Abstract
Main conclusion
Portulaca leaves serve as an alternative bioresource for edible PUFAs. Transcriptome data provide information to explore Portulaca as a model system for galactolipids, leaf lipid metabolism, and PUFA-rich designer lipids.
Poly-unsaturated fatty acids (PUFAs) are gaining importance due to their innumerable health benefits, and hence, understanding their biosynthesis in plants has attained prominence in recent years. The most common source of PUFAs is of marine origin. Although reports have identified Portulaca oleracea (purslane) as a leaf source of omega-3 fatty acids in the form of alpha-linolenic acid (ALA), the mechanism of ALA accumulation and its distribution into various lipids has not been elucidated. Here, we present the lipid profiles of leaves and seeds of several accessions of P. oleracea. Among the nineteen distinct accessions, the RR04 accession has the highest amount of ALA and is primarily associated with galactolipids. In addition, we report the transcriptome of RR04, and we have mapped the potential genes involved in lipid metabolism. Phosphatidylcholine (PC) is the major site of acyl editing, which is catalyzed by lysophosphatidylcholine acyltransferase (LPCAT), an integral membrane protein that plays a major role in supplying oleate to the PC pool for further unsaturation. Our investigations using mass spectrometric analysis of leaf microsomal fractions identified LPCAT as part of a membrane protein complex. Both native and recombinant LPCAT showed strong acyltransferase activity with various acyl-CoA substrates. Altogether, the results suggest that ALA-rich glycerolipid biosynthetic machinery is highly active in nutritionally important Portulaca leaves. Furthermore, lipidome, transcriptome, and mass spectrometric analyses of RR04 provide novel information for exploring Portulaca as a potential resource and a model system for studying leaf lipid metabolism.
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References
AnastáCio A, Carvalho IS (2012) Accumulation of fatty acids in purslane grown in hydroponic salt stress conditions. Int J Food Sci Nutr 64(2):235–242. https://doi.org/10.3109/09637486.2012.713915
Arroyo-Caro JM, Chileh T, Alonso DL, García-Maroto F (2013) molecular characterization of a lysophosphatidylcholine acyltransferase gene belonging to the MBOAT family in Ricinus communis L. Lipids 48(7):663–674. https://doi.org/10.1007/s11745-013-3797-z
Ayerza R, Coates W (2011) Protein content, oil content and fatty acid profiles as potential criteria to determine the origin of commercially grown chia (Salvia hispanica L.). Ind Crops Prod 34(2):1366–1371. https://doi.org/10.1016/j.indcrop.2010.12.007
Bankar KG, Todur VN, Shukla RN, Vasudevan M (2015) Ameliorated de novo transcriptome assembly using Illumina paired end sequence data with Trinity Assembler. Genom Data 5:352–359. https://doi.org/10.1016/j.gdata.2015.07.012
Bates PD, Browse J (2012) The significance of different diacylgycerol synthesis pathways on plant oil composition and bioengineering. Front Plant Sci. https://doi.org/10.3389/fpls.2012.00147
Bates PD, Ohlrogge JB, Pollard M (2007) Incorporation of newly synthesized fatty acids into cytosolic glycerolipids in pea leaves occurs via acyl editing. J Biol Chem 282(43):31206–31216. https://doi.org/10.1074/jbc.m705447200
Bates PD, Fatihi A, Snapp AR, Carlsson AS, Browse J, Lu C (2012) Acyl editing and headgroup exchange are the major mechanisms that direct polyunsaturated fatty acid flux into triacylglycerols. Plant Physiol 160(3):1530–1539. https://doi.org/10.1104/pp.112.204438
Bates PD, Stymne S, Ohlrogge J (2013) Biochemical pathways in seed oil synthesis. Curr Opin Plant Biol 16(3):358–364. https://doi.org/10.1016/j.pbi.2013.02.015
Browse J, Warwick N, Somerville CR, Slack CR (1986) Fluxes through the prokaryotic and eukaryotic pathways of lipid synthesis in the ‘16:3’ plant Arabidopsis thaliana. Biochem J 235(1):25–31. https://doi.org/10.1042/bj2350025
