[en] Nitrogen starvation induced changes in carbohydrate and lipid content is described in several algal species. Although these phenotypic changes are desirable, such manipulations also significantly deteriorate culture health, ultimately halting growth. To optimize biofuel production from algae, it is desirable to induce lipid accumulation without compromising cell growth and survival. In this study, we utilized an 8-plex iTRAQ-based proteomic approach to assess the model alga Chlamydomonas reinhardtii CCAP 11/32CW15+ under nitrogen starvation. First-dimension fractionation was conducted using HILIC and SCX. A total of 587 proteins were identified (≥3 peptides) of which 71 and 311 were differentially expressed at significant levels (p<0.05), during nitrogen stress induced carbohydrate and lipid production, respectively. Forty-seven percent more changes with significance were observed with HILIC compared to SCX. Several trends were observed including increase in energy metabolism, decrease in translation machinery, increase in cell wall production and a change of balance between photosystems I and II. These findings point to a severely compromised system where lipid is accumulated at the expense of normal functioning of the organism, suggesting that a more informed and controlled method of lipid induction than gross nutrient manipulation would be needed for development of sustainable processes.
Disciplines :
Biotechnology
Author, co-author :
LONGWORTH, Joseph ; ChELSI Institute, Department of Chemical and Biological Engineering, The University of Sheffield, Mappin Street, Sheffield, S1 3JD, United Kingdom
Noirel, Josselin; ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, S1 3JD, United Kingdom
Pandhal, Jagroop; ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, S1 3JD, United Kingdom
Wright, Phillip C; ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, S1 3JD, United Kingdom
Vaidyanathan, Seetharaman; ChELSI Institute, Department of Chemical and Biological Engineering, University of Sheffield, Sheffield, S1 3JD, United Kingdom
External co-authors :
yes
Language :
English
Title :
HILIC- and SCX-based quantitative proteomics of Chlamydomonas reinhardtii during nitrogen starvation induced lipid and carbohydrate accumulation.
Brennan, L.; Owende, P. Biofuels from microalgae-A review of technologies for production, processing, and extractions of biofuels and co-products Renewable Sustainable Energy Rev. 2010, 14 (2) 557-577
Chisti, Y. Biodiesel from microalgae beats bioethanol Trends Biotechnol. 2008, 26 (3) 126-131
Chisti, Y. Biodiesel from microalgae Biotechnol. Adv. 2007, 25 (3) 294-306
Sheehan, J.; Dunahay, T.; Benemann, J.; Roessler, P. A Look Back at the US Department of Energy's Aquatic Species Program: Biodiesel from Algae; NERL Report TP-580-24190; National Renewable Energy Laboratory: Golden, CO, 1998.
Gallagher, B. J. The economics of producing biodiesel from algae Renewable Energy 2011, 36 (1) 158-162
Radakovits, R.; Jinkerson, R. E.; Darzins, A.; Posewitz, M. C. Genetic engineering of algae for enhanced biofuel production Eukaryotic Cell 2010, 9 (4) 486-501
Shifrin, N. S.; Chisholm, S. W. Phytoplankton lipids: Interspecific differences and effects of nitrate, silicate and light-dark cycle J. Phycol. 1981, 17 (4) 374-384
