Abstract
Jatropha curcas L. is an excellent biofuel crop, which displays a high efficiency of carbon absorption, and seed oil of Jatropha can be efficiently processed to produce high-quality biodiesel. Plant phosphoenolpyruvate carboxylases (PEPCs) play important roles not only in initial fixation of atmospheric CO2 in C4 and Crassulacean acid metabolism (CAM) plants, but also in fatty acid biosynthesis in seeds of oil plants by regulating carbon partitioning. Here, we identified JcPEPC1 from J. curcas L. by homology cloning, and alignment analysis of protein sequence revealed JcPEPC1 was a plant C3-type PEPC, and shared high similarity to PEPC of castor oil plant Ricinus communis. We implemented detailed functional characterization of JcPEPC1 by expression analysis and transgenic tobacco. JcPEPC1 gene expressed in the leaves and seeds of J. curcas L., and remarkable increase of expression level was also detected at seed oil-accumulating stages. We overexpressed JcPEPC1 in tobacco, and showed the enzymatic activity of PEPC in transgenic plants was notably higher than wild type. Gas chromatography (GC) analysis elucidated the composition and total content of fatty acids were also altered. This study indicated JcPEPC1 played a fundamental role in fatty acid biosynthesis in Jatropha seeds. Our results proposed enhanced PEPC activity of Jatropha could improve biosynthesis of fatty acid, which implied critical functions in primary metabolism of non-photosynthetic PEPC.
Similar content being viewed by others
References
Abdulla R, Chan ES, Ravindra P (2011) Biodiesel production from Jatropha curcas: a critical review. Crit Rev Biotechnol 31(1):53–64. doi:10.3109/07388551.2010.487185
Alonso AP, Goffman FD, Ohlrogge JB, Shachar-Hill Y (2007) Carbon conversion efficiency and central metabolic fluxes in developing sunflower (Helianthus annuus L.) embryos. Plant J 52(2):296–308. doi:10.1111/j.1365-313X.2007.03235.x
Alonso AP, Dale VL, Shachar-Hill Y (2010) Understanding fatty acid synthesis in developing maize embryos using metabolic flux analysis. Metab Eng 12(5):488–497. doi:10.1016/j.ymben.2010.04.002
Alonso AP, Val DL, Shachar-Hill Y (2011) Central metabolic fluxes in the endosperm of developing maize seeds and their implications for metabolic engineering. Metab Eng 13(1):96–107. doi:10.1016/j.ymben.2010.10.002
Azocar L, Ciudad G, Heipieper HJ, Navia R (2010) Biotechnological processes for biodiesel production using alternative oils. Appl Microbiol Biotechnol 88(3):621–636. doi:10.1007/s00253-010-2804-z
Browse J, McCourt PJ, Somerville CR (1986) Fatty acid composition of leaf lipids determined after combined digestion and fatty acid methyl ester formation from fresh tissue. Anal Biochem 152(1):141–145. doi:10.1016/0003-2697(86)90132-6
Chen LM, Li KZ, Miwa T, Izui K (2004) Overexpression of a cyanobacterial phosphoenolpyruvate carboxylase with diminished sensitivity to feedback inhibition in Arabidopsis changes amino acid metabolism. Planta 219(3):440–449. doi:10.1007/s00425-004-1244-3
Costa GGL, Cardoso KC, Del Bem LEV, Lima AC, Cunha MAS, de Campos-Leite L, Vicentini R, Papes F, Moreira RC, Yunes JA, Campos FAP, Da Silva MJ (2010) Transcriptome analysis of the oil-rich seed of the bioenergy crop Jatropha curcas L. BMC Genomics 11(1):462. doi:10.1186/1471-2164-11-462
Debnath M, Bisen PS (2008) Jatropha curcas L., a multipurpose stress resistant plant with a potential for Ethnomedicine and renewable energy. Curr Pharm Biotechnol 9(4):288–306. doi:10.2174/138920108785161541
Dodds ED, McCoy MR, Rea LD, Kennish JM (2005) Gas chromatographic quantification of fatty acid methyl esters: flame ionization detection vs electron impact mass spectrometry. Lipids 40(4):419–428
Eastmond PJ, Dennis DT, Rawsthorne S (1997) Evidence that a malate inorganic phosphate exchange translocator imports carbon across the leucoplast envelope for fatty acid synthesis in developing castor seed endosperm. Plant Physiol 114(3):851–856
Endo T, Mihara Y, Furumoto T, Matsumura H, Kai Y, Izui K (2008) Maize C(4)-form phosphoenolpyruvate carboxylase engineered to be functional in C(3) plants: mutations for diminished sensitivity to feedback inhibitors and for increased substrate affinity. J Exp Bot 59(7):1811–1818. doi:10.1093/Jxb/Ern018
