 0027-8424y97y941142-6$2.00y0 WI 54409. PNAS is available online at http:yywww.pnas.org. §To whom reprint requests should be addressed. 1142 Downloaded by guest on September 24, 2021 Biochemistry: Spychalla et al. Proc. Natl. Acad. Sci. USA 94 (1997) 1143 purified, and labeled with [32P]dCTP using a random prime kit esters derived from phosphatidylcholine were separated on a (Promega). The denatured probe was used to screen a mixed 30 m 3 0.2 mm AT1000 column (Alltech Associates) in a stage C. elegans cDNA l phage (Uni-Zap XR) library (Strat- HP6890 Instrument (Hewlett–Packard). Oven temperature at agene). Nucleic acid hybridizations and high stringency washes injection was 1508C, and this was increased at 58Cymin to were performed as described (13). Hybridizing plaques were 2308C then held at 2308C for 10 min. Criteria for identification visualized by autoradiography. Positive clones were isolated of D5,8,11,14,17–20:5 in phosphatidylcholine from fat-1 trans- and excised from the phage vector according to the manufac- genic plants where the identification of a mass peak at myz 5 turer’s protocol to yield pBluescript plasmids. The plasmid 316, which corresponds to the expected molecular ion, and a clone with the longest insert, pCE8, was completely sequenced retention time (36.11 min) and fragmentation pattern that in both directions. Sequence analysis was carried out using the were identical to those of the authentic D5,8,11,14,17–20:5 programs available in the Genetics Computer Group package standard. No commercial standard was available for (18) using default settings for parameters unless indicated. The D8,11,14,17–20:4. A fatty acid methyl ester present in fat-1 DNA sequence corresponding to the ORF of the fat-1 cDNA transgenic plants sprayed with D8,11,14–20:3 but not in wild- was used to search the database of the C. elegans genome type control plants had a retention time of 35.74 min during gas sequencing project using the BLAST server accessed through chromatography–mass spectrometry. This compound showed the World Wide Web interface (15). No homologous sequence a mass peak at myz 5 318 (the expected molecular ion for 20:4) was found (highest BLAST score, 145; P 5 0.034), indicating that and a fragmentation pattern very similar to that of the the fat-1 genomic sequence had not yet been included by this authentic D5,8,11,14–20:4 standard. The retention time of project. D5,8,11,14–20:4 was 35.04 min for both the authentic standard Gene Construct for Plant Expression. The cauliflower and for the methyl esters recovered from plants sprayed with mosaic virus 35S promoterynopaline synthase terminator cas- soaps of this isomer. Therefore it was concluded that the new sette of Baulcombe et al. (19) was cloned into the XbaIyEcoRI compound detected only in fat-1 transgenic plants sprayed with sites of pBIN400 (20) to make the binary transformation vector D8,11,14–20:3 was an isomer of 20:4 and most probably pBIN420. The cDNA insert of pCE8 was released with a D8,11,14,17–20:4. EcoRIyKpnI double digest, end-filled with Klenow fragment, Nucleic Acid Analysis. Total RNA was extracted from leaves and blunt-ligated into the SmaI site of pBIN420 to make according to the method of Verwoerd et al. (24). Twenty-five pBIN420-CE8. These vectors contain the NPTII gene within micrograms of total RNA was separated on 1.2% agarose– their T-DNA, thus conferring kanamycin resistance to trans- formaldehyde gels and blot transferred to nylon membranes.
