Abstract
Artemisinin, a potent antimalarial sesquiterpene lactone, is produced in low quantities by the plant Artemisia annua L. We used inhibitors of both the mevalonate and nonmevalonate terpenoid pathways to study in both seedlings and hairy root cultures the source of isopentenyl diphosphate (IPP), the channeling of carbon from sterols to sesquiterpenes, and the role that sugars may play in controlling artemisinin biosynthesis. Together, our results indicated that artemisinin is likely biosynthesized from IPP pools originating in both the plastid and the cytosol and that channeling of carbon can be directed away from competing sterol pathways and toward sesquiterpenes. Although glucose stimulated artemisinin production, the response is very complex with ratios of glucose to fructose involved; artemisinin levels increased proportionate to increasing amounts of glucose. Disaccharides mainly inhibited artemisinin production, but the response was less definitive. Glucose also increased expression of some of the genes in the artemisinin biosynthetic pathway, thereby suggesting that this sugar is acting not only as a carbon source but also as a signal. As we develop a better understanding of the regulation of the artemisinin biosynthetic pathway, results suggest that many factors can possibly be harnessed to increase artemisinin production in A. annua.
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References
Adam K, Zapp J (1998) Biosynthesis of the isoprene units of chamomile sesquiterpenes. Phytochemistry 48:953–959
Adam K, Thiel R, Zapp J, Becker H (1998) Involvement of the mevalonic acid pathway and the glyceraldehyde-pyruvate pathway in terpenoid biosynthesis of the liverworts Ricciocarpos natans and Conocephalum conicum. Arch Biochem Biophys 354:181–187
Arsenault PR, Vail D, Wobbe KK, Weathers PJ (2010a) Effect of sugars on artemisinin production in Artemisia annua L.: transcription and metabolite measurements. Molecules 15:2302–2318
Arsenault PR, Vail D, Wobbe KK, Erickson K, Weathers PJ (2010b) Reproductive development modulates gene expression and metabolite levels with possible feedback inhibition of artemisinin in Artemisia annua. Plant Physiol 154:958–968
Asadollahi MA, Maury J, Møller K, Nielsen KF, Schalk M, Clark A, Nielsen J (2008) Production of plant sesquiterpenes in Saccharomyces cerevisiae: effect of ERG9 repression on sesquiterpene biosynthesis. Biotechnol Bioeng 99:666–677
Bach TJ, Lichtenthaler HK (1983) Inhibition by mevinolin of plant growth, sterol formation and pigment accumulation. Physiol Plant 59:50–60
Baldi A, Dixit VK (2008) Yield enhancement strategies for artemisinin production by suspension cultures of Artemisia annua. Bioresour Technol 99:4609–4614
Baskin TI, Remillong EL, Wilson JE (2001) The impact of mannose and other carbon sources on the elongation and diameter of the primary root of Arabidopsis thaliana. Aust J Plant Physiol 28:481–488
Biggs AR (1990) Managing wound-associated disease by understanding wound healing in the bark of woody plants. J Arboric 16:108–112
Cortès S, Gromova M, Evrard A, Roby C, Heyraud A, Rolin DB, Raymond P, Brouquisse RM (2003) In plants, 3-O-methylglucose is phosphorylated by hexokinase but not perceived as a sugar. Plant Physiol 131:824–837
D’Angelo JG, Bordón C, Posner GH, Yolken R, Jones-Brando L (2009) Artemisinin derivatives inhibit Toxoplasma gondii in vitro at multiple steps in the lytic cycle. J Antimicrob Chemother 63:146–150
de Vries PJ, Dien TK (1996) Clinical pharmacology and therapeutic potential of artemisinin and its derivatives in the treatment of malaria. Drugs 52:818–836
Dudareva N, Andersson S, Orlova I, Gatto N, Reichelt M, Rhodes D, Boland W, Gershenzon J (2005) The nonmevalonate pathway supports both monoterpene and sesquiterpene formation in snapdragon flowers. Proc Natl Acad Sci USA 102:933–938
Efferth T, Marschall M, Wang X, Huong SM, Hauber I, Olbrich A, Kronschnabl M, Stamminger T, Huang ES (2002) Activity of artesunate towards wild-type, recombinant GFP-expressing sensitive and ganciclovir-resistant human cytomegaloviruses. J Mol Med 80:233–242