Gennidakis S, Rao S, Greenham K, Uhrig RG, O’Leary B, Snedden WA, Lu C, Plaxton WC (2007) Bacterial- and plant-type phosphoenolpyruvate carboxylase polypeptides interact in the hetero-oligomeric Class-2 PEPC complex of developing castor oil seeds. Plant J 52(5):839–849. doi:10.1111/j.1365-313X.2007.03274.x
Goffman FD, Ruckle M, Ohlrogge J, Shachar-Hill Y (2004) Carbon dioxide concentrations are very high in developing oilseeds. Plant Physiol Biochem 42(9):703–708. doi:10.1016/j.plaphy.2004.07.003
Gu KY, Chiam H, Tian DS, Yin ZC (2011) Molecular cloning and expression of heteromeric ACCase subunit genes from Jatropha curcas. Plant Sci 180(4):642–649. doi:10.1016/j.plantsci.2011.01.007
Hudspeth RL, Grula JW, Dai Z, Edwards GE, Ku MSB (1992) Expression of maize phosphoenolpyruvate carboxylase in transgenic tobacco—effects on biochemistry and physiology. Plant Physiol 98(2):458–464. doi:10.1104/Pp.98.2.458
Izui K, Matsumura H, Furumoto T, Kai Y (2004) Phosphoenolpyruvate carboxylase: a new era of structural biology. Annu Rev Plant Biol 55:69–84. doi:10.1146/annurev.arplant.55.031903.141619
Jeanneau M, Vidal J, Gousset-Dupont A, Lebouteiller B, Hodges M, Gerentes D, Perez P (2002) Manipulating PEPC levels in plants. J Exp Bot 53(376):1837–1845. doi:10.1093/Jxb/Erf061
Johnson TS, Eswaran N, Sujatha M (2011) Molecular approaches to improvement of Jatropha curcas Linn. as a sustainable energy crop. Plant Cell Rep 30(9):1573–1591. doi:10.1007/s00299-011-1083-1
Kai Y, Matsumura H, Inoue T, Terada K, Nagara Y, Yoshinaga T, Kihara A, Tsumura K, Izui K (1999) Three-dimensional structure of phosphoenolpyruvate carboxylase: a proposed mechanism for allosteric inhibition. Proc Natl Acad Sci USA 96(3):823–828. doi:10.1073/pnas.96.3.823
Kant P, Wu SR (2011) The extraordinary collapse of Jatropha as a global biofuel. Environ Sci Technol 45(17):7114–7115. doi:10.1021/Es201943y
Liu H, Yang ZL, Yang MF, Shen SH (2011) The differential proteome of endosperm and embryo from mature seed of Jatropha curcas. Plant Sci 181(6):660–666. doi:10.1016/j.plantsci.2011.03.012
Meimoun P, Gousset-Dupont A, Lebouteiller B, Ambard-Bretteville F, Besin E, Lelarge C, Mauve C, Hodges M, Vidal J (2009) The impact of PEPC phosphorylation on growth and development of Arabidopsis thaliana: molecular and physiological characterization of PEPC kinase mutants. FEBS Lett 583(10):1649–1652. doi:10.1016/j.febslet.2009.04.030
Murmu J, Plaxton WC (2007) Phosphoenolpyruvate carboxylase protein kinase from developing castor oil seeds: partial purification, characterization, and reversible control by photosynthate supply. Planta 226(5):1299–1310. doi:10.1007/s00425-007-0551-x
Natarajan P, Parani M (2011) De novo assembly and transcriptome analysis of five major tissues of Jatropha curcas L. using GS FLX titanium platform of 454 pyrosequencing. BMC Genomics 12(1):191. doi:10.1186/1471-2164-12-191
Ohlrogge JB, Jaworski JG (1997) Regulation of fatty acid synthesis. Annu Rev Plant Phys 48:109–136. doi:10.1146/annurev.arplant.48.1.109
Ohlrogge J, Pollard M, Bao X, Focke M, Girke T, Ruuska S, Mekhedov S, Benning C (2000) Fatty acid synthesis: from CO2 to functional genomics. Biochem Soc T 28:567–574. doi:10.1042/0300-5127:0280567
O’Leary B, Park J, Plaxton WC (2011) The remarkable diversity of plant PEPC (phosphoenolpyruvate carboxylase): recent insights into the physiological functions and post-translational controls of non-photosynthetic PEPCs. Biochem J 436:15–34. doi:10.1042/Bj20110078
Park J, Khuu N, Howard ASM, Mullen RT, Plaxton WC (2012) Bacterial- and plant-type phosphoenolpyruvate carboxylase isozymes from developing castor oil seeds interact in vivo and associate with the surface of mitochondria. Plant J 71(2):251–262. doi:10.1111/j.1365-313X.2012.04985.x
Rademacher T, Hausler RE, Hirsch HJ, Zhang L, Lipka V, Weier D, Kreuzaler F, Peterhansel C (2002) An engineered phosphoenolpyruvate carboxylase redirects carbon and nitrogen flow in transgenic potato plants. Plant J 32(1):25–39. doi:10.1046/j.1365-313X.2002.01397.x
Ramkat RC, Calari A, Maghuly F, Laimer M (2011) Biotechnological approaches to determine the impact of viruses in the energy crop plant Jatropha curcas. Virol J 3(8):386. doi:10.1186/1743-422x-8-386
