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Advanced genetic tools enable synthetic biology in the oleaginous microalgae Nannochloropsis sp.

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Abstract

Nannochloropsis is a genus of fast-growing microalgae that are regularly used for biotechnology applications. Nannochloropsis species have a high triacylglycerol content and their polar lipids are rich in the omega-3 long-chain polyunsaturated fatty acid, eicosapentaenoic acid. Placed in the heterokont lineage, the Nannochloropsis genus has a complex evolutionary history. Genome sequences are available for several species, and a number of transcriptomic datasets have been produced, making this genus a facile model for comparative genomics. There is a growing interest in Nannochloropsis species as models for the study of microalga lipid metabolism and as a chassis for synthetic biology. Recently, techniques for gene stacking, and targeted gene disruption and repression in the Nannochloropsis genus have been developed. These tools enable gene-specific, mechanistic studies and have already allowed the engineering of improved Nannochloropsis strains with superior growth, or greater bioproduction.

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References

  • Agapakis CM (2014) Designing synthetic biology. ACS Synth Biol 3:121–128

    CAS  PubMed  Google Scholar 

  • Ajjawi I, Verruto J, Aqui M, Soriaga LB, Coppersmith J, Kwok K, Peach L, Orchard E, Kalb R, Xu W, Carlson TJ, Francis K, Konigsfeld K, Bartalis J, Schultz A, Lambert W, Schwartz AS, Brown R, Moellering ER (2017) Lipid production in Nannochloropsis gaditanais doubled by decreasing expression of a single transcriptional regulator. Nat Biotechnol 35:647–652

    CAS  PubMed  Google Scholar 

  • Alboresi A, Perin G, Vitulo N, Diretto G, Block MA, Jouhet J, Meneghesso A, Valle G, Giuliano G, Maréchal E, Morosinotto T (2016) Light remodels lipid biosynthesis in Nannochloropsis gaditanaby modulating carbon partitioning between organelles. Plant Physiol. https://doi.org/10.1104/pp.16.0059900599

  • Alboresi A, Le Quiniou C, Yadav SKN, Scholz M, Meneghesso A, Gerotto C, Simionato D, Hippler M, Boekema EJ, Croce R, Morosinotto T (2017) Conservation of core complex subunits shaped the structure and function of photosystem I in the secondary endosymbiont alga Nannochloropsis gaditana. New Phytol 213:714–726

    CAS  PubMed  Google Scholar 

  • Andersen RA, Brett RW, Potter D, Sexton JP (1998) Phylogeny of the Eustigmatophyceae based upon 18S rDNA, with emphasis on Nannochloropsis. Protist 149:61–74

    CAS  PubMed  Google Scholar 

  • Andrianantoandro E, Basu S, Karig DK, Weiss R (2006) Synthetic biology: new engineering rules for an emerging discipline. Mol Syst Biol 2

  • Arnold AA, Genard B, Zito F, Tremblay R, Warschawski DE, Marcotte I (2015) Identification of lipid and saccharide constituents of whole microalgal cells by 13C solid-state NMR. Biochim et Biophys Acta (BBA) Biomembr 1848:369–377

    CAS  Google Scholar 

  • Ashworth J, Coesel S, Lee A, Armbrust EV, Orellana MV, Baliga NS (2013) Genome-wide diel growth state transitions in the diatom Thalassiosira pseudonana. Proc Natl Acad Sci 110:7518–7523

    CAS  PubMed  PubMed Central  Google Scholar 

  • Baek K, Kim DH, Jeong J, Sim SJ, Melis A, Kim J-S, Jin E, Bae S (2016) DNA-free two-gene knockout in Chlamydomonas reinhardtiivia CRISPR-Cas9 ribonucleoproteins. Sci Rep 6:30620

    CAS  PubMed  PubMed Central  Google Scholar 

  • Beacham TA, Ali ST (2016) Growth dependent silencing and resetting of DGA1 transgene in Nannochloropsis salina. Algal Res 14:65–71

    Google Scholar 

  • Bolte K, Bullmann L, Hempel F, Bozarth A, Zauner S, Maier U-G (2009) Protein targeting into secondary plastids. J Eukaryot Microbiol 56:9–15

    CAS  PubMed  Google Scholar 

  • Bowler C, Allen AE, Badger JH, Grimwood J, Jabbari K, Kuo A, Maheswari U, Martens C, Maumus F, Otillar RP, Rayko E, Salamov A, Vandepoele K, Beszteri B, Gruber A, Heijde M, Katinka M, Mock T, Valentin K, Verret F, Berges JA, Brownlee C, Cadoret JP, Chiovitti A, Choi CJ, Coesel S, De Martino A, Detter JC, Durkin C, Falciatore A, Fournet J, Haruta M, Huysman MJJ, Jenkins BD, Jiroutova K, Jorgensen RE, Joubert Y, Kaplan A, Kroger N, Kroth PG, La Roche J, Lindquist E, Lommer M, Martin-Jezequel V, Lopez PJ, Lucas S, Mangogna M, McGinnis K, Medlin LK, Montsant A, Oudot-Le Secq MP, Napoli C, Obornik M, Parker MS, Petit JL, Porcel BM, Poulsen N, Robison M, Rychlewski L, Rynearson TA, Schmutz J, Shapiro H, Siaut M, Stanley M, Sussman MR, Taylor AR, Vardi A, von Dassow P, Vyverman W, Willis A, Wyrwicz LS, Rokhsar DS, Weissenbach J, Armbrust EV, Green BR, Van De Peer Y, Grigoriev IV (2008) The Phaeodactylum genome reveals the evolutionary history of diatom genomes. Nature 456:239–244

    CAS  PubMed  Google Scholar 

  • Braun R, Farre EM, Schurr U, Matsubara S (2014) Effects of light and circadian clock on growth and chlorophyll accumulation of Nannochloropsis gaditana. J Phycol 50:515–525