Griffiths, M. J.; Harrison, S. T. L. Lipid productivity as a key characteristic for choosing algal species for biodiesel production J. Appl. Phycol. 2009, 21 (5) 493-507
Hejazi, M. A.; Wijffels, R. H. Milking of microalgae Trends Biotechnol. 2004, 22 (4) 189-194
Ramachandra, T. V.; Mahapatra, D. M.; B, K.; Gordon, R. Milking diatoms for sustainable energy: Biochemical engineering versus gasoline-secreting diatom solar panels Ind. Eng. Chem. Res. 2009, 48 (19) 8769-8788
Merchant, S. S.; Prochnik, S. E.; Vallon, O.; Harris, E. H.; Karpowicz, S. J.; Witman, G. B.; Terry, A.; Salamov, A.; Fritz-Laylin, L. K.; Maréchal-Drouard, L.; Marshall, W. F.; Qu, L.-H.; Nelson, D. R.; Sanderfoot, A. A.; Spalding, M. H.; Kapitonov, V. V.; Ren, Q.; Ferris, P.; Lindquist, E.; Shapiro, H.; Lucas, S. M.; Grimwood, J.; Schmutz, J.; Cardol, P.; Cerutti, H.; Chanfreau, G.; Chen, C.-L.; Cognat, V.; Croft, M. T.; Dent, R.; Dutcher, S.; Fernández, E.; Ferris, P.; Fukuzawa, H.; González-Ballester, D.; González-Halphen, D.; Hallmann, A.; Hanikenne, M.; Hippler, M.; Inwood, W.; Jabbari, K.; Kalanon, M.; Kuras, R.; Lefebvre, P. A.; Lemaire, S. D.; Lobanov, A. V.; Lohr, M.; Manuell, A.; Meier, I.; Mets, L.; Mittag, M.; Mittelmeier, T.; Moroney, J. V.; Moseley, J.; Napoli, C.; Nedelcu, A. M.; Niyogi, K.; Novoselov, S. V.; Paulsen, I. T.; Pazour, G.; Purton, S.; Ral, J.-P.; Riaño-Pachón, D. M.; Riekhof, W.; Rymarquis, L.; Schroda, M.; Stern, D.; Umen, J.; Willows, R.; Wilson, N.; Zimmer, S. L.; Allmer, J.; Balk, J.; Bisova, K.; Chen, C.-J.; Elias, M.; Gendler, K.; Hauser, C.; Lamb, M. R.; Ledford, H.; Long, J. C.; Minagawa, J.; Page, M. D.; Pan, J.; Pootakham, W.; Roje, S.; Rose, A.; Stahlberg, E.; Terauchi, A. M.; Yang, P.; Ball, S.; Bowler, C.; Dieckmann, C. L.; Gladyshev, V. N.; Green, P.; Jorgensen, R.; Mayfield, S.; Mueller-Roeber, B.; Rajamani, S.; Sayre, R. T.; Brokstein, P.; Dubchak, I.; Goodstein, D.; Hornick, L.; Huang, Y. W.; Jhaveri, J.; Luo, Y.; Martínez, D.; Ngau, W. C. A.; Otillar, B.; Poliakov, A.; Porter, A.; Szajkowski, L.; Werner, G.; Zhou, K.; Grigoriev, I. V.; Rokhsar, D. S.; Grossman, A. R. The Chlamydomonas genome reveals the evolution of key animal and plant functions Science 2007, 318 (5848) 245-250
Esquível, M. G.; Amaro, H. M.; Pinto, T. S.; Fevereiro, P. S.; Malcata, F. X. Efficient H2 production via Chlamydomonas reinhardtii Trends Biotechnol. 2011, 29 (12) 595-600
Vijayaragh, K.; Karthik, R.; Kamala Nal, S. P. Hydrogen generation from algae: A review J. Plant Sci. 2010, 5 (1) 1-19
Wang, Z. T.; Ullrich, N.; Joo, S.; Waffenschmidt, S.; Goodenough, U. Algal lipid bodies: Stress induction, purification, and biochemical characterization in wild-type and starchless Chlamydomonas reinhardtii Eukaryotic Cell 2009, 8 (12) 1856-1868
Moellering, E. R.; Benning, C. RNA interference silencing of a major lipid droplet protein affects lipid droplet size in Chlamydomonas reinhardtii Eukaryotic Cell 2010, 9 (1) 97-106
Miller, R.; Wu, G.; Deshpande, R. R.; Vieler, A.; Gärtner, K.; Li, X.; Moellering, E. R.; Zäuner, S.; Cornish, A. J.; Liu, B.; Bullard, B.; Sears, B. B.; Kuo, M.-H.; Hegg, E. L.; Shachar-Hill, Y.; Shiu, S.-H.; Benning, C. Changes in transcript abundance in Chlamydomonas reinhardtii following nitrogen deprivation predict diversion of metabolism Plant Physiol. 2010, 154 (4) 1737-1752
