Engineered microorganisms unlock a new era of diterpenoid biomanufacturing: recent advances and future perspectives

Abstract

Diterpenoids are natural compounds composed of four isoprene units. They possess diverse biological activities and widespread applications in the cosmetics, food additives and pharmaceutical. With the rapid advancement of synthetic biology, the biomanufacturing of diterpenoids via microbial metabolism has witnessed substantial advancements. Microbial chassis such as Escherichia coli, Saccharomyces cerevisiae, Yarrowia lipolytica and Rhodosporidium toruloides, have been successfully engineered to enable efficient biosynthesis of these compounds, thereby demonstrating substantial potential for industrial-scale applications. In this review, the construction of diterpenoid biosynthetic pathways in microbial cell factories is summarized. The research progress and engineering strategies for efficient microbial synthesis of diterpenoids are discussed, and the key challenges and future directions facilitating the design of high-yield diterpenoid production platforms and their translation into industrial practice are explored.

Graphical abstract

Keywords

Diterpenoids / Synthetic biology / Biomanufacturing / Microbial cell factories

References

[1] Ajikumar PK, Xiao W-H, Tyo KEJ, Wang Y, Simeon F, Leonard E, Mucha O, Phon TH, Pfeifer B, Stephanopoulos G. Isoprenoid pathway optimization for taxol precursor overproduction in Escherichia coli. Science, 2010, 330: 70-74

[2] Arendt P, Miettinen K, Pollier J, De Rycke R, Callewaert N, Goossens A. An endoplasmic reticulum-engineered yeast platform for overproduction of triterpenoids. Metab Eng, 2017, 40: 165-175

[3] Arhar S, Natter K. Common aspects in the engineering of yeasts for fatty acid- and isoprene-based products. Biochimica et Biophysica Acta (BBA) - Mol Cell Biol Lipids, 2019, 1864(12): 158513

[4] Arnold FH. Directed evolution: bringing new chemistry to life. Angew Chem Int Ed Engl, 2018, 57: 4143-4148

[5] Asadollahi MA, Maury J, Møller K, Nielsen KF, Schalk M, Clark A, Nielsen J. Production of plant sesquiterpenes in Saccharomyces cerevisiae: effect of ERG9 repression on sesquiterpene biosynthesis. Biotechnol Bioeng, 2007, 99: 666-677

[6] Asadollahi MA, Maury J, Schalk M, Clark A, Nielsen J. Enhancement of farnesyl diphosphate pool as direct precursor of sesquiterpenes through metabolic engineering of the mevalonate pathway in Saccharomyces cerevisiae. Biotechnol Bioeng, 2010, 106: 86-96

[7] Bathe U, Tissier A. Cytochrome P450 enzymes: a driving force of plant diterpene diversity. Phytochemistry, 2019, 161: 149-162

[8] Bathe U, Frolov A, Porzel A, Tissier A. CYP76 oxidation network of abietane diterpenes in lamiaceae reconstituted in yeast. J Agric Food Chem, 2019, 67: 13437-13450

[9] Bian G, Deng Z, Liu T. Strategies for terpenoid overproduction and new terpenoid discovery. Curr Opin Biotechnol, 2017, 48: 234-241

[10] Bureau J-A, Oliva ME, Dong Y, Ignea C. Engineering yeast for the production of plant terpenoids using synthetic biology approaches. Nat Prod Rep, 2023, 40: 1822-1848

[11] Cao X, Yu W, Chen Y, Yang S, Zhao ZK, Nielsen J, Luan H, Zhou YJ. Engineering yeast for high-level production of diterpenoid sclareol. Metab Eng, 2023, 75: 19-28

[12] Castegna A, Scarcia P, Agrimi G, Palmieri L, Rottensteiner H, Spera I, Germinario L, Palmieri F. Identification and functional characterization of a novel mitochondrial carrier for citrate and oxoglutarate in Saccharomyces cerevisiae. J Biol Chem, 2010, 285: 17359-17370