Burdge GC, Calder PC (2005) α-Linolenic acid metabolism in adult humans: the effects of gender and age on conversion to longer-chain polyunsaturated fatty acids. Eur J Lipid Sci Technol 107(6):426–439. https://doi.org/10.1002/ejlt.200501145
Chapman KD, Ohlrogge JB (2012) Compartmentation of triacylglycerol accumulation in plants. J Biol Chem 287(4):2288–2294. https://doi.org/10.1074/jbc.R111.290072
Chapman KD, Dyer JM, Mullen RT (2013) Commentary: why don’t plant leaves get fat? Plant Sci 207:128–134. https://doi.org/10.1016/j.plantsci.2013.03.003
Cheng J, Zhu L-H, Salentijn EMJ, Huang B, Gruber J, Dechesne AC, Krens FA, Qi W, Visser RGF, van Loo EN (2013) Functional analysis of the omega-6 fatty acid desaturase (CaFAD2) gene family of the oil seed crop Crambe abyssinica. BMC Plant Biol 13(1):146. https://doi.org/10.1186/1471-2229-13-146
Cumberford G, Hebard A (2015) Ahiflower oil: a novel non-GM plant-based omega-3 + 6 source. Lipid Technol 27(9):207–210. https://doi.org/10.1002/lite.201500044
Dahlqvist A, Stahl U, Lenman M, Banas A, Lee M, Sandager L, Ronne H, Stymne S (2000) Phospholipid:diacylglycerol acyltransferase: an enzyme that catalyzes the acyl-CoA-independent formation of triacylglycerol in yeast and plants. Proc Natl Acad Sci 97(12):6487–6492. https://doi.org/10.1073/pnas.120067297
Fan J, Yan C, Zhang X, Xu C (2013) Dual role for phospholipid: diacylglycerol acyltransferase: enhancing fatty acid synthesis and diverting fatty acids from membrane lipids to triacylglycerol in Arabidopsis Leaves. Plant Cell 25(9):3506–3518. https://doi.org/10.1105/tpc.113.117358
Fan J, Yan C, Roston R, Shanklin J, Xu C (2014) Arabidopsis lipins, PDAT1 acyltransferase, and SDP1 triacylglycerol lipase synergistically direct fatty acids toward β-oxidation, thereby maintaining membrane lipid homeostasis. Plant Cell 26(10):4119–4134. https://doi.org/10.1105/tpc.114.130377
Gigon A, Matos A-R, Laffray D, Zuily-Fodil Y, Pham-Thi A-T (2004) Effect of drought stress on lipid metabolism in the leaves of Arabidopsis thaliana (Ecotype Columbia). Ann Bot 94(3):345–351. https://doi.org/10.1093/aob/mch150
Haas BJ, Papanicolaou A, Yassour M, Grabherr M, Blood PD, Bowden J, Couger MB, Eccles D, Li B, Lieber M, MacManes MD, Ott M, Orvis J, Pochet N, Strozzi F, Weeks N, Westerman R, William T, Dewey CN, Henschel R, LeDuc RD, Friedman N, Regev A (2013) De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis. Nat Protoc 8(8):1494–1512. https://doi.org/10.1038/nprot.2013.084
Hill J, Nelson E, Tilman D, Polasky S, Tiffany D (2006) Environmental, economic, and energetic costs and benefits of biodiesel and ethanol biofuels. Proc Natl Acad Sci 103(30):11206–11210. https://doi.org/10.1073/pnas.0604600103
Hoffmann M, Wagner M, Abbadi A, Fulda M, Feussner I (2008) Metabolic engineering of ω3-very long chain polyunsaturated fatty acid production by an exclusively acyl-CoA-dependent pathway. J Biol Chem 283(33):22352–22362. https://doi.org/10.1074/jbc.m802377200
Joyard J, Ferro M, Masselon C, Seigneurin-Berny D, Salvi D, Garin J, Rolland N (2010) Chloroplast proteomics highlights the subcellular compartmentation of lipid metabolism. Prog Lipid Res 49(2):128–158. https://doi.org/10.1016/j.plipres.2009.10.003
Kim YH, Choi J-S, Yoo JS, Park Y-M, Kim MS (1999) Structural identification of glycerolipid molecular species isolated from Cyanobacterium Synechocystis sp. PCC 6803 using fast atom bombardment tandem mass spectrometry. Anal Biochem 267(2):260–270. https://doi.org/10.1006/abio.1998.3041
Kris-Etherton PM, Taylor DS, Yu-Poth S, Huth P, Moriarty K, Fishell V, Hargrove RL, Zhao G, Etherton TD (2000) Polyunsaturated fatty acids in the food chain in the United States. Am J Clin Nutr 71(1 Suppl):179S–188S
