Abstract
Main conclusion
Overexpression of Artemisia annua jasmonic acid carboxyl methyltransferase (AaJMT) leads to enhanced artemisinin content in Artemisia annua.
Abstract
Artemisinin-based combination therapies remain the sole deterrent against deadly disease malaria and Artemisia annua remains the only natural producer of artemisinin. In this study, the 1101 bp gene S-adenosyl-l-methionine (SAM): Artemisia annua jasmonic acid carboxyl methyltransferase (AaJMT), was characterised from A. annua, which converts jasmonic acid (JA) to methyl jasmonate (MeJA). From phylogenetic analysis, we confirmed that AaJMT shares a common ancestor with Arabidopsis thaliana, Eutrema japonica and has a close homology with JMT of Camellia sinensis. Further, the Clustal Omega depicted that the conserved motif I, motif III and motif SSSS (serine) required to bind SAM and JA, respectively, are present in AaJMT. The relative expression of AaJMT was induced by wounding, MeJA and salicylic acid (SA) treatments. Additionally, we found that the recombinant AaJMT protein catalyses the synthesis of MeJA from JA with a Km value of 37.16 µM. Moreover, site-directed mutagenesis of serine-151 in motif SSSS to tyrosine, asparagine-10 to threonine and glutamine-25 to histidine abolished the enzyme activity of AaJMT, thus indicating their determining role in JA substrate binding. The GC–MS analysis validated that mutant proteins of AaJMT were unable to convert JA into MeJA. Finally, the artemisinin biosynthetic and trichome developmental genes were upregulated in AaJMT overexpression transgenic lines, which in turn increased the artemisinin content.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
Data availability
The AaJMT sequences were retrieved from the National Centre for Biotechnology Information under ID AIN76708.1
Abbreviations
- JA:
-
Jasmonic acid
- JMT:
-
Jasmonic acid carboxyl methyltransferase
- MeJA:
-
Methyl jasmonate
- SA:
-
Salicylic acid
- SAM:
-
S-adenosyl-L-methionine
References
Altschul SF, Stephen F, Gish W, Miller W, Myers EW, Lipman DJ (1990) Basic local alignment search tool. J Mol Biol 215:403–410. https://doi.org/10.1016/S0022-2836(05)80360-2
Ashihara H, Mizuno K, Yokota T, Crozier A (2017) Xanthine alkaloids: occurrence, biosynthesis, and function in plants. Prog Chem Org Nat Prod 105:1–88. https://doi.org/10.1007/978-3-319-49712-9_1
Baba SA, Vishwakarma RA, Ashraf N (2017) Functional characterization of CsBGlu12, a β-glucosidase from Crocus sativus, provides insights into its role in abiotic stress through accumulation of antioxidant flavonols. J Biol Chem 292(11):4700–4713. https://doi.org/10.1074/jbc.M116.762161
Barkman TJ, Martins TR, Sutton E, Stout JT (2007) Positive selection for single amino acid change promotes substrate discrimination of a plant volatile producing enzyme. Mol Biol Evol 24:1320–1329. https://doi.org/10.1093/molbev/msm053
Beale MH, Ward JL (1998) Jasmonates: key players in the plant defence. Natural Prod Rep 15(6):533–548. https://doi.org/10.1039/A815533Y
Chini A, Fonseca SGDC, Fernandez G, Adie B, Chico JM, Lorenzo O, Solano R (2007) The JAZ family of repressors is the missing link in jasmonate signalling. Nature 448(7154):666–671. https://doi.org/10.1038/nature06006
Clarke SM, Cristescu SM, Miersch O, Harren FJ, Wasternack C, Mur LA (2009) Jasmonates act with salicylic acid to confer basal thermotolerance in Arabidopsis thaliana. New Phytol 182(1):175–187. https://doi.org/10.1111/j.1469-8137.2008.02735.x
Creelman RA, Mullet JE (1995) Jasmonic acid distribution and action in plants: Regulation during development and response to biotic and abiotic stress. Proc Natl Acad Sci USA 92(10):4114–4119. https://doi.org/10.1073/pnas.92.10.411
Creelman RA, Mullet JE (1997) Biosynthesis and action of jasmonates in plants. Annu Rev Plant Biol 48(1):355–381. https://doi.org/10.1146/annurev.arplant.48.1.355