    CAS  PubMed  Google Scholar 

  • Buitrago-Flórez FJ, Restrepo S, Riaño-Pachón DM (2014) Identification of transcription factor genes and their correlation with the high diversity of stramenopiles. PLoS One 9:e111841

    PubMed  PubMed Central  Google Scholar 

  • Cao S, Zhang X, Xu D, Fan X, Mou S, Wang Y, Ye N, Wang W (2013) A transthylakoid proton gradient and inhibitors induce a non-photochemical fluorescence quenching in unicellular algae Nannochloropsissp. FEBS Lett 587:1310–1315

    CAS  PubMed  Google Scholar 

  • Cermak T, Curtin SJ, Gil-Humanes J, Čegan R, Kono TJY, Konečná E, Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF (2017) A multi-purpose toolkit to enable advanced genome engineering in plants. Plant Cell. https://doi.org/10.1105/tpc.16.00922

    Article  PubMed  PubMed Central  Google Scholar 

  • Cha T-S, Chen C-F, Yee W, Aziz A, Loh S-H (2011) Cinnamic acid, coumarin and vanillin: alternative phenolic compounds for efficient Agrobacterium-mediated transformation of the unicellular green alga, Nannochloropsissp. J Microbiol Methods 84:430–434

    CAS  PubMed  Google Scholar 

  • Chauton MS, Winge P, Brembu T, Vadstein O, Bones AM (2013) Gene regulation of carbon fixation, storage, and utilization in the diatom Phaeodactylum tricornutumacclimated to light/dark cycles. Plant Physiol 161:1034–1048

    CAS  PubMed  Google Scholar 

  • Cheah YE, Albers SC, Peebles CAM (2013) A novel counter-selection method for markerless genetic modification in Synechocystissp. PCC 6803. Biotechnol Prog 29:23–30

    CAS  PubMed  Google Scholar 

  • Chen HL, Li SS, Huang R, Tsai HJ (2008) Conditional production of a functional fish growth hormone in the transgenic line of Nannochloropsis Oculata(Eustigmatophyceae). J Phycol 44:768–776

    CAS  PubMed  Google Scholar 

  • Chen J-W, Liu W-J, Hu D-X, Wang X, Balamurugan S, Alimujiang A, Yang W-D, Liu J-S, Li H-Y (2017) Identification ofa malonyl CoA-acyl carrier protein transacylase and its regulatory role in fatty acid biosynthesis in oleaginous microalga Nannochloropsis oceanica: Nannochloropsis MCAT. Biotechnol Appl Biochem

  • Chew KW, Yap JY, Show PL, Suan NH, Juan JC, Ling TC, Lee D-J, Chang J-S (2017) Microalgae biorefinery: High value products perspectives. Biores Technol 229:53–62

    CAS  Google Scholar 

  • Chukhutsina VU, Fristedt R, Morosinotto T, Croce R (2017) Photoprotection strategies of the alga Nannochloropsis gaditana. Biochim et Biophys Acta (BBA) Bioenerg 1858:544–552

    CAS  Google Scholar 

  • Coesel S, Mangogna M, Ishikawa T, Heijde M, Rogato A, Finazzi G, Todo T, Bowler C, Falciatore A (2009) Diatom PtCPF1 is a new cryptochrome/photolyase family member with DNA repair and transcription regulation activity. EMBO Rep 10:655–661

    CAS  PubMed  PubMed Central  Google Scholar 

  • Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, Jiang W, Marraffini LA, Zhang F (2013) Multiplex genome engineering using CRISPR/Cas systems. Science 339:819–823

    CAS  PubMed  PubMed Central  Google Scholar 

  • Corteggiani Carpinelli E, Telatin A, Vitulo N, Forcato C, D’Angelo M, Schiavon R, Vezzi A, Giacometti GM, Morosinotto T, Valle G (2013) Chromosome scale genome assembly and transcriptome profiling of Nannochloropsis gaditana in nitrogen depletion. Mol Plant

  • Daboussi F, Leduc S, Maréchal A, Dubois G, Guyot V, Perez-Michaut C, Amato A, Falciatore A, Juillerat A, Beurdeley M, Voytas DF, Cavarec L, Duchateau P (2014) Genome engineering empowers the diatom Phaeodactylum tricornutum for biotechnology. Nat Commun 5

  • De Riso V, Raniello R, Maumus F, Rogato A, Bowler C, Falciatore A (2009) Gene silencing in the marine diatom Phaeodactylum tricornutum. Nucleic Acids Res 37:e96-e96

    Google Scholar 

  • Dehoff P, Soriaga L (2014) Nannochloropsis kozak consensus sequence. US9309523 B2

  • Dehoff P, Akella S, Soriaga L (2014) Autonomous replication sequences and episomal DNA molecules. US9447422 B2

  • Diner RE, Bielinski VA, Dupont CL, Allen AE, Weyman PD (2016a) Refinement of the diatom episome maintenance sequence and improvement of conjugation-based DNA delivery methods. Front Bioeng Biotechnol 4:65

    PubMed  PubMed Central  Google Scholar 

  • Diner RE, Schwenck SM, McCrow JP, Zheng H, Allen AE (2016b) Genetic manipulation of competition for nitrate between heterotrophic bacteria and diatoms. Fronti Microbiol 7

  • Diner RE, Noddings CM, Lian NC, Kang AK, McQuaid JB, Jablanovic J, Espinoza JL, Nguyen NA, Anzelmatti MA, Jansson J, Bielinski VA, Karas BJ, Dupont CL, Allen AE, Weyman PD (2017) Diatom centromeres suggest a mechanism for nuclear DNA acquisition. Proc Natl Acad Sci 114:E6015–E6024