Lager I, Yilmaz JL, Zhou X-R, Jasieniecka K, Kazachkov M, Wang P, Zou J, Weselake R, Smith MA, Bayon S, Dyer JM, Shockey JM, Heinz E, Green A, Banas A, Stymne S (2013) Plant acyl-CoA: lysophosphatidylcholine acyltransferases (LPCATs) have different specificities in their forward and reverse reactions. J Biol Chem 288(52):36902–36914. https://doi.org/10.1074/jbc.m113.521815
Langmead B, Salzberg SL (2012) Fast gapped-read alignment with Bowtie 2. Nat Methods 9(4):357–359. https://doi.org/10.1038/nmeth.1923
Li B, Dewey CN (2011) RSEM: accurate transcript quantification from RNA-Seq data with or without a reference genome. BMC Bioinform 12(1):323. https://doi.org/10.1186/1471-2105-12-323
Li-Beisson Y, Shorrosh B, Beisson F, Andersson MX, Arondel V, Bates PD, Baud S, Bird D, DeBono A, Durrett TP, Franke RB, Graham IA, Katayama K, Kelly AA, Larson T, Markham JE, Miquel M, Molina I, Nishida I, Rowland O, Samuels L, Schmid KM, Wada H, Welti R, Xu C, Zallot R, Ohlrogge J (2010) Acyl-lipid metabolism. Arabidopsis Book 8:e0133. https://doi.org/10.1199/tab.0133
Lin W, Oliver DJ (2008) Role of triacylglycerols in leaves. Plant Sci 175(3):233–237. https://doi.org/10.1016/j.plantsci.2008.04.003
Lung S-C, Weselake RJ (2006) Diacylglycerol acyltransferase: a key mediator of plant triacylglycerol synthesis. Lipids 41(12):1073–1088. https://doi.org/10.1007/s11745-006-5057-y
Melo N, Tavares RM, Morais F, Barroso JG, Pais MSS (1995) Lipid composition of thylakoid membranes from leaves and regreened spathes of Zantedeschia aethiopica. Phytochemistry 40(5):1367–1371. https://doi.org/10.1016/0031-9422(95)00506-3
Misurcova L, Ambrozova J, Samek D (2011) Seaweed lipids as nutraceuticals. Adv Food Nutr Res 64:339–355. https://doi.org/10.1016/B978-0-12-387669-0.00027-2
Murphy J, Bustin SA (2009) Reliability of real-time reverse-transcription PCR in clinical diagnostics: gold standard or substandard? Expert Rev Mol Diagn 9(2):187–197. https://doi.org/10.1586/14737159.9.2.187
Nichols PD, Petrie J, Singh S (2010) Long-chain omega-3 oils–an update on sustainable sources. Nutrients 2(12):572–585. https://doi.org/10.3390/nu2060572
Ohlrogge J, Browse J (1995) Lipid biosynthesis. Plant Cell 7(7):957. https://doi.org/10.2307/3870050
Omara-Alwala TR, Mebrahtu T, Prior DE, Ezekwe MO (1991) Omega-three fatty acids in Purslane (Portulaca oleracea) tissues. J Am Oil Chem Soc 68(3):198–199. https://doi.org/10.1007/bf02657769
Palaniswamy UR, McAvoy RJ, Bible BB (2001) Stage of harvest and polyunsaturated essential fatty acid concentrations in Purslane (Portulaca oleraceae) leaves. J Agric Food Chem 49(7):3490–3493. https://doi.org/10.1021/jf0102113
Pan X, Siloto RMP, Wickramarathna AD, Mietkiewska E, Weselake RJ (2013) Identification of a pair of phospholipid: diacylglycerol acyltransferases from developing flax (Linum usitatissimum L.) seed catalyzing the selective production of trilinolenin. J Biol Chem 288(33):24173–24188. https://doi.org/10.1074/jbc.m113.475699
Pan X, Chen G, Kazachkov M, Greer MS, Caldo KMP, Zou J, Weselake RJ (2015) In vivo and in vitro evidence for biochemical coupling of reactions catalyzed by lysophosphatidylcholine acyltransferase and diacylglycerol acyltransferase. J Biol Chem 290(29):18068–18078. https://doi.org/10.1074/jbc.m115.654798
Parthibane V, Rajakumari S, Venkateshwari V, Iyappan R, Rajasekharan R (2011) Oleosin is bifunctional enzyme that has both monoacylglycerol acyltransferase and phospholipase activities. J Biol Chem 287(3):1946–1954. https://doi.org/10.1074/jbc.m111.309955
Petrie JR, Shrestha P, Zhou X-R, Mansour MP, Liu Q, Belide S, Nichols PD, Singh SP (2012) Metabolic engineering plant seeds with fish oil-like levels of DHA. PLoS One 7(11):e49165. https://doi.org/10.1371/journal.pone.0049165
Petrie JR, Shrestha P, Belide S, Kennedy Y, Lester G, Liu Q, Divi UK, Mulder RJ, Mansour MP, Nichols PD, Singh SP (2014) Metabolic engineering camelina sativa with fish oil-like levels of DHA. PLoS One 9(1):e85061. https://doi.org/10.1371/journal.pone.0085061