Creelman RA, Tierney ML, Mullet JE (1992) Jasmonic acid/methyl jasmonate accumulate in wounded soybean hypocotyls and modulate wound gene expression. Proc Natl Acad Sci USA 89(11):4938–4941. https://doi.org/10.1073/pnas.89.11.4938
D’Auria JC, Chen F, Pichersky E (2003) The SABATH family of MTs in Arabidopsis thaliana and other plant species. Recent Adv Phytochem 37:253–283. https://doi.org/10.1016/S0079-9920(03)80026-6
Farmer EE, Ryan CA (1990) Interplant communication: airborne methyl jasmonate induces synthesis of proteinase inhibitors in plant leaves. Proc Natl Acad Sci USA 87(19):7713–7716. https://doi.org/10.1073/pnas.87.19.7713
Gavrilescu M (2021) Water, soil, and plants interactions in a threatened environment. Water 13(19):2746. https://doi.org/10.3390/w13192746
Harms K, Atzorn R, Brash A, Kuhn H, Wasternack C, Willmitzer L, Pena-Cortes H (1995) Expression of a flax allene oxide synthase cDNA leads to increased endogenous jasmonic acid (JA) levels in transgenic potato plants but not to a corresponding activation of JA-responding genes. Plant Cell 7(10):1645–1654. https://doi.org/10.1105/tpc.7.10.1645
Heil M, Ton J (2008) Long-distance signalling in plant defence. Trends Plant Sci 13(6):264–272. https://doi.org/10.1016/j.tplants.2008.03.005
Hussain A (2023) Recent trends on production sources, biosynthesis pathways and antiviral efficacies of artemisinin: a candidate phytomedicine against SARS-CoV-2. Curr Pharm Biotechnol 24:1859–1880. https://doi.org/10.2174/1389201024666230327082051
Ibrahim RK, Bruneau A, Bantignies B (1998) Plant O-methyltransferases: molecular analysis, common signature and classification. Plant Mol Biol 36:1–10. https://doi.org/10.1023/a:1005939803300
Karban R, Baldwin IT, Baxter KJ, Laue G, Felton GW (2000) Communication between plants: induced resistance in wild tobacco plants following clipping of neighboring sagebrush. Oecologia 125:66–71. https://doi.org/10.1007/PL00008892
Khan RA, Kumar A, Abbas N (2024) AaGL3-like is jasmonate-induced bHLH transcription factor that positively regulates trichome density in Artemisia annua. Gene 904:148213. https://doi.org/10.1016/j.gene.2024.148213
Kim BG, Kim DH, Hur HG, Lim J, Lim YH, Ahn JH (2005) O-methyltransferases from Arabidopsis thaliana. J Appl Biol Chem 48(3):113–119
Koeduka T, Suzuki H, Taguchi G, Matsui K (2020) Biochemical characterization of the jasmonic acid methyltransferase gene from wasabi (Eutrema japonicum). Plant Biotechnol 37(3):389–392. https://doi.org/10.5511/plantbiotechnology.20.0622a
Kost C, Heil M (2008) The defensive role of volatile emission and extrafloral nectar secretion for lima bean in nature. J Chem Ecol 34:2–13. https://doi.org/10.1007/s10886-007-9404-0
Liao Z, Liu X, Zheng J, Zhao C, Wang D, Xu Y, Sun C (2023) A multifunctional true caffeoyl coenzyme AO-methyltransferase enzyme participates in the biosynthesis of polymethoxylated flavones in citrus. Plant Physiol 192(3):2049–2066. https://doi.org/10.1093/plphys/kiad249
Lu X, Zhang L, Zhang F, Jiang W, Shen Q, Zhang L, Lv Z, Wang G, Tang K (2013) AaORA, a trichome-specific AP 2/ERF transcription factor of Artemisia annua, is a positive regulator in the artemisinin biosynthetic pathway and in disease resistance to Botrytis cinerea. New Phytol 198(4):1191–1202. https://doi.org/10.1111/nph.12207
Luo Q, Li N, Xu JW (2022) A methyltransferase LaeA regulates ganoderic acid biosynthesis in Ganoderma lingzhi. Front Microbiol 13:1025983. https://doi.org/10.3389/fmicb.2022.1025983
Majid I, Kumar A, Abbas N (2019) A basic helix loop helix transcription factor, AaMYC2-Like positively regulates artemisinin biosynthesis in Artemisia annua L. Ind Crops Prod 128:115–125. https://doi.org/10.1016/j.indcrop.2018.10.083