    CAS  PubMed  PubMed Central  Google Scholar 

  • Dolch L-J, Rak C, Perin G, Tourcier G, Broughton R, Leterrier M, Morosinotto T, Tellier F, Faure J-D, Falconet D, Jouhet J, Sayanova O, Beaudoin F, Maréchal E (2017) A palmitic acid elongase affects eicosapentaenoic acid and plastidial monogalactosyldiacylglycerol levels in Nannochloropsis. Plant Physiol 173:742–759

    CAS  PubMed  Google Scholar 

  • Fábregas J, Maseda A, Domínguez A, Ferreira M, Otero A (2002) Changes in the cell composition of the marine microalga, Nannochloropsis gaditana, during a light:dark cycle. Biotechnol Lett 24:1699–1703

    Google Scholar 

  • Fawley MW, Jameson I, Fawley KP (2015) The phylogeny of the genus Nannochloropsis (Monodopsidaceae, Eustigmatophyceae), with descriptions of N. australis sp. nov. and Microchloropsis gen nov. Phycologia 54:545–552

    CAS  Google Scholar 

  • Fortunato AE, Jaubert M, Enomoto G, Bouly J-P, Raniello R, Thaler M, Malviya S, Bernardes JS, Rappaport F, Gentili B, Huysman MJJ, Carbone A, Bowler C, d’Alcalà MR, Ikeuchi M, Falciatore A (2016) Diatom phytochromes reveal the existence of far-red-light-based sensing in the ocean. Plant Cell 28:616–628

    CAS  PubMed  PubMed Central  Google Scholar 

  • Galdzicki M, Clancy KP, Oberortner E, Pocock M, Quinn JY, Rodriguez CA, Roehner N, Wilson ML, Adam L, Anderson JC, Bartley BA, Beal J, Chandran D, Chen J, Densmore D, Endy D, Grunberg R, Hallinan J, Hillson NJ, Johnson JD, Kuchinsky A, Lux M, Misirli G, Peccoud J, Plahar HA, Sirin E, Stan GB, Villalobos A, Wipat A, Gennari JH, Myers CJ, Sauro HM (2014) The synthetic biology open language (SBOL) provides a community standard for communicating designs in synthetic biology. Nat Biotechnol 32:545–550

    CAS  PubMed  Google Scholar 

  • Gallie DR, Sleat DE, Watts JW, Turner PC, Wilson TMA (1987) The 5′-leader sequence of tobacco mosaic virus RNA enhances the expression of foreign gene transcripts in vitro and in vivo. Nucleic Acids Res 15:3257–3273

    CAS  PubMed  PubMed Central  Google Scholar 

  • Gee CW, Niyogi KK (2017) The carbonic anhydrase CAH1 is an essential component of the carbon-concentrating mechanism in Nannochloropsis oceanica. Proc Natl Acad Sci 114:4537–4542

    CAS  Google Scholar 

  • Gschloessl B, Guermeur Y, Cock JM (2008) HECTAR: a method to predict subcellular targeting in heterokonts. BMC Bioinformatics 9:393

    PubMed  PubMed Central  Google Scholar 

  • Hall MP, Unch J, Binkowski BF, Valley MP, Butler BL, Wood MG, Otto P, Zimmerman K, Vidugiris G, Machleidt T, Robers MB, Benink HA, Eggers CT, Slater MR, Meisenheimer PL, Klaubert DH, Fan F, Encell LP, Wood KV (2012) Engineered luciferase reporter from a deep sea shrimp utilizing a novel imidazopyrazinone substrate. ACS Chem Biol 7:1848–1857

    CAS  PubMed  PubMed Central  Google Scholar 

  • Hamilton ML, Haslam RP, Napier JA, Sayanova O (2014) Metabolic engineering of Phaeodactylum tricornutumfor the enhanced accumulation of omega-3 long chain polyunsaturated fatty acids. Metab Eng 22:3–9

    CAS  PubMed  PubMed Central  Google Scholar 

  • Hibberd DJ (1981) Notes on the taxonomy and nomenclature of the algal classes Eustigmatophyceae and Tribophyceae (synonym Xanthophyceae). Bot J Linn Soc 82:93–119

    Google Scholar 

  • Hildebrand M, Manandhar-Shrestha K, Abbriano R (2017) Effects of chrysolaminarin synthase knockdown in the diatom Thalassiosira pseudonana: Implications of reduced carbohydrate storage relative to green algae. Algal Res 23:66–77

    Google Scholar 

  • Hu J, Wang D, Li J, Jing G, Ning K, Xu J (2014) Genome-wide identification of transcription factors and transcription-factor binding sites in oleaginous microalgae Nannochloropsis. Sci Rep 4

  • Huang J, Liu J, Li Y, Chen F (2008) Isolation and characterization of the Phytoene Desaturase Gene as a Potential selective marker for genetic engineering of the astaxanthin-producing green alga Chlorella Zofingiensis(Chlorophyta). J Phycol 44:684–690

    CAS  PubMed  Google Scholar 

  • Huysman MJ, Martens C, Vandepoele K, Gillard J, Rayko E, Heijde M, Bowler C, Inze D, Van de Peer Y, De Veylder L, Vyverman W (2010) Genome-wide analysis of the diatom cell cycle unveils a novel type of cyclins involved in environmental signaling. Genome Biol 11:R17

    PubMed  PubMed Central  Google Scholar 

  • Huysman MJ, Fortunato AE, Matthijs M, Costa BS, Vanderhaeghen R, Van den Daele H, Sachse M, Inze D, Bowler C, Kroth PG, Wilhelm C, Falciatore A, Vyverman W, De Veylder L (2013) AUREOCHROME1a-mediated induction of the diatom-specific cyclin dsCYC2 controls the onset of cell division in diatoms (Phaeodactylum tricornutum). Plant Cell 25:215–228

    CAS  PubMed  PubMed Central  Google Scholar 

  • Iwai M, Hori K, Sasaki-Sekimoto Y, Shimojima M, Ohta H (2015) Manipulation of oil synthesis in Nannochloropsis strain NIES-2145 with a phosphorus starvation–inducible promoter from Chlamydomonas reinhardtii. Front Microbiol 6