Proctor CA, Gaussoin RE, Reicher ZJ (2011) Vegetative reproduction potential of common Purslane (Portulaca oleracea). Weed Technol 25(04):694–697. https://doi.org/10.1614/wt-d-11-00045.1
Qi B, Fraser T, Mugford S, Dobson G, Sayanova O, Butler J, Napier JA, Stobart AK, Lazarus CM (2004) Production of very long chain polyunsaturated omega-3 and omega-6 fatty acids in plants. Nat Biotechnol 22(6):739–745. https://doi.org/10.1038/nbt972
Rajasekharan R, Nachiappan V (1994) Use of photoreactive substrates for characterization of lysophosphatidate acyltransferases from developing soybean cotyledons. Arch Biochem Biophys 311(2):389–394. https://doi.org/10.1006/abbi.1994.1253
Robert SS, Singh SP, Zhou X-R, Petrie JR, Blackburn SI, Mansour PM, Nichols PD, Liu Q, Green AG (2005) Metabolic engineering of Arabidopsis to produce nutritionally important DHA in seed oil. Funct Plant Biol 32(6):473. https://doi.org/10.1071/fp05084
Ruiz-Lopez N, Haslam RP, Usher SL, Napier JA, Sayanova O (2013) Reconstitution of EPA and DHA biosynthesis in Arabidopsis: iterative metabolic engineering for the synthesis of n − 3 LC-PUFAs in transgenic plants. Metab Eng 17:30–41. https://doi.org/10.1016/j.ymben.2013.03.001
Saha S, Enugutti B, Rajakumari S, Rajasekharan R (2006) Cytosolic triacylglycerol biosynthetic pathway in oilseeds. Molecular cloning and expression of peanut cytosolic diacylglycerol acyltransferase. Plant Physiol 141(4):1533–1543. https://doi.org/10.1104/pp.106.082198
Sayanova O, Haslam RP, Calerón MV, López NR, Worthy C, Rooks P, Allen MJ, Napier JA (2011) Identification and functional characterisation of genes encoding the omega-3 polyunsaturated fatty acid biosynthetic pathway from the coccolithophore Emiliania huxleyi. Phytochemistry 72(7):594–600. https://doi.org/10.1016/j.phytochem.2011.01.022
Schaller GE (2017) Isolation of endoplasmic reticulum and its membrane. Methods Mol Biol 1511:119–129. https://doi.org/10.1007/978-1-4939-6533-5_10
Siloto RMP, Truksa M, He X, McKeon T, Weselake RJ (2009) Simple methods to detect triacylglycerol biosynthesis in a yeast-based recombinant system. Lipids 44(10):963–973. https://doi.org/10.1007/s11745-009-3336-0
Simopoulos AP, Norman HA, Gillaspy JE (1995) Purslane in human nutrition and its potential for world agriculture. World Rev Nutr Diet 77:47–74
Simpson JP, Ohlrogge JB (2016) A novel pathway for triacylglycerol biosynthesis is responsible for the accumulation of massive quantities of glycerolipids in the surface wax of bayberry (Myrica pensylvanica) fruit. Plant Cell 28(1):248–264. https://doi.org/10.1105/tpc.15.00900
Simopoulos AP, Norman HA, Gillaspy JE, Duke JA (1992) Common purslane: a source of omega-3 fatty acids and antioxidants. J Am Coll Nutr 11(4):374–382
Singer SD, Chen G, Mietkiewska E, Tomasi P, Jayawardhane K, Dyer JM, Weselake RJ (2016) Arabidopsis GPAT9 contributes to synthesis of intracellular glycerolipids but not surface lipids. J Exp Bot 67(15):4627–4638. https://doi.org/10.1093/jxb/erw242
Siriwardhana N, Kalupahana NS, Moustaid-Moussa N (2012) Health benefits of n − 3 polyunsaturated fatty acids. Marine medicinal foods—implications and applications—animals and microbes. Elsevier, New York. https://doi.org/10.1016/b978-0-12-416003-3.00013-5
Slocombe SP, Cornah J, Pinfield-Wells H, Soady K, Zhang Q, Gilday A, Dyer JM, Graham IA (2009) Oil accumulation in leaves directed by modification of fatty acid breakdown and lipid synthesis pathways. Plant Biotechnol J 7(7):694–703. https://doi.org/10.1111/j.1467-7652.2009.00435.x
Strijbosch RAM, Lee S, Arsenault DA, Andersson C, Gura KM, Bistrian BR, Puder M (2008) Fish oil prevents essential fatty acid deficiency and enhances growth: clinical and biochemical implications. Metabolism 57(5):698–707. https://doi.org/10.1016/j.metabol.2008.01.008