Ferreira JF, Simon JE, Janick J (1995) Relationship of artemisinin content of tissue-cultured, greenhouse-grown, and field-grown plants of Artemisia annua. Planta Med 61:351–355
Gibson SI (2000) Plant sugar-response pathways: part of a complex regulatory web. Plant Physiol 124:1532–1539
Gonzali S, Alpi A, Blando F, Bellis LD (2002) Arabiodopsis (HXK1 and HXK2) and yeast (HXK2) hexokinases over expressed in transgenic lines are characterized by different catalytic properties. Plant Sci 163:943–954
Hampel D, Mosandl A, Wust M (2005) Biosynthesis of mono- and sesquiterpenes in carrot roots and leaves (Daucus carota L.): metabolic cross talk of cytosolic mevalonate and plastidial methylerythritol phosphate pathways. Phytochemistry 66:305–311
Jang JC, Sheen J (1994) Sugar sensing in higher plants. Plant Cell 6:1665–1679
Jones-Brando L, D’Angelo J, Posner GH, Yolken R (2006) In vitro inhibition of Toxoplasma gondii by four new derivatives of artemisinin. Antimicrob Agents Chemother 50:4206–4208
Kim P (2004) Current studies on biological tagatose production using L-arabinose isomerase: a review and future perspective. Appl Microbiol Biotechnol 65:243–249
Kudakasseril GJ, Lam L, Staba EJ (1987) Effect of sterol inhibitors on the incorporation of 14C-isopentenyl pyrophosphate into artemisinin by a cell-free system from Artemisia annua tissue cultures and plants. Planta Med 53:280–284
Larronde F, Krisa S, Decendit A, Chèze C, Deffieux G, Mérillon JM (1998) Regulation of polyphenol production in Vitis vinifera cell suspension cultures by sugars. Plant Cell Rep 17:946–950
Laule O, Fürholz A, Chang HS, Zhu T, Wang X, Heifetz PB, Gruissem W, Lange BM (2003) Crosstalk between cytosolic and plastidial pathways of isoprenoid biosynthesis in Arabidopsis thaliana. Proc Natl Acad Sci USA 100:6866–6871
Maier W, Schneider B, Strack D (1998) Biosynthesis of sesquiterpenoid cyclohexenone derivatives in mycorrhizal barley roots proceeds via the glyceraldehyde 3-phosphate/pyruvate pathway. Tetrahedron Lett 39:521–524
Mandels M, Reese ET (1965) Inhibition of cellulases. Annu Rev Phytopathol 3:85–102
Mannan A, Liu CZ, Arsenault PR, Towler MJ, Vail DR, Lorence A, Weathers PJ (2010) DMSO triggers the generation of ROS leading to an increase in artemisinin and dihydroartemisinic acid in Artemisia annua shoot cultures. Plant Cell Rep 29:143–152
Merali S, Meshnick SR (1991) Susceptibility of Pneumocystis carinii to artemisinin in vitro. Antimicrob Agents Chemother 35:1225–1227
Nam W, Tak J, Ryu JK, Jung M, Yook JI, Kim HJ, Cha IH (2007) Effects of artemisinin and its derivatives on growth inhibition and apoptosis of oral cancer cells. Head Neck 29:335–340
Nguyen KT, Arsenault PR, Weathers PJ (2011) Trichomes + roots + ROS = artemisinin: regulating artemisinin biosynthesis in Artemisia annua L. In Vitro Cell Dev Biol Plant 47:329–338
Pego JV, Weisbeek PJ, Smeekens SCM (1999) Mannose inhibits Arabidopsis germination via a hexokinase-mediated step. Plant Physiol 119:1017–1023
Putalun W, Luealon W, De-Eknamkul W, Tanaka H, Shoyama Y (2007) Improvement of artemisinin production by chitosan in hairy root cultures of Artemisia annua L. Biotechnol Lett 29:1143–1146
Rédei GP (1974) ‘Fructose effect’ in higher plants. Ann Bot 38:287–297
Ro DK, Paradise EM, Ouellet M, Fisher KJ, Newman KL, Ndungu JM, Ho KA, Eachus RA, Ham TS, Kirby J, Chang MC, Withers ST, Shiba Y, Sarpong R, Keasling JD (2006) Production of the antimalarial drug precursor artemisinic acid in engineered yeast. Nature 440:940–943
Rodríguez-Concepción M, Forés O, Martínez-García JF, González V, Phillips MA, Ferrer A, Boronat A (2004) Distinct light-mediated pathways regulate the biosynthesis and exchange of isoprenoid precursors during Arabidopsis seedling development. Plant Cell 16:144–156
Rolland F, Baena-Gonzalez E, Sheen J (2006) Sugar sensing and signaling in plants: conserved and novel mechanisms. Annu Rev Plant Biol 57:675–709