  • Janouskovec J, Horak A, Obornik M, Lukes J, Keeling PJ (2010) A common red algal origin of the apicomplexan, dinoflagellate, and heterokont plastids. Proc Natl Acad Sci 107:10949–10954

    PubMed  PubMed Central  Google Scholar 

  • Jeong SW, Nam SW, HwangBo K, Jeong WJ, Jeong B-r, Chang YK, Park Y-I (2017) Transcriptional regulation of cellulose biosynthesis during the early phase of nitrogen deprivation in Nannochloropsis salina. Sci Rep 7

  • Jia J, Han D, Gerken HG, Li Y, Sommerfeld M, Hu Q, Xu J (2015) Molecular mechanisms for photosynthetic carbon partitioning into storage neutral lipids in Nannochloropsis oceanicaunder nitrogen-depletion conditions. Algal Res 7:66–77

    Google Scholar 

  • Jia B, Zheng Y, Xiao K, Wu M, Lei Y, Huang Y, Hu Z (2016) A vector for multiple gene co-expression in Chlamydomonas reinhardtii. Algal Res 20:53–56

    Google Scholar 

  • Jinkerson RE, Radakovits R, Posewitz MC (2013) Genomic insights from the oleaginous model alga Nannochloropsis gaditana. Bioengineered 4:37–43

    PubMed  PubMed Central  Google Scholar 

  • Kang NK, Choi G-G, Kim EK, Shin S-E, Jeon S, Park MS, Jeong KJ, Jeong B-r, Chang YK, Yang J-W, Lee B (2015a) Heterologous overexpression of sfCherry fluorescent protein in Nannochloropsis salina. Biotechnol Rep 8:10–15

    Google Scholar 

  • Kang NK, Jeon S, Kwon S, Koh HG, Shin S-E, Lee B, Choi G-G, Yang J-W, Jeong B-r, Chang YK (2015b) Effects of overexpression of a bHLH transcription factor on biomass and lipid production in Nannochloropsis salina. Biotechnol Biofuels 8

  • Kang NK, Kim EK, Kim YU, Lee B, Jeong W-J, Jeong B-r, Chang YK (2017) Increased lipid production by heterologous expression of AtWRI1 transcription factor in Nannochloropsis salina. Biotechnol Biofuels 10

  • Karas BJ, Diner RE, Lefebvre SC, McQuaid J, Phillips AP, Noddings CM, Brunson JK, Valas RE, Deerinck TJ, Jablanovic J, Gillard JT, Beeri K, Ellisman MH, Glass JI, Hutchison CA, 3rd, Smith HO, Venter JC, Allen AE, Dupont CL, Weyman PD (2015) Designer diatom episomes delivered by bacterial conjugation. Nat Commun 6:6925

    CAS  PubMed  Google Scholar 

  • Kaye Y, Grundman O, Leu S, Zarka A, Zorin B, Didi-Cohen S, Khozin-Goldberg I, Boussiba S (2015) Metabolic engineering toward enhanced LC-PUFA biosynthesis in Nannochloropsis oceanica: Overexpression of endogenous ∆12 desaturase driven by stress-inducible promoter leads to enhanced deposition of polyunsaturated fatty acids in TAG. Algal Res 11:387–398

    Google Scholar 

  • Keeling PJ (2009) Chromalveolates and the evolution of plastids by secondary endosymbiosis. J Eukaryot Microbiol 56:1–8

    CAS  PubMed  Google Scholar 

  • Kilian O, Benemann CSE, Niyogi KK, Vick B (2011) High-efficiency homologous recombination in the oil-producing alga Nannochloropsissp. Proc Natl Acad Sci USA 108:21265–21269

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kindle KL, Schnell RA, Fernández E, Lefebvre PA (1989) Stable nuclear transformation of Chlamydomonas using the Chlamydomonas gene for nitrate reductase. J Cell Biol 109:2589–2601

    CAS  PubMed  Google Scholar 

  • Kleinstiver BP, Pattanayak V, Prew MS, Tsai SQ, Nguyen NT, Zheng Z, Joung JK (2016) High-fidelity CRISPR–Cas9 nucleases with no detectable genome-wide off-target effects. Nature 529:490–495

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kroth P, Strotmann H (1999) Diatom plastids: secondary endocytobiosis, plastid genome and protein import. Physiol Plant 107:136–141

    CAS  Google Scholar 

  • Kwon S, Kang NK, Koh HG, Shin S-E, Lee B, Jeong B-r, Chang YK (2017) Enhancement of biomass and lipid productivity by overexpression of a bZIP transcription factor in Nannochloropsis salina: Engineering of Nannochloropsis with bZIP TF. Biotechnol Bioeng

  • Li F, Gao D, Hu H (2014a) High-efficiency nuclear transformation of the oleaginous marine Nannochloropsisspecies using PCR product. Biosci Biotechnol Biochem 78:812–817

    CAS  PubMed  Google Scholar 

  • Li J, Han D, Wang D, Ning K, Jia J, Wei L, Jing X, Huang S, Chen J, Li Y, Hu Q, Xu J (2014b) Choreography of transcriptomes and lipidomes of Nannochloropsisreveals the mechanisms of oil synthesis in microalgae. Plant Cell 26:1645–1665

    CAS  PubMed  PubMed Central  Google Scholar 

  • Li D-W, Cen S-Y, Liu Y-H, Balamurugan S, Zheng X-Y, Alimujiang A, Yang W-D, Liu J-S, Li H-Y (2016a) A type 2 diacylglycerol acyltransferase accelerates the triacylglycerol biosynthesis in heterokont oleaginous microalga Nannochloropsis oceanica. J Biotechnol 229:65–71