Load more

Recommended publications
  • Fatty Acid Metabolism and Desaturation in the Pathogenesis of Leukemic Stem Cells in Acute Myeloid Leukemia
    Fatty acid metabolism and desaturation in the pathogenesis of leukemic stem cells in Acute Myeloid Leukemia Rachel Culp-Hill1,2, Courtney Jones3, Brett Stevens3, Daniel Pollyea3, Shanshan Pei3, Craig Jordan3, Angelo D’Alessandro1,2 1Graduate Program in Structural Biology and Biochemistry, 2Department of Biochemistry & Molecular Genetics, 3Blood Cancer and BMT Program, Division of Hematology, University of Colorado AMC, Aurora, CO, USA Abstract Ven/Aza targets de novo LSC metabolism… Dysregulation of fatty acid desaturation in relapse p53 controls FA metabolism and desaturation p53 knockout in an AML cell line results in Background: Acute myeloid leukemia (AML) is a cancer of bone marrow- LSCs blasts 510 06 Vehicle Ven+Aza 1.210 7 Saturated Lipids Unsaturated Lipids derived blood cells, where leukemic blasts build up and block function and Newly diagnosed (de novo) ALA ALA increased unsaturated fatty acids, including 1.0 10 7 development of myeloid progenitors. Conventional therapy eliminates bulk LSCs uptake amino acids 410 06 ARG ARG Diagnosis docosapentaenoic acid (22:5) and tumor cells but leukemic stem cells (LSCs) survive, leading to disease ASP more quickly than leukemic 6 ASP 8.010 Relapse docosahexaenoic acid (22:6). Loss of p53 progression and relapse. LSCs uniquely rely on oxidative phosphorylation CYS blasts, and when amino GLU 310 06 6 increases unsaturated fatty acids, similarly (OXPHOS), metabolically driven by amino acid and fatty acid metabolism. C-C GLN 6.010 acids are removed, 06 GLU HIS 210 to the relapsed AML phenotype. 4.010 6 Aim: We have successfully targeted amino acid metabolism in LSCs, but the GLN OXPHOS is reduced.
    [Show full text]
  • Fatty Acid Desaturases: Uncovering Their Involvement in Grapevine Defence Against Downy Mildew
    International Journal of Molecular Sciences Article Fatty Acid Desaturases: Uncovering Their Involvement in Grapevine Defence against Downy Mildew Gonçalo Laureano * , Ana Rita Cavaco , Ana Rita Matos and Andreia Figueiredo * Biosystems & Integrative Sciences Institute (BioISI), Faculdade de Ciências, Universidade de Lisboa, Campo Grande, 1749-016 Lisboa, Portugal; [email protected] (A.R.C.); [email protected] (A.R.M.) * Correspondence: [email protected] (G.L.); aafi[email protected] (A.F.) Abstract: Grapevine downy mildew, caused by the biotrophic oomycete Plasmopara viticola, is one of the most severe and devastating diseases in viticulture. Unravelling the grapevine defence mechanisms is crucial to develop sustainable disease control measures. Here we provide new insights concerning fatty acid’s (FA) desaturation, a fundamental process in lipid remodelling and signalling. Previously, we have provided evidence that lipid signalling is essential in the establishment of the incompatible interaction between grapevine and Plasmopara viticola. In the first hours after pathogen challenge, jasmonic acid (JA) accumulation, activation of its biosynthetic pathway and an accumulation of its precursor, the polyunsaturated α-linolenic acid (C18:3), were observed in the leaves of the tolerant genotype, Regent. This work was aimed at a better comprehension of the desaturation processes occurring after inoculation. We characterised, for the first time in Vitis vinifera, the gene family of the FA desaturases and evaluated their involvement in Regent response to Plasmopara viticola. Upon pathogen challenge, an up-regulation of the expression of plastidial FA desaturases genes was observed, resulting in a higher content of polyunsaturated fatty acids (PUFAs) of chloroplast lipids. This study highlights FA desaturases as key players in membrane remodelling Citation: Laureano, G.; Cavaco, A.R.; and signalling in grapevine defence towards biotrophic pathogens.
    [Show full text]
  • Fatty Acid Desaturases, Polyunsaturated Fatty Acid Regulation, and Biotechnological Advances
    Review Fatty Acid Desaturases, Polyunsaturated Fatty Acid Regulation, and Biotechnological Advances Je Min Lee 1,†, Hyungjae Lee 2,†, SeokBeom Kang 3 and Woo Jung Park 4,* Received: 11 October 2015; Accepted: 17 December 2015; Published: 4 January 2016 1 Department of Horticultural Science, Kyungpook National University, Daegu 41566, Korea; [email protected] 2 Department of Food Engineering, Dankook University, Cheonan, Chungnam 31116, Korea; [email protected] 3 Citrus Research Station, National Institute of Horticultural & Herbal Science, RDA, Seogwipo 63607, Korea; [email protected] 4 Department of Marine Food Science and Technology, Gangneung-Wonju National University, Gangneung, Gangwon 25457, Korea * Correspondence: [email protected] or [email protected]; Tel.: +82-33-640-2857; Fax: +82-33-640-2850 † These authors contributed equally to this work. Abstract: Polyunsaturated fatty acids (PUFAs) are considered to be critical nutrients to regulate human health and development, and numerous fatty acid desaturases play key roles in synthesizing PUFAs. Given the lack of delta-12 and -15 desaturases and the low levels of conversion to PUFAs, humans must consume some omega-3 and omega-6 fatty acids in their diet. Many studies on fatty acid desaturases as well as PUFAs have shown that fatty acid desaturase genes are closely related to different human physiological conditions. Since the first front-end desaturases from cyanobacteria were cloned, numerous desaturase genes have been identified and animals and plants have been genetically engineered to produce PUFAs such as eicosapentaenoic acid and docosahexaenoic acid. Recently, a biotechnological approach has been used to develop clinical treatments for human physiological conditions, including cancers and neurogenetic disorders.