Stymne S, Stobart AK (1987) Triacylglycerol biosynthesis. Lipids structure and function. Elsevier, New York. https://doi.org/10.1016/b978-0-12-675409-4.50014-9
Taylor NL, Stroher E, Millar AH (2014) Arabidopsis organelle isolation and characterization. Methods Mol Biol 1062:551–572. https://doi.org/10.1007/978-1-62703-580-4_29
Tjellström H, Strawsine M, Ohlrogge JB (2015) Tracking synthesis and turnover of triacylglycerol in leaves. J Exp Bot 66(5):1453–1461. https://doi.org/10.1093/jxb/eru500
Tumaney AW, Rajasekharan R (1999) Synthesis of azidophospholipids and labeling of lysophosphatidylcholine acyltransferase from developing soybean cotyledons. Biochim Biophys Acta Mol Cell Biol Lipids 1439(1):47–56. https://doi.org/10.1016/s1388-1981(99)00073-6
Tumaney AW, Shekar S, Rajasekharan R (2001) Identification, purification, and characterization of monoacylglycerol acyltransferase from developing peanut cotyledons. J Biol Chem 276(14):10847–10852. https://doi.org/10.1074/jbc.m100005200
Umate P (2012) Comparative genomics of the lipid-body-membrane proteins oleosin, caleosin and steroleosin in magnoliophyte, lycophyte and bryophyte. Genom Proteom Bioinform 10(6):345–353. https://doi.org/10.1016/j.gpb.2012.08.006
Wang L, Shen W, Kazachkov M, Chen G, Chen Q, Carlsson AS, Stymne S, Weselake RJ, Zou J (2012) Metabolic Interactions between the lands cycle and the Kennedy pathway of glycerolipid synthesis in Arabidopsis developing seeds. Plant Cell 24(11):4652–4669. https://doi.org/10.1105/tpc.112.104604
Wang F, Chen H, Li X, Wang N, Wang T, Yang J, Guan L, Yao N, Du L, Wang Y, Liu X, Chen X, Wang Z, Dong Y, Li H (2015) Mining and identification of polyunsaturated fatty acid synthesis genes active during camelina seed development using 454 pyrosequencing. BMC Plant Biol. https://doi.org/10.1186/s12870-015-0513-6
Wanjie SW, Welti R, Moreau RA, Chapman KD (2005) Identification and quantification of glycerolipids in cotton fibers: reconciliation with metabolic pathway predictions from DNA databases. Lipids 40(8):773–785. https://doi.org/10.1007/s11745-005-1439-4
Welti R, Li W, Li M, Sang Y, Biesiada H, Zhou H-E, Rajashekar CB, Williams TD, Wang X (2002) Profiling membrane lipids in plant stress responses. J Biol Chem 277(35):31994–32002. https://doi.org/10.1074/jbc.m205375200
Winichayakul SCR, Scott R, Zhou X, Zou X, Roldan M, Richardson K, Roberts N (2008) Delivery of grasses with high levels of unsaturated, protected fatty acids. Proc N Z Grassl Assoc 70:211–216
Ulven SM, Holven KB (2015) Comparison of bioavailability of krill oil versus fish oil and health effect. Vascu Health Risk Manag. https://doi.org/10.2147/vhrm.s85165
Zheng Q, Li JQ, Kazachkov M, Liu K, Zou J (2012) Identification of Brassica napus lysophosphatidylcholine acyltransferase genes through yeast functional screening. Phytochemistry 75:21–31. https://doi.org/10.1016/j.phytochem.2011.11.022
Acknowledgements
We thank the Kansas Lipidomics Research Center Analytical Laboratory, Kansas State University for analyzing the plant lipid samples presented in this paper. We acknowledge Bionivid Technology Private Limited Bangalore, India for sequencing and analysis.
Funding
This work was supported by LIPIC, 12th five year plan, CSIR, New Delhi, India. Additional support was provided by JC Bose National Fellowship from the Department of Science and Technology, New Delhi for R.R. V.V. was supported by CSIR-Senior Research Fellowship.
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Venkateshwari, V., Vijayakumar, A., Vijayakumar, A.K. et al. Leaf lipidome and transcriptome profiling of Portulaca oleracea: characterization of lysophosphatidylcholine acyltransferase. Planta 248, 347–367 (2018). https://doi.org/10.1007/s00425-018-2908-8
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DOI: https://doi.org/10.1007/s00425-018-2908-8