    CAS  PubMed  Google Scholar 

  • Li X, Zhang R, Patena W, Gang SS, Blum SR, Ivanova N, Yue R, Robertson JM, Lefebvre PA, Fitz-Gibbon ST, Grossman AR, Jonikas MC (2016b) An indexed, mapped mutant library enables reverse genetics studies of biological processes in Chlamydomonas reinhardtii. Plant Cell 28:367–387

    CAS  PubMed  PubMed Central  Google Scholar 

  • Lin G, Wang Y, Guo L, Ding H, Hu Y, Liang S, Zhang Z, Yang G (2017) Verification of mutagen function of Zeocin in Nannochloropsis oceanicathrough transcriptome analysis. J Ocean Univ China 16:501–508

    CAS  Google Scholar 

  • Loira N, Mendoza S, Paz Cortés M, Rojas N, Travisany D, Genova AD, Gajardo N, Ehrenfeld N, Maass A (2017) Reconstruction of the microalga Nannochloropsis salina genome-scale metabolic model with applications to lipid production. BMC Syst Biol 11

  • Lubián L (1982) Nannochloropsis gaditana sp. nov., una nueva Eustigmatophyceae marina. Lazaroa 4

  • Ma X, Pan K, Zhang L, Zhu B, Yang G, Zhang X (2016) Genetic transformation of Nannochloropsis oculatawith a bacterial phleomycin resistance gene as dominant selective marker. J Ocean Univ China 15:351–356

    CAS  Google Scholar 

  • Ma X, Yao L, Yang B, Lee YK, Chen F, Liu J (2017) RNAi-mediated silencing of a pyruvate dehydrogenase kinase enhances triacylglycerol biosynthesis in the oleaginous marine alga Nannochloropsis salina. Sci Rep 7

  • Mann M, Serif M, Jakob T, Kroth PG, Wilhelm C (2017) PtAUREO1a and PtAUREO1b knockout mutants of the diatom Phaeodactylum tricornutumare blocked in photoacclimation to blue light. J Plant Physiol 217:44–48

    CAS  PubMed  Google Scholar 

  • McCarthy JK, Smith SR, McCrow JP, Tan M, Zheng H, Beeri K, Roth R, Lichtle C, Goodenough U, Bowler CP, Dupont CL, Allen AE (2017) Nitrate reductase knockout uncouples nitrate transport from nitrate assimilation and drives repartitioning of carbon flux in a model pennate diatom. Plant Cell 29:2047–2070

    CAS  PubMed  PubMed Central  Google Scholar 

  • Meng Y, Jiang J, Wang H, Cao X, Xue S, Yang Q, Wang W (2015) The characteristics of TAG and EPA accumulation in Nannochloropsis oceanicaIMET1 under different nitrogen supply regimes. Biores Technol 179:483–489

    CAS  Google Scholar 

  • Miller R, Wu G, Deshpande RR, Vieler A, Gartner K, Li X, Moellering ER, Zauner S, Cornish AJ, Liu B, Bullard B, Sears BB, Kuo MH, Hegg EL, Shachar-Hill Y, Shiu SH, Benning C (2010) Changes in transcript abundance in Chlamydomonas reinhardtiifollowing nitrogen deprivation predict diversion of metabolism. Plant Physiol 154:1737–1752

    CAS  PubMed  PubMed Central  Google Scholar 

  • Moellering ER, Benning C (2010) RNA interference silencing of a major lipid droplet protein affects lipid droplet size in Chlamydomonas reinhardtii. Eukaryot Cell 9:97–106

    CAS  PubMed  Google Scholar 

  • Moog D, Stork S, Reislöhner S, Grosche C, Maier U-G (2015) In vivo localization studies in the stramenopile alga Nannochloropsis oceanica. Protist 166:161–171

    CAS  PubMed  Google Scholar 

  • Mühlroth A, Li K, Røkke G, Winge P, Olsen Y, Hohmann-Marriott M, Vadstein O, Bones A (2013) Pathways of lipid metabolism in marine algae, co-expression network, bottlenecks and candidate genes for enhanced production of EPA and DHA in species of chromista. Mar Drugs 11:4662–4697

    PubMed  PubMed Central  Google Scholar 

  • Mühlroth A, Winge P, Assimi AE, Jouhet J, Marechal E, Hohmann-Marriott MF, Vadstein O, Bones AM (2017) Mechanisms of phosphorus acquisition and lipid class remodelling under P limitation in a marine microalga. Plant Physiology

  • Müller N, Wenzel S, Zou Y, Künzel S, Sasso S, Weiß D, Prager K, Grossman A, Kottke T, Mittag M (2017) A plant cryptochrome controls key features of the Chlamydomonascircadian clock and its life cycle. Plant Physiol 174:185–201

    PubMed  PubMed Central  Google Scholar 

  • Murakami R, Hashimoto H (2009) Unusual nuclear division in Nannochloropsis oculata(Eustigmatophyceae, Heterokonta) which may ensure faithful transmission of secondary plastids. Protist 160:41–49

    PubMed  Google Scholar 

  • Nobusawa T, Hori K, Mori H, Kurokawa K, Ohta H (2017) Differently localized lysophosphatidic acid acyltransferases crucial for triacylglycerol biosynthesis in the oleaginous alga Nannochloropsis. Plant J 90:547–559

    CAS  PubMed  Google Scholar 

  • Nymark M, Sharma AK, Sparstad T, Bones AM, Winge P (2016) A CRISPR/Cas9 system adapted for gene editing in marine algae. Sci Rep 6:24951

    CAS  PubMed  PubMed Central  Google Scholar 

  • Pal D, Khozin-Goldberg I, Didi-Cohen S, Solovchenko A, Batushansky A, Kaye Y, Sikron N, Samani T, Fait A, Boussiba S (2013) Growth, lipid production and metabolic adjustments in the euryhaline eustigmatophyteNannochloropsis oceanicaCCALA 804 in response to osmotic downshift. Appl Microbiol Biotechnol 97:8291–8306