    [Show full text]
  • Ricinus Communis L.) Mutant
    This is a post-peer-review, pre-copyedit version of an article published in Planta. The final authenticated version is available online at: https://doi.org/10.1007/s00425-016-2508-4 Molecular and biochemical characterization of the OLE-1 high-oleic castor seed (Ricinus communis L.) mutant Mónica Venegas-Calerón*, Rosario Sánchez, Joaquín J. Salas, Rafael Garcés and Enrique Martínez-Force Instituto de la Grasa (CSIC), Edificio 46, Campus Universitario Pablo de Olavide, Carretera de Utrera Km 1, 41013 Sevilla, Spain. *To whom correspondence should be addressed Running title: hydroxylase from a high-oleic castor bean Corresponding author: Mónica Venegas-Calerón Email: [email protected] Tlf: +34 954611550-259 Main conclusion: The natural OLE-1 high oleic castor mutant has been characterized, demonstrating that point mutations in the FAH12 gene are responsible for the high oleic phenotype. The contribution of each mutation was evaluated by heterologous expression in yeast and lipid studies in developing OLE-1 seeds provided new evidence of unusual fatty acids channelling into TAGs. 1 Abstract Ricinus communis L. is a plant of the Euphorbiaceae family well known for producing seeds whose oil has a very high ricinoleic (12-hydroxy-octadecenoic) acid content. Castor oil is considered the only commercially renewable source of hydroxylated fatty acids, which have many applications as chemical reactants. Accordingly, there has been great interest in the field of plant lipid biotechnology to define how ricinoleic acid is synthesised, which could also provide information that might serve to increase the content of other unusual fatty acids in oil crops. Accordingly, we set out to study the biochemistry of castor oil synthesis by characterizing a natural castor bean mutant deficient in ricinoleic acid synthesis (OLE- 1).
    [Show full text]
  • The Key Roles of Elongases and Desaturases in Mammalian Fatty Acid Metabolism: Insights from Transgenic Mice
    Progress in Lipid Research 49 (2010) 186–199 Contents lists available at ScienceDirect Progress in Lipid Research journal homepage: www.elsevier.com/locate/plipres Review The key roles of elongases and desaturases in mammalian fatty acid metabolism: Insights from transgenic mice Hervé Guillou a, Damir Zadravec b, Pascal G.P. Martin a, Anders Jacobsson b,* a Integrative Toxicology and Metabolism, Pôle de Toxicologie Alimentaire, Laboratoire de Pharmacologie et Toxicologie, Institut National de la Recherche Agronomique INRA UR66, Toulouse Cedex 3, France b Dept. of Physiology, The Wenner-Gren Institute, The Arrhenius Laboratories F3, Stockholm University, SE-10691 Stockholm, Sweden article info abstract Article history: In mammalian cells, elongases and desaturases play critical roles in regulating the length and degree of Received 19 November 2009 unsaturation of fatty acids and thereby their functions and metabolic fates. In the past decade, a great Received in revised form 9 December 2009 deal has been learnt about these enzymes and the first part of this review summarizes our current knowl- Accepted 10 December 2009 edge concerning these enzymes. More recently, several transgenic mouse models lacking either an elong- ase (Elovl3À/À, Elovl4À/À, Elovl5À/À, Elovl6À/À) or a desaturase (Scd-1À/À, Scd-2À/À, Fads2À/À) have been developed and the second part of this review focuses on the insights gained from studies with these mice, Keywords: as well as from investigations on cell cultures. Elongase Ó 2009 Elsevier Ltd. All rights reserved. Desaturase VLCFA PUFA Knockout mice Contents 1. Introduction ......................................................................................................... 187 2. Mammalian elongases and desaturases . ...................................................................... 188 2.1. Elongases . ............................................................................................... 188 2.2.