[13] Chatzivasileiou AO, Ward V, Edgar SM, Stephanopoulos G. Two-step pathway for isoprenoid synthesis. Proc Natl Acad Sci U S A, 2019, 116: 506-511

[14] Chen Y, Siewers V, Nielsen J. Profiling of cytosolic and peroxisomal acetyl-CoA metabolism in Saccharomyces cerevisiae. PLoS One, 2012, 7 e42475

[15] Chen X, Zaro JL, Shen W-C. Fusion protein linkers: property, design and functionality. Adv Drug Deliv Rev, 2013, 65: 1357-1369

[16] Chen L, Pang Y, Luo Y, Cheng X, Lv B, Li C. Separation and purification of plant terpenoids from biotransformation. Eng Life Sci, 2021, 21: 724-738

[17] Chen R, Wang J, Xu J, Nie S, Chen C, Li Y, Li Y, He J, Li W, Wen M, Qiao J. Heterologous biosynthesis of kauralexin A1 in Saccharomyces cerevisiae through metabolic and enzyme engineering. J Agric Food Chem, 2024, 72: 7308-7317

[18] Chen J, Huang L, Ye BC, Zhou Y. Combinatorial metabolic engineering of Yarrowia lipolytica for high-level production of the plant-derived diterpenoid sclareol. Microb Cell Fact, 2025, 24: 110

[19] Choi BH, Kang HJ, Kim SC, Lee PC. Organelle engineering in yeast: enhanced production of protopanaxadiol through manipulation of peroxisome proliferation in Saccharomyces cerevisiae. Microorganisms, 2022

[20] Dekker L, Polizzi KM. Sense and sensitivity in bioprocessing-detecting cellular metabolites with biosensors. Curr Opin Chem Biol, 2017, 40: 31-36

[21] Delatte TL, Scaiola G, Molenaar J, de Sousa Farias K, Alves Gomes Albertti L, Busscher J, Verstappen F, Carollo C, Bouwmeester H, Beekwilder J. Engineering storage capacity for volatile sesquiterpenes in Nicotiana benthamiana leaves. Plant Biotechnol J, 2018, 16: 1997-2006

[22] Dinday S, Ghosh S. Recent advances in triterpenoid pathway elucidation and engineering. Biotechnol Adv, 2023, 68 108214

[23] Donald KA, Hampton RY, Fritz IB. Effects of overproduction of the catalytic domain of 3-hydroxy-3-methylglutaryl coenzyme A reductase on squalene synthesis in Saccharomyces cerevisiae. Appl Environ Microbiol, 1997, 63: 3341-3344

[24] Duran L, López JM, Avalos JL. ¡Viva la mitochondria!: harnessing yeast mitochondria for chemical production. FEMS Yeast Res, 2020

[25] Dusseaux S, Wajn WT, Liu Y, Ignea C, Kampranis SC. Transforming yeast peroxisomes into microfactories for the efficient production of high-value isoprenoids. Proc Natl Acad Sci U S A, 2020, 117: 31789-31799

[26] Einhaus A, Steube J, Freudenberg RA, Barczyk J, Baier T, Kruse O. Engineering a powerful green cell factory for robust photoautotrophic diterpenoid production. Metab Eng, 2022, 73: 82-90

[27] Engels B, Dahm P, Jennewein S. Metabolic engineering of taxadiene biosynthesis in yeast as a first step towards taxol (paclitaxel) production. Metab Eng, 2008, 10: 201-206

[28] Englund E, Andersen-Ranberg J, Miao R, Hamberger B, Lindberg P. Metabolic engineering of Synechocystis sp. PCC 6803 for production of the plant diterpenoid manoyl oxide. ACS Synth Biol, 2015, 4: 1270-1278