    CAS  PubMed  Google Scholar 

  • Perin G, Bellan A, Segalla A, Meneghesso A, Alboresi A, Morosinotto T (2015) Generation of random mutants to improve light-use efficiency of Nannochloropsis gaditanacultures for biofuel production. Biotechnol Biofuels 8:161

    PubMed  PubMed Central  Google Scholar 

  • Plucinak TM, Horken KM, Jiang W, Fostvedt J, Nguyen ST, Weeks DP (2015) Improved and versatile viral 2A platforms for dependable and inducible high-level expression of dicistronic nuclear genes in Chlamydomonas reinhardtii. Plant J 82:717–729

    CAS  PubMed  Google Scholar 

  • Poliner E, Panchy N, Newton L, Wu G, Lapinsky A, Bullard B, Zienkiewicz A, Benning C, Shiu S-H, Farré EM (2015) Transcriptional coordination of physiological responses in Nannochloropsis oceanicaCCMP1779 under light/dark cycles. Plant J 83:1097–1113

    CAS  PubMed  Google Scholar 

  • Poliner E, Pulman JA, Zienkiewicz K, Childs K, Benning C, Farre EM (2017) A toolkit for Nannochloropsis oceanica CCMP1779 enables gene stacking and genetic engineering of the eicosapentaenoic acid pathway for enhanced long-chain polyunsaturated fatty acid production. Plant Biotechnol J

  • Radakovits R, Jinkerson RE, Fuerstenberg SI, Tae H, Settlage RE, Boore JL, Posewitz MC (2012) Draft genome sequence and genetic transformation of the oleaginous alga Nannochloropsis gaditana. Nat Commun 3:686

    PubMed  Google Scholar 

  • Ran FA, Hsu Patrick D, Lin C-Y, Gootenberg Jonathan S, Konermann S, Trevino AE, Scott David A, Inoue A, Matoba S, Zhang Y, Zhang F (2013) Double nicking by RNA-guided CRISPR Cas9 for enhanced genome editing specificity. Cell 154:1380–1389

    CAS  PubMed  PubMed Central  Google Scholar 

  • Rayko E, Maumus F, Maheswari U, Jabbari K, Bowler C (2010) Transcription factor families inferred from genome sequences of photosynthetic stramenopiles. New Phytol 188:52–66

    CAS  PubMed  Google Scholar 

  • Rodolfi L, Chini Zittelli G, Bassi N, Padovani G, Biondi N, Bonini G, Tredici MR (2009) Microalgae for oil: Strain selection, induction of lipid synthesis and outdoor mass cultivation in a low-cost photobioreactor. Biotechnol Bioeng 102:100–112

    CAS  PubMed  Google Scholar 

  • Rohr J, Sarkar N, Balenger S, Jeong B-r, Cerutti H (2004) Tandem inverted repeat system for selection of effective transgenic RNAi strains inChlamydomonas: Tandem inverted repeat system for efficient RNAi. Plant J 40:611–621

    CAS  PubMed  Google Scholar 

  • Schellenberger Costa B, Sachse M, Jungandreas A, Bartulos CR, Gruber A, Jakob T, Kroth PG, Wilhelm C (2013) Aureochrome 1a is involved in the photoacclimation of the diatom Phaeodactylum tricornutum. PLoS One 8:e74451

    PubMed  PubMed Central  Google Scholar 

  • Schneider JC, Roessler P (1994) Radiolabeling studies of lipids and fatty acids in Nannochloropsis(Eustigmatophyceae), an oleaginous marine alga1. J Phycol 30:594–598

    CAS  Google Scholar 

  • Schneider JC, Livne A, Sukenik A, Roessler PG (1995) A mutant of Nannochloropsisdeficient in eicosapentaenoic acid production. Phytochemistry 40:807–814

    CAS  Google Scholar 

  • Scholz MJ, Weiss TL, Jinkerson RE, Jing J, Roth R, Goodenough U, Posewitz MC, Gerken HG (2014) Ultrastructure and Composition of the Nannochloropsis gaditanaCell Wall. Eukaryot Cell 13:1450–1464

    PubMed  PubMed Central  Google Scholar 

  • Schonknecht G, Chen WH, Ternes CM, Barbier GG, Shrestha RP, Stanke M, Brautigam A, Baker BJ, Banfield JF, Garavito RM, Carr K, Wilkerson C, Rensing SA, Gagneul D, Dickenson NE, Oesterhelt C, Lercher MJ, Weber APM (2013) Gene transfer from bacteria and archaea facilitated evolution of an extremophilic eukaryote. Science 339:1207–1210

    PubMed  Google Scholar 

  • Sharma P, Yan F, Doronina VA, Escuin-Ordinas H, Ryan MD, Brown JD (2012) 2A peptides provide distinct solutions to driving stop-carry on translational recoding. Nucleic Acids Res 40:3143–3151

    CAS  PubMed  Google Scholar 

  • Sheehan J, Dunahay T, Benemann J, Roessler P (1998) Look back at the U.S. Department of Energy’s Aquatic Species Program: Biodiesel from Algae; Close-Out Report

  • Shih C-H, Chen H-Y, Lee H-C, Tsai H-J (2015) Purple chromoprotein gene serves as a new selection marker for transgenesis of the microalga Nannochloropsis oculata. Plos One 10:e0120780

    PubMed  PubMed Central  Google Scholar 

  • Simionato D, Block MA, La Rocca N, Jouhet J, Marechal E, Finazzi G, Morosinotto T (2013) The response of Nannochloropsis gaditana to nitrogen starvation includes de novo biosynthesis of triacylglycerols, a decrease of chloroplast galactolipids, and reorganization of the photosynthetic apparatus. Eukaryot Cell 12:665–676

    CAS  PubMed  PubMed Central  Google Scholar 

  • Slaymaker IM, Gao L, Zetsche B, Scott DA, Yan WX, Zhang F (2016) Rationally engineered Cas9 nucleases with improved specificity. Science 351:84–88