Sanchez R, Cejudo FJ (2003) Identification and expression analysis of a gene encoding a bacterial-type phosphoenolpyruvate carboxylase from Arabidopsis and rice. Plant Physiol 132(2):949–957. doi:10.1104/pp.102.019653
Sangwan RS, Singh N, Plaxton WC (1992) Phosphoenolpyruvate carboxylase activity and concentration in the endosperm of developing and germinating castor-oil seeds. Plant Physiol 99(2):445–449. doi:10.1104/Pp.99.2.445
Sato S, Hirakawa H, Isobe S, Fukai E, Watanabe A, Kato M, Kawashima K, Minami C, Muraki A, Nakazaki N, Takahashi C, Nakayama S, Kishida Y, Kohara M, Yamada M, Tsuruoka H, Sasamoto S, Tabata S, Aizu T, Toyoda A, Shin-i T, Minakuchi Y, Kohara Y, Fujiyama A, Tsuchimoto S, Kajiyama S, Makigano E, Ohmido N, Shibagaki N, Cartagena JA, Wada N, Kohinata T, Atefeh A, Yuasa S, Matsunaga S, Fukui K (2011) Sequence analysis of the genome of an oil-bearing tree Jatropha curcas L. DNA Res 18(1):65–76. doi:10.1093/dnares/dsq030
Sayre RT, Kennedy RA (1979) Photosynthetic enzyme activities and localization in Mollugoverticillata populations differing in the leaves of C3 and C4 cycle operation. Plant Physiol 64(2):293–299. doi:10.1104/pp.64.2.293
Sebei K, Ouerghi Z, Kallel H, Boukhchina S (2006) Evolution of phosphoenolpyruvate carboxylase activity and lipid content during seed maturation of two spring rapeseed cultivars (Brassica napus L.). CR Biol 329(9):719–725. doi:10.1016/j.crvi.2006.06.002
Smith RG, Gauthier DA, Dennis DT, Turpin DH (1992) Malate- and pyruvate-dependent fatty acid synthesis in leucoplasts from developing castor endosperm. Plant Physiol 98(4):1233–1238. doi:10.1104/pp.98.4.1233
Sullivan S, Jenkins GI, Nimmo HG (2004) Roots, cycles and leaves. Expression of the phosphoenolpyruvate carboxylase kinase gene family in soybean. Plant Physiol 135(4):2078–2087. doi:10.1104/pp.104.042762
Suzuki S, Murai N, Burnell JN, Arai M (2000) Changes in photosynthetic carbon flow in transgenic rice plants that express C4-type phosphoenolpyruvate carboxykinase from Urochloa panicoides. Plant Physiol 124(1):163–172. doi:10.1104/Pp.124.1.163
Tang MJ, Liu XF, Deng HP, Shen SH (2011) Over-expression of JcDREB, a putative AP2/EREBP domain-containing transcription factor gene in woody biodiesel plant Jatropha curcas, enhances salt and freezing tolerance in transgenic Arabidopsis thaliana. Plant Sci 181(6):623–631. doi:10.1016/j.plantsci.2011.06.014
Troncoso-Ponce MA, Kilaru A, Cao X, Durrett TP, Fan JL, Jensen JK, Thrower NA, Pauly M, Wilkerson C, Ohlrogge JB (2011) Comparative deep transcriptional profiling of four developing oilseeds. Plant J 68(6):1014–1027. doi:10.1111/j.1365-313X.2011.04751.x
Wang QY, Guan YC, Wu YR, Chen HL, Chen F, Chu CC (2008) Overexpression of a rice OsDREB1F gene increases salt, drought, and low temperature tolerance in both Arabidopsis and rice. Plant Mol Biol 67:589–602
Xu WX, Sato SJ, Clemente TE, Chollet R (2007) The PEP-carboxylase kinase gene family in Glycine max (GmPpcK1-4): an in-depth molecular analysis with nodulated, non-transgenic and transgenic plants. Plant J 49(5):910–923. doi:10.1111/j.1365-313X.2006.03006.x
Acknowledgments
This work was supported by the funds from Key Projects in the National Science and Technology Pillar Program during the Twelfth Five-year Plan Period (No.2012BAD01B0703). We also acknowledge the Nonprofit Research Projects (RISF6153, RISF6141) and Country’s international cooperation projects (2011DFA30490). We thank Dr. Renying Zhuo (Chinese Academy of Forestry) for providing seeds of wild type tobacco. We are grateful to Dr. Peng Gao (National Research Council Canada) for the help of fatty acid analysis. We greatly appreciate two anonymous reviewers and the editor for critical reading and helpful comments.
Author information
Authors and Affiliations
Corresponding author
Additional information
Communicated by U. Feller.
Rights and permissions
About this article
Cite this article
Fan, Z., Li, J., Lu, M. et al. Overexpression of phosphoenolpyruvate carboxylase from Jatropha curcas increases fatty acid accumulation in Nicotiana tabacum . Acta Physiol Plant 35, 2269–2279 (2013). https://doi.org/10.1007/s11738-013-1264-3
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11738-013-1264-3