[29] Furubayashi M, Ikezumi M, Kajiwara J, Iwasaki M, Fujii A, Li L, Saito K, Umeno D. A high-throughput colorimetric screening assay for terpene synthase activity based on substrate consumption. PLoS ONE, 2014, 9 e93317

[30] Gao S, Tong Y, Zhu L, Ge M, Zhang Y, Chen D, Jiang Y, Yang S. Iterative integration of multiple-copy pathway genes in Yarrowia lipolytica for heterologous β-carotene production. Metab Eng, 2017, 41: 192-201

[31] Gao K, Zha W-L, Zhu J-X, Zheng C, Zi J-C. A review: biosynthesis of plant-derived labdane-related diterpenoids. Chin J Nat Med, 2021, 19: 666-674

[32] Gardenour KR, Levy J, Lopes JM. Identification of novel dominant INO2c mutants with an Opi- phenotype. Mol Microbiol, 2004, 52: 1271-1280

[33] Geiselman GM, Zhuang X, Kirby J, Tran-Gyamfi MB, Prahl J-P, Sundstrom ER, Gao Y, Munoz Munoz N, Nicora CD, Clay DM, Papa G, Burnum-Johnson KE, Magnuson JK, Tanjore D, Skerker JM, Gladden JM. Production of ent-kaurene from lignocellulosic hydrolysate in Rhodosporidium toruloides. Microb Cell Fact, 2020

[34] Ghodasara A, Voigt CA. Balancing gene expression without library construction via a reusable sRNA pool. Nucleic Acids Res, 2017, 45: 8116-8127

[35] Guerfal M, Claes K, Knittelfelder O, De Rycke R, Kohlwein SD, Callewaert N. Enhanced membrane protein expression by engineering increased intracellular membrane production. Microb Cell Fact, 2013, 12: 122

[36] Hafner J, Payne J, MohammadiPeyhani H, Hatzimanikatis V, Smolke C. A computational workflow for the expansion of heterologous biosynthetic pathways to natural product derivatives. Nat Commun, 2021, 12 1760

[37] Hammer SK, Avalos JL. Harnessing yeast organelles for metabolic engineering. Nat Chem Biol, 2017, 13: 823-832

[38] He B, Bai X, Tan Y, Xie W, Feng Y, Yang GY. Glycosyltransferases: mining, engineering and applications in biosynthesis of glycosylated plant natural products. Synth Syst Biotechnol, 2022, 7: 602-620

[39] Hu Z, He B, Ma L, Sun Y, Niu Y, Zeng B. Recent advances in ergosterol biosynthesis and regulation mechanisms in Saccharomyces cerevisiae. Indian J Microbiol, 2017, 57: 270-277

[40] Hu T, Zhou J, Tong Y, Su P, Li X, Liu Y, Liu N, Wu X, Zhang Y, Wang J, Gao L, Tu L, Lu Y, Jiang Z, Zhou YJ, Gao W, Huang L. Engineering chimeric diterpene synthases and isoprenoid biosynthetic pathways enables high-level production of miltiradiene in yeast. Metab Eng, 2020, 60: 87-96

[41] Huang ZY, Ye RY, Yu HL, Li AT, Xu JH. Mining methods and typical structural mechanisms of terpene cyclases. Bioresour Bioprocess, 2021, 8: 66

[42] Huang M, Zhang G, Chen h, Liu Q, Jia K-Z, Dai Z, Wang Q. Enhanced biosynthesis of sclareol in Yarrowia lipolytica via systematic metabolic engineering. ACS Synth Biol, 2026

[43] Hunkeler M, Hagmann A, Stuttfeld E, Chami M, Guri Y, Stahlberg H, Maier T. Structural basis for regulation of human acetyl-CoA carboxylase. Nature, 2018, 558: 470-474

[44] Ignea C, Ioannou E, Georgantea P, Loupassaki S, Trikka FA, Kanellis AK, Makris AM, Roussis V, Kampranis SC. Reconstructing the chemical diversity of labdane-type diterpene biosynthesis in yeast. Metab Eng, 2015, 28: 91-103