    [Show full text]
  • FADS1 (Fatty Acid Desaturase 1) Genotype Associates with Aortic Valve FADS Mrna Expression, Fatty Acid Content and Calcification Oscar Plunde, MD; Susanna C
    Circulation: Genomic and Precision Medicine ORIGINAL ARTICLE FADS1 (Fatty Acid Desaturase 1) Genotype Associates With Aortic Valve FADS mRNA Expression, Fatty Acid Content and Calcification Oscar Plunde, MD; Susanna C. Larsson , PhD; Gonzalo Artiach, MSc; George Thanassoulis, MD; Miguel Carracedo, PhD; Anders Franco-Cereceda, MD, PhD; Per Eriksson, PhD; Magnus Bäck , MD, PhD BACKGROUND: Aortic stenosis (AS) contributes to cardiovascular mortality and morbidity but disease mechanisms remain largely unknown. Recent evidence associates a single nucleotide polymorphism rs174547 within the FADS1 gene, encoding FADS1 (fatty acid desaturase 1), with risk of several cardiovascular outcomes, including AS. FADS1 encodes a rate-limiting enzyme for ω-3 and ω-6 fatty acid metabolism. The aim of this study was to decipher the local transcriptomic and lipidomic consequences of rs174547 in tricuspid aortic valves from patients with AS. METHODS: Expression quantitative trait loci study was performed using data from Illumina Human610-Quad BeadChip, Infinium Global Screening Arrays, and Affymetrix Human Transcriptome 2.0 arrays in calcified and noncalcified aortic valve tissue from 58 patients with AS (mean age, 74.2; SD, 5.9). Fatty acid content was assessed in aortic valves from 25 patients with AS using gas chromatography. Δ5 and Δ6 desaturase activity was assessed by the product-to-precursor ratio. RESULTS: The minor C-allele of rs174547, corresponding to the protective genotype for AS, was associated with higher FADS2 mRNA levels in calcified valve tissue, whereas FADS1 mRNA and other transcripts in proximity of the single nucleotide Downloaded from http://ahajournals.org by on September 23, 2020 polymorphism were unaltered. In contrast, the FADS1 Δ5-desaturase activity and the FADS2 Δ6-desaturase activity were decreased.
    [Show full text]
  • Genome of Wild Olive and the Evolution of Oil Biosynthesis
    Genome of wild olive and the evolution of PNAS PLUS oil biosynthesis Turgay Unvera,1,2,3, Zhangyan Wub,1, Lieven Sterckc,d, Mine Turktase, Rolf Lohausc,d, Zhen Lic,d, Ming Yangb, Lijuan Heb, Tianquan Dengb, Francisco Javier Escalantef, Carlos Llorensg, Francisco J. Roigg, Iskender Parmaksizh, Ekrem Dundari, Fuliang Xiej, Baohong Zhangj, Arif Ipeke, Serkan Uranbeyk, Mustafa Eraymanl, Emre Ilhanl, Oussama Badadm, Hassan Ghazaln, David A. Lightfooto, Pavan Kasarlao, Vincent Colantonioo, Huseyin Tombuloglup, Pilar Hernandezq, Nurengin Meter, Oznur Cetinr, Marc Van Montaguc,d,3, Huanming Yangb, Qiang Gaob, Gabriel Dorados, and Yves Van de Peerc,d,t,3 aIzmir_ International Biomedicine and Genome Institute, Dokuz Eylül University, 35340 Izmir,_ Turkey; bBGI Shenzhen, 518038 Shenzhen, China; cDepartment of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Ghent, Belgium; dCenter for Plant Systems Biology, VIB, 9052 Ghent, Belgium; eDepartment of Biology, Faculty of Science, Cankiri Karatekin University, 18100 Cankiri, Turkey; fPlataforma de Genómica y Bioinformática de Andalucía, 41013 Sevilla, Spain; gBiotechvana, 46980 Paterna (Valencia), Spain; hDepartment of Molecular Biology and Genetics, Faculty of Science, Gaziosmanpasa University, 60250 Tokat, Turkey; iDepartment of Molecular Biology and Genetics, Faculty of Science, Balikesir University, 10145 Balikesir, Turkey; jDepartment of Biology, East Carolina University, Greenville, NC 27858; kDepartment of Field Crops, Faculty of Agriculture, Ankara University, 06120 Ankara, Turkey;
    [Show full text]