Rogers, S.; Girolami, M.; Kolch, W.; Waters, K. M.; Liu, T.; Thrall, B.; Wiley, S. H. Investigating the correspondence between transcriptomic and proteomic expression profiles using coupled cluster models Bioinformatics 2008, 24 (24) 2894-2900
Jamers, A.; Blust, R.; De Coen, W. Omics in algae: Paving the way for a systems biological understanding of algal stress phenomena? Aquat. Toxicol. 2009, 92 (3) 114-121
Jinkerson, R. E.; Subramanian, V.; Posewitz, M. C. Improving biofuel production in phototrophic microorganisms with systems biology Biofuels 2011, 2 (2) 125-144
Stauber, E. J.; Hippler, M. Chlamydomonas reinhardtii proteomics Plant Physiol. Biochem. 2004, 42 (12) 989-1001
Rolland, N.; Atteia, A.; Decottignies, P.; Garin, J.; Hippler, M.; Kreimer, G.; Lemaire, S. D.; Mittag, M.; Wagner, V. Chlamydomonas proteomics Curr. Opin. Microbiol. 2009, 12 (3) 285-291
Keller, L. C.; Marshall, W. F. Isolation and proteomic analysis of Chlamydomonas centrioles Methods Mol. Biol. 2008, 432, 289-300
Atteia, A.; Adrait, A.; BrugieIre, S.; Tardif, M.; van Lis, R.; Deusch, O.; Dagan, T.; Kuhn, L.; Gontero, B.; Martin, W.; Garin, J.; Joyard, J.; Rolland, N. A proteomic survey of Chlamydomonas reinhardtii mitochondria sheds new light on the metabolic plasticity of the organelle and on the nature of the alpha-proteobacterial mitochondrial ancestor Mol. Biol. Evol. 2009, 26 (7) 1533-1548
Heide, H.; Nordhues, A.; Drepper, F.; Nick, S.; Schulz-Raffelt, M.; Haehnel, W.; Schroda, M. Application of quantitative immunoprecipitation combined with knockdown and cross-linking to Chlamydomonas reveals the presence of vesicle-inducing protein in plastids 1 in a common complex with chloroplast HSP90C Proteomics 2009, 9 (11) 3079-3089
Chen, M.; Zhao, L.; Sun, Y.-L.; Cui, S.-X.; Zhang, L.-F.; Yang, B.; Wang, J.; Kuang, T.-Y.; Huang, F. Proteomic analysis of hydrogen photoproduction in sulfur-deprived Chlamydomonas cells J. Proteome Res. 2010, 9 (8) 3854-3866
Cid, C.; Garcia-Descalzo, L.; Casado-Lafuente, V.; Amils, R.; Aguilera, A. Proteomic analysis of the response of an acidophilic strain of Chlamydomonas sp. (Chlorophyta) to natural metal-rich water Proteomics 2010, 10 (10) 2026-2036
Mathy, G.; Cardol, P.; Dinant, M.; Blomme, A.; Gérin, S.; Cloes, M.; Ghysels, B.; DePauw, E.; Leprince, P.; Remacle, C.; Sluse-Goffart, C.; Franck, F.; Matagne, R. F.; Sluse, F. E. Proteomic and functional characterization of a Chlamydomonas reinhardtii mutant lacking the mitochondrial alternative oxidase 1 J. Proteome Res. 2010, 9 (6) 2825-2838
Wagner, V.; Mittag, M. Probing circadian rhythms in Chlamydomonas rheinhardtii by functional proteomics Methods Mol. Biol. 2009, 479, 173-188
Gygi, S. P.; Corthals, G. L.; Zhang, Y.; Rochon, Y.; Aebersold, R. Evaluation of two-dimensional gel electrophoresis-based proteome analysis technology Proc. Natl. Acad. Sci. U.S.A. 2000, 97 (17) 9390-9395
Ong, S.-E.; Blagoev, B.; Kratchmarova, I.; Kristensen, D. B.; Steen, H.; Pandey, A.; Mann, M. Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics Mol. Cell. Proteomics 2002, 1 (5) 376-386
Ross, P. L.; Huang, Y. N.; Marchese, J. N.; Williamson, B.; Parker, K.; Hattan, S.; Khainovski, N.; Pillai, S.; Dey, S.; Daniels, S.; Purkayastha, S.; Juhasz, P.; Martin, S.; Bartlet-Jones, M.; He, F.; Jacobson, A.; Pappin, D. J. Multiplexed protein quantitation in Saccharomyces cerevisiae using amine-reactive isobaric tagging reagents Mol. Cell. Proteomics 2004, 3 (12) 1154-1169