Mohammad, Hurrah IM, Kumar A, Abbas N (2023) Synergistic interaction of two bHLH transcription factors positively regulates artemisinin biosynthetic pathway in Artemisia annua L. Physiologia Plantarum 175(1):e13849. https://doi.org/10.1111/ppl.13849
Munemasa S, Mori IC, Murata Y (2011) Methyl jasmonate signaling and signal crosstalk between methyl jasmonate and abscisic acid in guard cells. Plant Signal Behav 6(7):939–941. https://doi.org/10.4161/psb.6.7.15439
Murfitt LM, Kolosova N, Mann CJ, Dudareva N (2000) Purification and characterization of S-adenosyl-l-methionine: benzoic acid carboxyl methyltransferase, the enzyme responsible for biosynthesis of the volatile ester methyl benzoate in flowers of Antirrhinum majus. Arch Biochem Biophys 382:145–151. https://doi.org/10.1006/abbi.2000.2008
Park SW, Kaimoyo E, Kumar D, Mosher S, Klessig DF (2007) Methyl salicylate is a critical mobile signal for plant systemic acquired resistance. Science 318(5847):113–116. https://doi.org/10.1126/science.1147113
Park HL, Bhoo SH, Lee SW, Cho MH (2024) Biochemical characterization of a regiospecific flavonoid 3′-O-methyltransferase from orange. Appl Biol Chem 67(1):4. https://doi.org/10.1186/s13765-023-00853-8
Qin GJ, Gu HY, Zhao YD, Ma ZQ, Shi GL, Yang Y, Pichersky E, Chen HD, Liu MH, Chen ZL, Qu LJ (2005) An indole-3-acetic acid carboxyl methyltransferase regulates Arabidopsis leaf development. Plant Cell 17:2693–2704. https://doi.org/10.1105/tpc.105.034959
Rosales-Campos AL, Gutiérrez-Ortega A (2019) Agrobacterium-mediated transformation of Nicotiana tabacum cv. Xanthi leaf explants. Bio-Protoc. https://doi.org/10.21769/BioProtoc.3150
Ross JR, Nam KH, D’Auria JC, Pichersky E (1999) S-adenosyl-lmethionine: salicylic acid carboxyl methyltransferase, an enzyme involved in floral scent production and plant defense represents a new class of plant methyltransferases. Arch Biochem Biophys 367:9–16. https://doi.org/10.1006/abbi.1999.1255
Ruan J, Zhou Y, Zhou M, Yan J, Khurshid M, Weng W, Zhang K (2019) Jasmonic acid signaling pathway in plants. Int J Mol Sci 20(10):2479. https://doi.org/10.3390/ijms20102479
Seo HS, Song JT, Cheong JJ, Lee YH, Lee YW, Hwang I, Lee JS, Choi YD (2001) Jasmonic acid carboxyl methyltransferase: a key enzyme for jasmonate-regulated plant responses. Proc Natl Acad Sci USA 98:4788–4793. https://doi.org/10.1073/pnas.081557298
Shen Q, Lu X, Yan T, Fu X, Lv Z, Zhang F, Tang K (2016) The jasmonate-responsive AaMYC 2 transcription factor positively regulates artemisinin biosynthesis in Artemisia annua. New Phytol 210(4):1269–1281. https://doi.org/10.1111/nph.13874
Sohn HB, Lee HY, Seo JS, Jung C, Jeon JH, Kim JH, Choi YD (2011) Overexpression of jasmonic acid carboxyl methyltransferase increases tuber yield and size in transgenic potato. Plant Biotechnol Rep 5:27–34. https://doi.org/10.1007/s11816-010-0153-0
Song MS, Kim DG, Lee SH (2005) Isolation and characterization of a jasmonic acid carboxyl methyltransferase gene from hot pepper (Capsicum annuum L.). J Plant Biol 48:292–297. https://doi.org/10.1007/BF03030525
Stintzi A, Browse J (2000) The Arabidopsis male-sterile mutant, opr3, lacks the 12-oxophytodienoic acid reductase required for jasmonate synthesis. Proc Natl Acad Sci USA 97(19):10625–10630. https://doi.org/10.1073/pnas.190264497
Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30(12):2725–2729. https://doi.org/10.1093/molbev/mst197
Varbanova M, Yamaguchi S, Yang Y, McKelvey K, Hanada A, Borochov R, Yu F, Jikumaru Y, Ross J, Cortes D, Ma C, Noel JP, Mander L, Shulaev V, Kamiya Y, Rodermel S, Weiss D, Pichersky E (2007) Methylation of gibberellins by Arabidopsis GAMT1 and GAMT2. Plant Cell 19:32–45. https://doi.org/10.1105/tpc.106.044602
Vogt T, Jones P (2000) Glycosyltransferases in plant natural product synthesis: characterization of a supergene family. Trends Plant Sci 5(9):380–386. https://doi.org/10.1016/s1360-1385(00)01720-9