  • Production of Eicosapentaenoic Acid by Application of a Delta-6
    Shi et al. Microb Cell Fact (2018) 17:7 https://doi.org/10.1186/s12934-018-0857-3 Microbial Cell Factories RESEARCH Open Access Production of eicosapentaenoic acid by application of a delta‑6 desaturase with the highest ALA catalytic activity in algae Haisu Shi, Xue Luo, Rina Wu* and Xiqing Yue* Abstract: Dunaliella salina is a unicellular green alga with a high α-linolenic acid (ALA) level, but a low eicosapentaenoic acid (EPA) level. In a previous analysis of the catalytic activity of delta 6 fatty acid desaturase (FADS6) from various spe- cies, FADS6 from Thalassiosira pseudonana (TpFADS6), a marine diatom, showed the highest catalytic activity for ALA. In this study, to enhance EPA production in D. salina, FADS6 from D. salina (DsFADS6) was identifed, and substrate specifcities for DsFADS6 and TpFADS6 were characterized. Furthermore, a plasmid harboring the TpFADS6 gene was constructed and overexpressed in D. salina. Our results revealed that EPA production reached 21.3 1.5 mg/L in D. salina transformants. To further increase EPA production, myoinositol (MI) was used as a growth-promoting± agent; it increased the dry cell weight of D. salina transformants, and EPA production reached 91.3 11.6 mg/L. The combina- ± tion of 12% ­CO2 aeration with glucose/KNO3 in the medium improved EPA production to 192.9 25.7 mg/L in the Ds-TpFADS6 transformant. We confrmed that the increase in ALA was optimal at 8 °C; the EPA percentage± reached 41.12 4.78%. The EPA yield was further increased to 554.3 95.6 mg/L by supplementation with 4 g/L perilla seed ± ± meal (PeSM), 500 mg/L MI, and 12% ­CO2 aeration with glucose/KNO3 at varying temperatures.
    [Show full text]
  • RT² Profiler PCR Array (Rotor-Gene® Format) Chicken Fatty Acid Metabolism
    RT² Profiler PCR Array (Rotor-Gene® Format) Chicken Fatty Acid Metabolism Cat. no. 330231 PAGG-007ZR For pathway expression analysis Format For use with the following real-time cyclers RT² Profiler PCR Array, Rotor-Gene Q, other Rotor-Gene cyclers Format R Description The Chicken Fatty Acid Metabolism RT² Profiler PCR Array profiles the expression of 84 key genes involved in the regulation and enzymatic pathways of fatty acid metabolism. Cells, particularly in skeletal muscle and adipose tissue, primarily store energy as triacylglycerols and, when needed, break them down again into glycerol and fatty acids for activation and transport into the mitochondria. The process of β-oxidation then metabolizes these activated fatty acids yielding acetyl-CoA, the initial metabolite necessary for the TCA cycle and ketogenesis. During resting states, cells store excess energy by re-synthesizing fatty acids in a process tightly regulated by hormones. Alterations in the expression of genes involved in fatty acid metabolism, such as CRAT, often associate with metabolic syndrome and insulin resistance. These two syndromes are risk factors for multiple diseases including diabetes and obesity as well as other prevalent health problems such as cardiovascular disease. A set of controls present on each array enables data analysis using the ∆∆CT method of relative quantification, assessment of reverse transcription performance, genomic DNA contamination, and PCR performance. Using real-time PCR, you can easily and reliably analyze the expression of a focused panel of genes involved in fatty acid metabolism with this array. For further details, consult the RT² Profiler PCR Array Handbook. Shipping and storage RT² Profiler PCR Arrays in the Rotor-Gene format are shipped at ambient temperature, on dry ice, or blue ice packs depending on destination and accompanying products.