[45] Ignea C, Trikka FA, Nikolaidis AK, Georgantea P, Ioannou E, Loupassaki S, Kefalas P, Kanellis AK, Roussis V, Makris AM, Kampranis SC. Efficient diterpene production in yeast by engineering Erg20p into a geranylgeranyl diphosphate synthase. Metab Eng, 2015, 27: 65-75

[46] Jia M, Zhou K, Tufts S, Schulte S, Peters RJ. A pair of residues that interactively affect diterpene synthase product outcome. ACS Chem Biol, 2017, 12: 862-867

[47] Jiang GZ, Yao MD, Wang Y, Zhou L, Song TQ, Liu H, Xiao WH, Yuan YJ. Manipulation of GES and ERG20 for geraniol overproduction in Saccharomyces cerevisiae. Metab Eng, 2017, 41: 57-66

[48] Jin B, Xu K, Guo J, Ma Y, Yang J, Chen N, Zeng T, Wang J, Liu J, Tian M, Ma Q, Zhang H, Peters RJ, Cui G, Wu R, Huang L. From functional plasticity of two diterpene synthases (IrTPS2/IrKSL3a) to enzyme evolution. ACS Catal, 2024, 14: 2959-2970

[49] Ju H, Zhang C, Lu W. Progress in heterologous biosynthesis of forskolin. J Ind Microbiol Biotechnol, 2021

[50] Ju H, Zhang C, He S, Nan W, Lu W. Construction and optimization of Saccharomyces cerevisiae for synthesizing forskolin. Appl Microbiol Biotechnol, 2022, 106: 1933-1944

[51] Kildegaard KR, Arnesen JA, Adiego-Pérez B, Rago D, Kristensen M, Klitgaard AK, Hansen EH, Hansen J, Borodina I. Tailored biosynthesis of gibberellin plant hormones in yeast. Metab Eng, 2021, 66: 1-11

[52] Kim EM, Woo HM, Tian T, Yilmaz S, Javidpour P, Keasling JD, Lee TS. Autonomous control of metabolic state by a quorum sensing (QS)-mediated regulator for bisabolene production in engineered E. coli. Metab Eng, 2017, 44: 325-336

[53] Kim JE, Jang IS, Son SH, Ko YJ, Cho BK, Kim SC, Lee JY. Tailoring the Saccharomyces cerevisiae endoplasmic reticulum for functional assembly of terpene synthesis pathway. Metab Eng, 2019, 56: 50-59

[54] Kirby J, Nishimoto M, Park JG, Withers ST, Nowroozi F, Behrendt D, Rutledge EJ, Fortman JL, Johnson HE, Anderson JV, Keasling JD. Cloning of casbene and neocembrene synthases from Euphorbiaceae plants and expression in Saccharomyces cerevisiae. Phytochemistry, 2010, 71: 1466-1473

[55] Kong MK, Kang H-J, Kim JH, Oh SH, Lee PC. Metabolic engineering of the Stevia rebaudiana ent-kaurene biosynthetic pathway in recombinant Escherichia coli. J Biotechnol, 2015, 214: 95-102

[56] Kunhao L, Xiaoli W, Zedong H, Bin Z, Zhiqing M, Chuanshu Z. Efficient biosynthesis of the plant-derived diterpenoid 14-hydroxy-dehydroabietadiene in Saccharomyces cerevisiae. Microb Cell Fact, 2026, 25 41

[57] Lauchli R, Rabe KS, Kalbarczyk KZ, Tata A, Heel T, Kitto RZ, Arnold FH. High-throughput screening for terpene‐synthase‐cyclization activity and directed evolution of a terpene synthase. Angew Chem Int Ed Engl, 2013, 52: 5571-5574