Wang B, Wang S, Wang Z (2017) Genome-wide comprehensive analysis the molecular phylogenetic evaluation and tissue-specific expression of SABATH gene family in Salvia miltiorrhiza. Genes 8(12):365. https://doi.org/10.3390/genes8120365
Wang Y, Mostafa S, Zeng W, Jin B (2021) Function and mechanism of jasmonic acid in plant responses to abiotic and biotic stresses. Int J Mol Sci 22(16):8568. https://doi.org/10.3390/ijms22168568
Wang Y, Chen Y, Zhang J, Zhang C (2022) Overexpression of llm1 affects the synthesis of secondary metabolites of Aspergillus cristatus. Microorganisms 10(9):1707. https://doi.org/10.3390/microorganisms10091707
World Health Organization (2022) World malaria report 2022. World Health Organization
Yu X, Zhang W, Zhang Y, Zhang X, Lang D, Zhang X (2018) The roles of methyl jasmonate to stress in plants. Funct Plant Biol 46(3):197–212. https://doi.org/10.1071/FP18106
Zhang J, Jia H, Zhu B, Li J, Yang T, Zhang ZZ, Deng WW (2021) Molecular and biochemical characterization of jasmonic acid carboxyl methyltransferase involved in aroma compound production of methyl jasmonate during black tea processing. J Agric Food Chem 69(10):3154–3164. https://doi.org/10.1021/acs.jafc.0c06248
Zhao N, Guan J, Lin H, Chen F (2007) Molecular cloning and biochemical characterization of indole-3-acetic acid methyltransferase from poplar. Phytochemistry 68:1537–1544. https://doi.org/10.1016/j.phytochem.2007.03.041
Zhao N, Ferrer JL, Ross J, Guan J, Yang Y, Pichersky E, Noel JP, Chen F (2008) Structural, biochemical, and phylogenetic analyses suggest that indole-3-acetic acid methyltransferase is an evolutionarily ancient member of the SABATH family. Plant Physiol 146:455–467. https://doi.org/10.1104/pp.107.110049
Zhao N, Yao J, Chaiprasongsuk M, Li G, Guan J, Tschaplinski TJ, Chen F (2013) Molecular and biochemical characterization of the jasmonic acid methyltransferase gene from black cottonwood (Populus trichocarpa). Phytochemistry 94:74–81. https://doi.org/10.1016/j.phytochem.2013.06.014
Zhou MZ, Yan CY, Zeng Z, Luo L, Zeng W, Huang YH (2020) N-methyltransferases of caffeine biosynthetic pathway in plants. J Agric Food Chem 68(52):15359–15372. https://doi.org/10.1021/acs.jafc.0c06167
Zia R, Nawaz MS, Siddique MJ, Hakim S, Imran A (2021) Plant survival under drought stress: Implications, adaptive responses, and integrated rhizosphere management strategy for stress mitigation. Microbiol Res 242:126626. https://doi.org/10.1016/j.micres.2020.126626
Zubieta C, Ross JR, Koscheski P, Yang Y, Pichersky E, Noel JP (2003) Structural basis for substrate recognition in the salicylic acid carboxyl methyltransferase family. Plant Cell 15(8):1704–1716. https://doi.org/10.1105/tpc.014548
Acknowledgements
This work was supported by INSPIRE Faculty Project grant (GAP-2129), Women SERB Excellence Award from DST (GAP-2186) both from DST Govt. of India, and CSIR-IIIM Jammu lab reserve fund. We sincerely acknowledge ICMR for the first author’s fellowship. The article bears institutional manuscript number CSIR-IIIM/IPR/00588.
Author information
Authors and Affiliations
Contributions
NA conceived the idea, supervised the experiments and finalised the draft of the manuscript. IM performed the experiments and wrote the draft of the manuscript. AK performed GC–MS analysis.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare no conflict of interest.
Additional information
Communicated by Dorothea Bartels.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Hurrah, I.M., Kumar, A. & Abbas, N. Functional characterisation of Artemisia annua jasmonic acid carboxyl methyltransferase: a key enzyme enhancing artemisinin biosynthesis. Planta 259, 152 (2024). https://doi.org/10.1007/s00425-024-04433-y
Received:
Accepted:
Published:
DOI: https://doi.org/10.1007/s00425-024-04433-y