    CAS  PubMed  Google Scholar 

  • Starkenburg SR, Kwon KJ, Jha RK, McKay C, Jacobs M, Chertkov O, Twary S, Rocap G, Cattolico R (2014) A pangenomic analysis of the Nannochloropsisorganellar genomes reveals novel genetic variations in key metabolic genes. BMC Genom 15:212

    Google Scholar 

  • Stormo GD (2000) DNA binding sites: representation and discovery. Bioinformatics 16:16–23

    CAS  PubMed  Google Scholar 

  • Suda S, Atsumi M, Miyashita H (2002) Taxonomic characterization of a marine Nannochloropsis species, N. oceanicasp. nov. (Eustigmatophyceae). Phycologia 41:273–279

    Google Scholar 

  • Sukenik A, Carmeli Y (1990) Lipid synthesis and fatty acid composition in NannochloropsisSp. (Eustigmatophyceae) grown in a light-dark cycle. J Phycol 26:463–469

    CAS  Google Scholar 

  • Sukenik A, Carmeli Y, Berner T (1989) Regulation of fatty acid composition by irradiance level in the Eustigmatophyte NannochloropsisSp. J Phycol 25:686–692

    CAS  Google Scholar 

  • Suzuki K, Kimura T, Shinoda H, Bai G, Daniels MJ, Arai Y, Nakano M, Nagai T (2016) Five colour variants of bright luminescent protein for real-time multicolour bioimaging. Nat Commun 7:13718

    CAS  PubMed  PubMed Central  Google Scholar 

  • Thiriet-Rupert S, Carrier G, Chénais B, Trottier C, Bougaran G, Cadoret J-P, Schoefs B, Saint-Jean B (2016) Transcription factors in microalgae: genome-wide prediction and comparative analysis.BMC Genom 17

  • Tian J, Zheng M, Yang G, Zheng L, Chen J, Yang B (2013) Cloning and stress-responding expression analysis of malonyl CoA-acyl carrier protein transacylase gene of Nannochloropsis gaditana. Gene 530:33–38

    CAS  PubMed  Google Scholar 

  • Trentacoste EM, Shrestha RP, Smith SR, Gle C, Hartmann AC, Hildebrand M, Gerwick WH (2013) Metabolic engineering of lipid catabolism increases microalgal lipid accumulation without compromising growth. Proc Natl Acad Sci 110:19748–19753

    CAS  PubMed  PubMed Central  Google Scholar 

  • Tsai CH, Warakanont J, Takeuchi T, Sears BB, Moellering ER, Benning C (2014) The protein compromised hydrolysis of triacylglycerols 7 (CHT7) acts as a repressor of cellular quiescence in Chlamydomonas. Proc Natl Acad Sci USA 111:15833–15838

    CAS  PubMed  PubMed Central  Google Scholar 

  • Umetani I, Kunugi M, Yokono M, Takabayashi A, Tanaka A (2017) Evidence of the supercomplex organization of photosystem II and light-harvesting complexes in Nannochloropsis granulata. Photosynth Res

  • Unkefer CJ, Sayre RT, Magnuson JK, Anderson DB, Baxter I, Blaby IK, Brown JK, Carleton M, Cattolico RA, Dale T, Devarenne TP, Downes CM, Dutcher SK, Fox DT, Goodenough U, Jaworski J, Holladay JE, Kramer DM, Koppisch AT, Lipton MS, Marrone BL, McCormick M, Molnár I, Mott JB, Ogden KL, Panisko EA, Pellegrini M, Polle J, Richardson JW, Sabarsky M, Starkenburg SR, Stormo GD, Teshima M, Twary SN, Unkefer PJ, Yuan JS, Olivares JA (2017) Review of the algal biology program within the national alliance for advanced biofuels and bioproducts. Algal Res 22:187–215

    Google Scholar 

  • Vieler A, Wu G, Tsai C-H, Bullard B, Cornish AJ, Harvey C, Reca I-B, Thornburg C, Achawanantakun R, Buehl CJ, Campbell MS, Cavalier D, Childs KL, Clark TJ, Deshpande R, Erickson E, Armenia Ferguson A, Handee W, Kong Q, Li X, Liu B, Lundback S, Peng C, Roston RL, Sanjaya, Simpson JP, TerBush A, Warakanont J, Zäuner S, Farre EM, Hegg EL, Jiang N, Kuo M-H, Lu Y, Niyogi KK, Ohlrogge J, Osteryoung KW, Shachar-Hill Y, Sears BB, Sun Y, Takahashi H, Yandell M, Shiu S-H, Benning C (2012) Genome, functionalgene annotation, and nuclear transformation of the heterokont oleaginous alga Nannochloropsis oceanicaCCMP1779. PLoS Genet 8:e1003064

    CAS  PubMed  PubMed Central  Google Scholar 

  • Wang D, Ning K, Li J, Hu J, Han D, Wang H, Zeng X, Jing X, Zhou Q, Su X, Chang X, Wang A, Wang W, Jia J, Wei L, Xin Y, Qiao Y, Huang R, Chen J, Han B, Yoon K, Hill RT, Zohar Y, Chen F, Hu Q, Xu J (2014) Nannochloropsisgenomes reveal evolution of microalgal oleaginous traits. PLoS Genet 10:e1004094

    PubMed  PubMed Central  Google Scholar 

  • Wang Q, Lu Y, Xin Y, Wei L, Huang S, Xu J (2016) Genome editing of model oleaginous microalgae Nannochloropsisspp. by CRISPR/Cas9. Plant J 88:1071–1081

    CAS  PubMed  Google Scholar 

  • Wei L, Xin Y, Wang D, Jing X, Zhou Q, Su X, Jia J, Ning K, Chen F, Hu Q, Xu J (2013) Nannochloropsisplastid and mitochondrial phylogenomes reveal organelle diversification mechanism and intragenus phylotyping strategy in microalgae. BMC Genom 14:534