Wu, W. W.; Wang, G.; Baek, S. J.; Shen, R.-F. Comparative study of three proteomic quantitative methods, DIGE, cICAT, and iTRAQ, using 2D Gel- or LC-MALDI TOF/TOF J. Proteome Res. 2006, 5 (3) 651-658
Mühlhaus, T.; Weiss, J.; Hemme, D.; Sommer, F.; Schroda, M. Quantitative shotgun proteomics using a uniform 15N-labeled standard to monitor proteome dynamics in time course experiments reveals new insights into the heat stress response of Chlamydomonas reinhardtii Mol. Cell. Proteomics 2011, 10, 9
Terashima, M.; Specht, M.; Naumann, B.; Hippler, M. Characterizing the anaerobic response of Chlamydomonas reinhardtii by quantitative proteomics Mol. Cell. Proteomics 2010, 9 (7) 1514-1532
Naumann, B.; Busch, A.; Allmer, J.; Ostendorf, E.; Zeller, M.; Kirchhoff, H.; Hippler, M. Comparative quantitative proteomics to investigate the remodeling of bioenergetic pathways under iron deficiency in Chlamydomonas reinhardtii Proteomics 2007, 7 (21) 3964-3979
Wienkoop, S.; Weiss, J.; May, P.; Kempa, S.; Irgang, S.; Recuenco-Munoz, L.; Pietzke, M.; Schwemmer, T.; Rupprecht, J.; Egelhofer, V.; Weckwerth, W. Targeted proteomics for Chlamydomonas reinhardtii combined with rapid subcellular protein fractionation, metabolomics and metabolic flux analyses Mol. BioSyst. 2010, 6 (6) 1018-1031
Wang, H.; Alvarez, S.; Hicks, L. M. Comprehensive comparison of iTRAQ and label-free LC-based quantitative proteomics approaches using two Chlamydomonas reinhardtii strains of interest for biofuels engineering J. Proteome Res. 2011, 11 (1) 487-501
Lee, D. Y.; Park, J.-J.; Barupal, D. K.; Fiehn, O. System response of metabolic networks in Chlamydomonas reinhardtii to total available ammonium Mol. Cell. Proteomics 2012, 11 (10) 973-988
Siaut, M.; Cuiné, S.; Cagnon, C.; Fessler, B.; Nguyen, M.; Carrier, P.; Beyly, A.; Beisson, F.; TriantaphylideIs, C.; Li-Beisson, Y.; Peltier, G. Oil accumulation in the model green alga Chlamydomonas reinhardtii: characterization, variability between common laboratory strains and relationship with starch reserves BMC Biotechnol. 2011, 11 (1) 7
Neupert, J.; Shao, N.; Lu, Y.; Bock, R. Genetic transformation of the model green alga C hlamydomonas reinhardtii Methods Mol. Biol. 2012, 847, 35-47
Scholz, M.; Hoshino, T.; Johnson, D.; Riley, M. R.; Cuello, J. Flocculation of wall-deficient cells of Chlamydomonas reinhardtii mutant cw15 by calcium and methanol Biomass Bioenergy 2011, 35 (12) 4835-4840
Ladygin, V. G.; Boutanaev, A. M. Transformation of Chlamydomonas reinhardtii CW-15 with the hygromycin phosphotransferase gene as a selectable marker Russ. J. Genet. 2002, 38 (9) 1009-1014
Bonente, G.; Pippa, S.; Castellano, S.; Bassi, R.; Ballottari, M. Acclimation of Chlamydomonas reinhardtii to different growth irradiances J. Biol. Chem. 2012, 287 (8) 5833-5847
Chankova, S. G.; Yurina, N. Micro-algae as a Model System for Studying of Genotype Resistance to Oxidative Stress and Adaptive Response. In Radiobiology and Environmental Security; Mothersill, C. E.; Korogodina, V.; Seymour, C. B., Eds.; NATO Science for Peace and Security Series C: Environmental Security; Springer: The Netherlands, 2012; pp 19-30.