[58] Lauersen KJ, Wichmann J, Baier T, Kampranis SC, Pateraki I, Møller BL, Kruse O. Phototrophic production of heterologous diterpenoids and a hydroxy-functionalized derivative from Chlamydomonas reinhardtii. Metab Eng, 2018, 49: 116-127

[59] Lee JW, Na D, Park JM, Lee J, Choi S, Lee SY. Systems metabolic engineering of microorganisms for natural and non-natural chemicals. Nat Chem Biol, 2012, 8: 536-546

[60] Lee HJ, Kim C, Heo YB, Kim S-E, Woo HM. Bacterial biosynthesis of abietane-type diterpene ferruginol from glucose. Microb Cell Fact, 2025, 24: 67-79

[61] Leonard E, Ajikumar PK, Thayer K, Xiao W-H, Mo JD, Tidor B, Stephanopoulos G, Prather KLJ. Combining metabolic and protein engineering of a terpenoid biosynthetic pathway for overproduction and selectivity control. Proc Natl Acad Sci U S A, 2010, 107: 13654-13659

[62] Li L, Wang X, Li X, Shi H, Wang F, Zhang Y, Li X. Combinatorial engineering of mevalonate pathway and diterpenoid synthases in Escherichia coli for cis-abienol production. J Agric Food Chem, 2019, 67: 6523-6531

[63] Li Z, Zhang L, Xu K, Jiang Y, Du J, Zhang X, Meng LH, Wu Q, Du L, Li X, Hu Y, Xie Z, Jiang X, Tang YJ, Wu R, Guo RT, Li S. Molecular insights into the catalytic promiscuity of a bacterial diterpene synthase. Nat Commun, 2023, 14 4001

[64] Li G, Liang H, Gao R, Qin L, Xu P, Huang M, Zong MH, Cao Y, Lou WY. Yeast metabolism adaptation for efficient terpenoids synthesis via isopentenol utilization. Nat Commun, 2024, 15 9844

[65] Li L, Fu J, Liu N. Advances in the structures, pharmacological activities, and biosynthesis of plant diterpenoids. J Microbiol Biotechnol, 2024, 34: 1563-1579

[66] Li S, Luo S, Yin X, Zhao X, Wang X, Gao S, Xu S, Lu J, Zhou J. Screening of ent-copalyl diphosphate synthase and metabolic engineering to achieve de novo biosynthesis of ent-copalol in Saccharomyces cerevisiae. Synth Syst Biotechnol, 2024, 9: 784-792

[67] Li Z, Jindani S, Kojasoy V, Ortega T, Marshall EM, Abboud KA, Loesgen S, Tantillo DJ, Rudolf JD. Computation-guided scaffold exploration of 2E,6E-1,10-trans/cis-eunicellanes. Beilstein J Org Chem, 2024, 20: 1320-1326

[68] Liang H, Deng X, Ji Q, Sun F, Shen T, He C. The Pseudomonas aeruginosa global regulator VqsR directly inhibits QscR to control quorum-sensing and virulence gene expression. J Bacteriol, 2012, 194: 3098-3108

[69] Liang X, Fan B, Wang J, Qin Y, Yu T, Xu P, Wang G, An T, Chen S. Advances in semi-rational design of terpene synthases and their modifying enzymes. AMB Express, 2025

[70] Liao P, Hemmerlin A, Bach TJ, Chye ML. The potential of the mevalonate pathway for enhanced isoprenoid production. Biotechnol Adv, 2016, 34: 697-713

[71] Lipko A, Pączkowski C, Perez-Fons L, Fraser PD, Kania M, Hoffman-Sommer M, Danikiewicz W, Rohmer M, Poznanski J, Swiezewska E. Divergent contribution of the MVA and MEP pathways to the formation of polyprenols and dolichols in Arabidopsis. Biochem J, 2023, 480: 495-520