    CAS  Google Scholar 

  • Wei H, Shi Y, Ma X, Pan Y, Hu H, Li Y, Luo M, Gerken H, Liu J (2017a) A type-I diacylglycerol acyltransferase modulates triacylglycerol biosynthesis and fatty acid composition in the oleaginous microalga, Nannochloropsis oceanica. Biotechnol Biofuels 10

  • Wei L, Wang Q, Xin Y, Lu Y, Xu J (2017b) Enhancing photosynthetic biomass productivity of industrial oleaginous microalgae by overexpression of RuBisCO activase. Algal Res 27:366–375

    Google Scholar 

  • Wei L, Xin Y, Wang Q, Yang J, Hu H, Xu J (2017c) RNAi-based targeted gene knockdown in the model oleaginous microalgae Nannochloropsis oceanica. Plant J 89:1236–1250

    CAS  PubMed  Google Scholar 

  • Wingender E (2008) The TRANSFAC project as an example of framework technology that supports the analysis of genomic regulation. Brief Bioinform 9:326–332

    CAS  PubMed  Google Scholar 

  • Work VH, Radakovits R, Jinkerson RE, Meuser JE, Elliott LG, Vinyard DJ, Laurens LML, Dismukes GC, Posewitz MC (2010) Increased lipid accumulation in the Chlamydomonas reinhardtiista7-10 starchless isoamylase mutant and increased carbohydrate synthesis in complemented strains. Eukaryot Cell 9:1251–1261

    CAS  PubMed  PubMed Central  Google Scholar 

  • Xiao Y, Zhang J, Cui J, Yao X, Sun Z, Feng Y, Cui Q (2015) Simultaneous accumulation of neutral lipids and biomass in Nannochloropsis oceanicaIMET1 under high light intensity and nitrogen replete conditions. Algal Res 11:55–62

    Google Scholar 

  • Xie Y, Wang D, Lan F, Wei G, Ni T, Chai R, Liu D, Hu S, Li M, Li D, Wang H, Wang Y (2017) An episomal vector-based CRISPR/Cas9 system for highly efficient gene knockout in human pluripotent stem cells. Sci Rep 7

  • Xin Y, Lu Y, Lee Y-Y, Wei L, Jia J, Wang Q, Wang D, Bai F, Hu H, Hu Q, Liu J, Li Y, Xu J (2017) Producing designer oils in industrial microalgae by rational modulation of co-evolving type-2 diacylglycerol acyltransferases. Mol Plant

  • Xue Z, Sharpe PL, Hong S-P, Yadav NS, Xie D, Short DR, Damude HG, Rupert RA, Seip JE, Wang J, Pollak DW, Bostick MW, Bosak MD, Macool DJ, Hollerbach DH, Zhang H, Arcilla DM, Bledsoe SA, Croker K, McCord EF, Tyreus BD, Jackson EN, Zhu Q (2013) Production of omega-3 eicosapentaenoic acid by metabolic engineering of Yarrowia lipolytica. Nat Biotechnol 31:734–740

    CAS  PubMed  Google Scholar 

  • Xue J, Niu Y-F, Huang T, Yang W-D, Liu J-S, Li H-Y (2015) Genetic improvement of the microalga Phaeodactylum tricornutumfor boosting neutral lipid accumulation. Metab Eng 27:1–9

    PubMed  Google Scholar 

  • Yaakob Z, Ali E, Zainal A, Mohamad M, Takriff M (2014) An overview: biomolecules from microalgae for animal feed and aquaculture. J Biol Res Thessaloniki 21:6

    Google Scholar 

  • Zaslavskaia LA, Lippmeier JC, Kroth PG, Grossman AR, Apt KE (2001) Transformation of the diatom Phaeodactylum tricornutum(Bacillariophyceae) with a variety of selectable marker and reporter genes. J Phycol 36:379–386

    Google Scholar 

  • Zienkiewicz K, Zienkiewicz A, Poliner E, Du Z-Y, Vollheyde K, Herrfurth C, Marmon S, Farré EM, Feussner I, Benning C (2017) Nannochloropsis, a rich source of diacylglycerol acyltransferases for engineering of triacylglycerol content in different hosts. Biotechnol Biofuels 10

  • Zou N, Zhang C, Cohen Z, Richmond A (2000) Production of cell mass and eicosapentaenoic acid (EPA) in ultrahigh cell density cultures of Nannochloropsissp. (Eustigmatophyceae). Eur J Phycol 35:127–133

    Google Scholar 

  • Zou Y, Wenzel S, Müller N, Prager K, Jung E-M, Kothe E, Kottke T, Mittag M (2017) An animal-like cryptochrome controls the Chlamydomonassexual cycle. Plant Physiol 174:1334–1347

    CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by a National Science Foundation grant (IOS-1354721) to EF. In addition, parts of this work were supported by the Division of Chemical Sciences, Geosciences and Biosciences, Office of Basic Energy Sciences of the United States Department of Energy (DE-FG02-91ER20021) and MSU-AgBioResearch to CB.

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  1. Cell and Molecular Biology Program, Michigan State University, East Lansing, MI, USA

    Eric Poliner

  2. MSU-DOE Plant Research Laboratory, Michigan State University, East Lansing, MI, USA

    Eric Poliner & Christoph Benning

  3. Department of Plant Biology, Michigan State University, East Lansing, MI, USA

    Eva M. Farré & Christoph Benning

  4. Department of Biochemistry and Molecular Biology, Michigan State University, East Lansing, MI, USA

    Christoph Benning

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Correspondence to Christoph Benning.

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Poliner, E., Farré, E.M. & Benning, C. Advanced genetic tools enable synthetic biology in the oleaginous microalgae Nannochloropsis sp.. Plant Cell Rep 37, 1383–1399 (2018). https://doi.org/10.1007/s00299-018-2270-0

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