Davies, D. R.; Plaskitt, A. Genetical and structural analyses of cell-wall formation in Chlamydomonas reinhardtii Genet. Res. 1971, 17 (01) 33-43
Harris, E. H. Chlamydomonas as a model organism Annu. Rev. Plant Physiol. Plant Mol. Biol. 2001, 52, 363-406
Harris, E. H.; Stern, D. B.; Witman, G. The Chlamydomonas Sourcebook: Introduction to Chlamydomonas and Its Laboratory Use; Academic Press: San Diego, CA, 2009.
Wellburn, R. W. The spectral determination of chlorophylls a and b, as well as total carotenoids, using various solvents with spectrophotometers of different resolution J. Plant Physiol. 1994, 144 (3) 307-313
Gerhardt, P.; Murray, R. G. E.; Wood, W. A.; Krieg, N. R. Methods for General and Molecular Bacteriology; American Society for Microbiology: Washington, DC, 1994; Vol. 40.
Chen, W.; Zhang, C.; Song, L.; Sommerfeld, M.; Hu, Q. A high throughput Nile red method for quantitative measurement of neutral lipids in microalgae J. Microbiol. Methods 2009, 77 (1) 41-47
Ow, S. Y.; Salim, M.; Noirel, J.; Evans, C.; Rehman, I.; Wright, P. C. iTRAQ underestimation in simple and complex mixtures: "The good, the bad and the ugly" J. Proteome Res. 2009, 8 (11) 5347-5355
R Development Core Team. R: A Language and Environment for Statistical Computing; R Foundation for Statistical Computing: Vienna, Austria, 2011.
Muller, J.; Szklarczyk, D.; Julien, P.; Letunic, I.; Roth, A.; Kuhn, M.; Powell, S.; von Mering, C.; Doerks, T.; Jensen, L. J.; Bork, P. eggNOG v2.0: Extending the evolutionary genealogy of genes with enhanced non-supervised orthologous groups, species and functional annotations Nucleic Acids Res. 2010, 38, D190-D195
Tatusov, R. L.; Koonin, E. V.; Lipman, D. J. A genomic perspective on protein families Science 1997, 278 (5338) 631-637
Ow, S. Y.; Salim, M.; Noirel, J.; Evans, C.; Wright, P. C. Minimising iTRAQ ratio compression through understanding LC-MS elution dependence and high-resolution HILIC fractionation Proteomics 2011, 11 (11) 2341-2346
Wang, Y.; Spalding, M. H. An inorganic carbon transport system responsible for acclimation specific to air levels of CO2 in Chlamydomonas Reinhardtii Proc. Natl. Acad. Sci. U.S.A. 2006, 103 (26) 10110-10115
Miura, K.; Yamano, T.; Yoshioka, S.; Kohinata, T.; Inoue, Y.; Taniguchi, F.; Asamizu, E.; Nakamura, Y.; Tabata, S.; Yamato, K. T.; Ohyama, K.; Fukuzawa, H. Expression profiling-based identification of CO2-responsive genes regulated by CCM1 controlling a carbon-concentrating mechanism in Chlamydomonas reinhardtii Plant Physiol. 2004, 135 (3) 1595-1607
Abe, J.; Kubo, T.; Takagi, Y.; Saito, T.; Miura, K.; Fukuzawa, H.; Matsuda, Y. The transcriptional program of synchronous gametogenesis in Chlamydomonas reinhardtii Curr. Genet. 2004, 46 (5) 304-315
Kanehisa, M.; Goto, S. KEGG: kyoto encyclopedia of genes and genomes Nucleic Acids Res. 2000, 28 (1) 27-30
Kanehisa, M.; Goto, S.; Sato, Y.; Furumichi, M.; Tanabe, M. KEGG for integration and interpretation of large-scale molecular data sets Nucleic Acids Res. 2012, 40 (D1) D109-D114