[72] Liu D, Mannan AA, Han Y, Oyarzún DA, Zhang F. Dynamic metabolic control: towards precision engineering of metabolism. J Ind Microbiol Biotechnol, 2018, 45: 535-543

[73] Liu T, Zhang C, Lu W. Heterologous production of levopimaric acid in Saccharomyces cerevisiae. Microb Cell Fact, 2018

[74] Liu Z, Li J, Sun Y, Zhang P, Wang Y. Structural insights into the catalytic mechanism of a plant diterpene glycosyltransferase SrUGT76G1. Plant Commun, 2020, 1 100004

[75] Liu Q, Liu Y, Li G, Savolainen O, Chen Y, Nielsen J. De novo biosynthesis of bioactive isoflavonoids by engineered yeast cell factories. Nat Commun, 2021, 12: 6085

[76] Liu F, Liu SC, Qi YK, Liu Z, Chen J, Wei LJ, Hua Q. Enhancing trans-nerolidol productivity in Yarrowia lipolytica by improving precursor supply and optimizing nerolidol synthase activity. J Agric Food Chem, 2022, 70: 15157-15165

[77] Liu Q, Zhang G, Su L, Liu P, Jia S, Wang Q, Dai Z. Reprogramming the metabolism of oleaginous yeast for sustainably biosynthesizing the anticarcinogen precursor germacrene A. Green Chem, 2023, 25: 7988-7997

[78] Liu Y, Chen X, Zhang C. Sustainable biosynthesis of valuable diterpenes in microbes. Eng Microbiol, 2023, 3 100058

[79] Liu Z, Xie L, Chen W. Advancement of uridine diphosphate-dependent glycosyltransferases (UGTs) in the glycosylation modification of natural products and their protein engineering. Food Qual Saf, 2025

[80] Lu Y, Yang Q, Lin Z, Yang X. A modular pathway engineering strategy for the high-level production of β-ionone in Yarrowia lipolytica. Microb Cell Fact, 2020, 19 49

[81] Ma Y, Li W, Mai J, Wang J, Wei Y, Ledesma-Amaro R, Ji X-J. Engineering Yarrowia lipolytica for sustainable production of the chamomile sesquiterpene (–)-α-bisabolol. Green Chem, 2021, 23: 780-787

[82] Moser S, Pichler H. Identifying and engineering the ideal microbial terpenoid production host. Appl Microbiol Biotechnol, 2019, 103: 5501-5516

[83] Nielsen J. Synthetic biology for engineering acetyl coenzyme A metabolism in yeast. mBio, 2014, 5 e02153

[84] Nielsen J, Keasling JD. Engineering cellular metabolism. Cell, 2016, 164: 1185-1197

[85] Nowrouzi B, Li RA, Walls LE, d’Espaux L, Malcı K, Liang L, Jonguitud-Borrego N, Lerma-Escalera AI, Morones-Ramirez JR, Keasling JD, Rios-Solis L. Enhanced production of taxadiene in Saccharomyces cerevisiae. Microb Cell Fact, 2020, 19 200

[86] Nowrouzi B, Lungang L, Rios-Solis L. Exploring optimal Taxol(R) CYP725A4 activity in Saccharomyces cerevisiae. Microb Cell Fact, 2022, 21: 197

[87] O’Leary K. AlphaFold gets an upgrade (and a Nobel). Nat Med, 2024, 30: 3393

[88] Okumoto K, Tamura S, Honsho M, Fujiki Y. Peroxisome: metabolic functions and biogenesis. Adv Exp Med Biol, 2020, 1299: 3-17

[89] Pan X, Rudolf JD, Dong L-B. Class II terpene cyclases: structures, mechanisms, and engineering. Nat Prod Rep, 2024, 41: 402-433

[90] Paramasivan K, Mutturi S. Progress in terpene synthesis strategies through engineering of Saccharomyces cerevisiae. Crit Rev Biotechnol, 2017, 37: 974-989