Romero MR, Efferth T, Serrano MA, Castaño B, Macias RIR, Briz O, Marin JJG (2005) Effect of artemisinin/artesunate as inhibitors of hepatitis B virus production in an “in vitro” replicative system. Antiviral Res 68:75–83
Ryan JF (1996) The examination of culture conditions, biomass optimization, and terpene elicitation with chitosan, heavy metals, and light exposure of transformed roots cultures of Artemisia annua. M.S. thesis (Biotechnology), Worcester Polytechnic Institute, Worcester
Schenk N, Hsiao K-C, Bornman CH (1991) Avoidance of precipitation and carbohydrate breakdown in autoclaved plant tissue culture medium. Plant Cell Rep 10:115–119
Singh NP, Lai HC (2004) Artemisinin induces apoptosis in human cancer cells. Anticancer Res 24:2277–2280
Sinha AK, Hofmann MG, Römer U, Köckenberger W, Elling L, Roitsch T (2002) Metabolizable and non-metabolizable sugars activate different signal transduction pathways in tomato. Plant Physiol 128:1480–1489
Sivakumar G, Liu C, Towler MJ, Weathers PJ (2010) Biomass production of hairy roots of Artemisia annua and Arachis hypogaea in a scaled-up mist bioreactor. Biotechnol Bioeng 107:802–813
Souret FF, Kim Y, Wyslouzil BE, Wobbe KK, Weathers PJ (2003) Scale-up of Artemisia annua L. hairy root cultures produces complex patterns of terpenoid gene expression. Biotechnol Bioeng 83:653–667
Steliopoulos P, Wüst M, Adam K-P, Mosandl A (2002) Biosynthesis of the sesquiterpene germacrene D in Solidago canadensis: 13C and 2H labeling studies. Phytochemistry 60:13–20
Towler MJ, Weathers PJ (2007) Evidence of artemisinin production from IPP stemming from both the mevalonate and the nonmevalonate pathways. Plant Cell Rep 26:2129–2136
Utzinger J, Xiao S, N’Goran EK, Bergquist R, Tanner M (2001) The potential of artemether for the control of schistosomiasis. Int J Parasitol 31:1549–1562
Vitrac X, Larronde F, Krisa S, Decendit A, Deffieux G, Mérillon JM (2000) Sugar sensing and Ca2+-calmodulin requirement in Vitis vinifera cells producing anthocyanins. Phytochemistry 53:659–665
Vögeli U, Chappell J (1988) Induction of sesquiterpene cyclase and suppression of squalene synthetase activities in plant cell cultures treated with fungal elicitor. Plant Physiol 88:1291–1296
Wang JW, Tan RX (2002) Artemisinin production in Artemisia annua hairy root cultures with improved growth by altering the nitrogen source in the medium. Biotechnol Lett 24:1153–1156
Wang Y, Weathers PJ (2007) Sugars proportionately affect artemisinin production. Plant Cell Rep 26:1073–1081
Wang Y, Zhang H, Zhao B, Yuan X (2001) Improved growth of Artemisia annua L. roots and artemisinin production under red light conditions. Biotechnol Lett 23:1971–1973
Weathers PJ, DeJesus-Gonzalez L, Kim YJ, Souret FF, Towler MJ (2004) Alteration of biomass and artemisinin production in Artemisia annua hairy roots by media sterilization method and sugars. Plant Cell Rep 23:414–418
Weathers PJ, Bunk G, McCoy MC (2005) The effect of phytohormones on growth and artemisinin production in Artemisia annua hairy roots. In Vitro Cell Dev Biol Plant 41:47–53
Woerdenbag HJ, Lüers JFJ, Uden W, Pras N, Malingré T, Alfermann AW (1993) Production of the new antimalarial drug artemisinin in shoot cultures of Artemisia annua L. Plant Cell Tissue Cult 32:247–257
Zarn JA, Brüschweiler BJ, Schlatter JR (2003) Azole fungicides affect mammalian steroidogenesis by inhibiting sterol 14α-demethylase and aromatase. Environ Health Perspect 111:255–261
Acknowledgments
The authors are indebted to Worcester Polytechnic Institute, the Arkansas Bioscience Institute, and NIH 2R15GM069562-03 for partial financial support of this work. Special thanks to Dan Vail (WPI) for Fig. 8.7.
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Weathers, P., Towler, M., Wang, Y., Wobbe, K.K. (2012). Artemisinin: Controlling Its Production in Artemisia annua . In: Bach, T., Rohmer, M. (eds) Isoprenoid Synthesis in Plants and Microorganisms. Springer, New York, NY. https://doi.org/10.1007/978-1-4614-4063-5_8
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