    [Show full text]
  • Omega-3 Versus Omega-6 Polyunsaturated Fatty Acids in the Prevention and Treatment of Inflammatory Skin Diseases
    International Journal of Molecular Sciences Review Omega-3 Versus Omega-6 Polyunsaturated Fatty Acids in the Prevention and Treatment of Inflammatory Skin Diseases Anamaria Bali´c 1 , Domagoj Vlaši´c 2, Kristina Žužul 3, Branka Marinovi´c 1 and Zrinka Bukvi´cMokos 1,* 1 Department of Dermatology and Venereology, University Hospital Centre Zagreb, School of Medicine University of Zagreb, Šalata 4, 10 000 Zagreb, Croatia; [email protected] (A.B.); [email protected] (B.M.) 2 Department of Ophtalmology and Optometry, General Hospital Dubrovnik, Ulica dr. Roka Mišeti´ca2, 20000 Dubrovnik, Croatia; [email protected] 3 School of Medicine, University of Zagreb, Šalata 3, 10000 Zagreb, Croatia; [email protected] * Correspondence: [email protected] Received: 31 December 2019; Accepted: 21 January 2020; Published: 23 January 2020 Abstract: Omega-3 (!-3) and omega-6 (!-6) polyunsaturated fatty acids (PUFAs) are nowadays desirable components of oils with special dietary and functional properties. Their therapeutic and health-promoting effects have already been established in various chronic inflammatory and autoimmune diseases through various mechanisms, including modifications in cell membrane lipid composition, gene expression, cellular metabolism, and signal transduction. The application of !-3 and !-6 PUFAs in most common skin diseases has been examined in numerous studies, but their results and conclusions were mostly opposing and inconclusive. It seems that combined !-6, gamma-linolenic acid (GLA), and
    [Show full text]
  • The Omega-6/Omega-3 Fatty Acid Ratio: Health Implications
    NUTRITION – SANTÉ The omega-6/omega-3 fatty acid ratio: health implications Artemis P. SIMOPOULOS Abstract: Today, Western diets are characterized by a higher omega-6 and a lower omega-3 fatty acid intake, whereas during the Paleolithic period when human’s genetic profile was established, there was a The Center for Genetics, balance between omega-6 and omega-3 fatty acids. Their balance is an important determinant for Nutrition and Health, brain development and in decreasing the risk for coronary heart disease (CHD), hypertension, cancer, 2001 S Street, NW, Suite 530, diabetes, arthritis, and other autoimmune and possibly neurodegenerative diseases. Both omega-6 and Washington DC 20009 omega-3 fatty acids influence gene expression. Because of single nucleotide polymorphisms (SNPs) in USA their metabolic pathways, blood levels of omega-6 and omega-3 fatty acids are determined by both en- <[email protected]> dogenous metabolism and dietary intake making the need of balanced dietary intake essential for health and disease prevention. Whether an omega-6/omega-3 ratio of 3:1 to 4:1 could prevent the pathogenesis of many diseases induced by today’s Western diets (AFSSA, 2010), a target of 1:1 to 2:1 appears to be consistent with studies on evolutionary aspects of diet, neurodevelopment, and genetics. A target of omega-6/omega-3 fatty acid ratio of 1:1 to 2:1 appears to be consistent with studies on evolutionary aspects of diet, neurodevelopment and genetics. A balanced ratio of omega-6/omega-3 fatty acids is important for health and in the prevention of CHD and possibly other chronic diseases.
    [Show full text]
  • Production of Novel Oils in Plants Denis J Murphy
    175 Production of novel oils in plants Denis J Murphy We have now isolated the great majority of genes encoding unexpected ways. We have seen the isolation of some enzymes of storage oil biosynthesis in plants. In the past two potentially key genes that contribute both to the quantity years, particular progress has been made with acyltransferases, and quality of seed storage oils. There has also been an ketoacyl-acyl carrier protein synthetases and with desaturases increasing appreciation of the importance of fatty acids, and their relatives. In some cases, these enzymes have been re- not only as storage or structural components, but also act- engineered to create novel products. Nevertheless, the single or ing as, or giving rise to, important signalling molecules that multiple insertion of such transgenes into oil crops has not always regulate many aspects of plant development [1,2]. This led to the desired phenotype. We are only now beginning to illustrates the importance of ensuring that novel fatty acids appreciate some of the complexities of storage and membrane in transgenic oil crops are correctly targeted to the storage lipid formation, such as acyl group remodelling and the turnover oil and are hence unable to adversely affect membrane or of unusual fatty acids. This understanding will be vital for future signalling functions. The purpose of this article is to attempts at the rational engineering of transgenic oil crops. In review some of the recent progress in understanding the parallel with this, the domestication of plants already synthesising mechanism and regulation of storage oil formation in useful fatty acids should be considered as a real alternative to the plants, and how this may impact on its biotechnological transgenic approach to producing novel oil crops.
    [Show full text]