[91] Pateraki I, Andersen-Ranberg J, Hamberger B, Heskes AM, Martens HJ, Zerbe P, Bach SS, Møller BL, Bohlmann J, Hamberger B. Manoyl oxide (13R), the biosynthetic precursor of forskolin, is synthesized in specialized root cork cells in Coleus forskohlii. Plant Physiol, 2014, 164: 1222-1236

[92] Peralta-Yahya PP, Ouellet M, Chan R, Mukhopadhyay A, Keasling JD, Lee TS. Identification and microbial production of a terpene-based advanced biofuel. Nat Commun, 2011, 2: 483

[93] Pickens LB, Tang Y, Chooi YH. Metabolic engineering for the production of natural products. Annu Rev Chem Biomol Eng, 2011, 2: 211-236

[94] Portela RM, Vogl T, Kniely C, Fischer JE, Oliveira R, Glieder A. Synthetic core promoters as universal parts for fine-tuning expression in different yeast species. ACS Synth Biol, 2017, 6: 471-484

[95] Reider Apel A, d'Espaux L, Wehrs M, Sachs D, Li RA, Tong GJ, Garber M, Nnadi O, Zhuang W, Hillson NJ, Keasling JD, Mukhopadhyay A. A Cas9-based toolkit to program gene expression in Saccharomyces cerevisiae. Nucleic Acids Res, 2017, 45: 496-508

[96] Ren J, Wu Y, Zhu Z, Chen R, Zhang L. Biosynthesis and regulation of diterpenoids in medicinal plants. Chin J Nat Med, 2022, 20: 761-772

[97] Rodriguez S, Denby CM, Van Vu T, Baidoo EE, Wang G, Keasling JD. ATP citrate lyase mediated cytosolic acetyl-CoA biosynthesis increases mevalonate production in Saccharomyces cerevisiae. Microb Cell Fact, 2016, 15: 48

[98] Sheldon RA, Pereira PC. Biocatalysis engineering: the big picture. Chem Soc Rev, 2017, 46: 2678-2691

[99] Shen X, Wang J, Li C, Yuan Q, Yan Y. Dynamic gene expression engineering as a tool in pathway engineering. Curr Opin Biotechnol, 2019, 59: 122-129

[100] Shoji T, Tanaka Y, Nakashima Y, Mizohata E, Komaki M, Sugawara S, Takaya J, Yonekura-Sakakibara K, Morita H, Saito K, Hirai T. Enhanced production of rebaudioside D and rebaudioside M through V155T substitution in the glycosyltransferase UGT91D2 from Stevia rebaudiana. J Agric Food Chem, 2025, 73: 2019-2032

[101] Sun Y, Chen Z, Wang G, Lv H, Mao Y, Ma K, Wang Y. De novo production of versatile oxidized kaurene diterpenes in Escherichia coli. Metab Eng, 2022, 73: 201-213

[102] Sun M-L, Han Y, Yu X, Wang K, Lin L, Ledesma-Amaro R, Ji X-J. Constructing a green oleaginous yeast cell factory for sustainable production of the plant-derived diterpenoid sclareol. Green Chem, 2024, 26: 5202-5210

[103] Tai M, Stephanopoulos G. Engineering the push and pull of lipid biosynthesis in oleaginous yeast Yarrowia lipolytica for biofuel production. Metab Eng, 2013, 15: 1-9

[104] Tang MC, Zou Y, Watanabe K, Walsh CT, Tang Y. Oxidative cyclization in natural product biosynthesis. Chem Rev, 2017, 117: 5226-5333

[105] Tholl D. Biosynthesis and biological functions of terpenoids in plants. Adv Biochem Eng Biotechnol, 2015, 148: 63-106

[106] Tokuhiro K, Muramatsu M, Ohto C, Kawaguchi T, Obata S, Muramoto N, Hirai M, Takahashi H, Kondo A, Sakuradani E, Shimizu S. Overproduction of geranylgeraniol by metabolically engineered Saccharomyces cerevisiae. Appl Environ Microbiol, 2009, 75: 5536-5543

[107] Tsao CY, Hooshangi S, Wu HC, Valdes JJ, Bentley WE. Autonomous induction of recombinant proteins by minimally rewiring native quorum sensing regulon of E. coli. Metab Eng, 2010, 12: 291-297

[108] Walls LE, Malcı K, Nowrouzi B, Li RA, d'Espaux L, Wong J, Dennis JA, Semião AJC, Wallace S, Martinez JL, Keasling JD, Rios-Solis L. Optimizing the biosynthesis of oxygenated and acetylated Taxol precursors in Saccharomyces cerevisiae using advanced bioprocessing strategies. Biotechnol Bioeng, 2020, 118: 279-293

[109] Wang J, Ledesma-Amaro R, Wei Y, Ji B, Ji XJ. Metabolic engineering for increased lipid accumulation in Yarrowia lipolytica-a review. Bioresour Technol, 2020, 313 123707

[110] Wang K, Yin M, Sun ML, Zhao Q, Ledesma-Amaro R, Ji XJ, Lin L. Engineering Yarrowia lipolytica for efficient synthesis of geranylgeraniol. J Agric Food Chem, 2024, 72: 20568-20581

[111] Wang S, Liang M, Chen W, Wan H, Meng X, Zhu X, Lu Y, Shen Q, Jiang C, Xie N, Chen S, Jia M, Sun W. Completing the biosynthesis of the clinically important diterpenoid andrographolide in Andrographis paniculata. Angew Chem Int Ed, 2025

[112] Weaver BA. How taxol/paclitaxel kills cancer cells. Mol Biol Cell, 2014, 25: 2677-2681

[113] Wei P, Zhang C, Bian X, Lu W. Metabolic engineering of Saccharomyces cerevisiae for heterologous carnosic acid production. Front Bioeng Biotechnol, 2022, 10 916605

[114] Wendt KU, Schulz GE. Isoprenoid biosynthesis: manifold chemistry catalyzed by similar enzymes. Structure, 1998, 6: 127-133

[115] Wilder CN, Diggle SP, Schuster M. Cooperation and cheating in Pseudomonas aeruginosa: the roles of the las, rhl and pqs quorum-sensing systems. ISME J, 2011, 5: 1332-1343

[116] Wong J, de Rond T, d’ Espaux L, van der Horst C, Dev I, Rios-Solis L, Kirby J, Scheller H, Keasling J. High-titer production of lathyrane diterpenoids from sugar by engineered Saccharomyces cerevisiae. Metab Eng, 2018, 45: 142-148

[117] Wu Z, Gao J, Gao N, Zhao Y, Zhou YJ. Engineering chronological lifespan toward a robust yeast cell factory. Proc Natl Acad Sci U S A, 2025, 122 e2515324122

[118] Xie W, Ye L, Lv X, Xu H, Yu H. Sequential control of biosynthetic pathways for balanced utilization of metabolic intermediates in Saccharomyces cerevisiae. Metab Eng, 2015, 28: 8-18

[119] Xu B, Tantillo DJ, Rudolf JD. Mechanistic insights into the formation of the 6,10-bicyclic eunicellane skeleton by the bacterial diterpene synthase Bnd4. Angew Chem Int Ed Engl, 2021, 60: 23159-23163

[120] Xu Y, Wang X, Zhang C, Zhou X, Xu X, Han L, Lv X, Liu Y, Liu S, Li J, Du G, Chen J, Ledesma-Amaro R, Liu L. De novo biosynthesis of rubusoside and rebaudiosides in engineered yeasts. Nat Commun, 2022, 13 3040

[121] Xu M, Xie W, Luo Z, Li C-X, Hua Q, Xu J-H. Improving solubility and copy number of taxadiene synthase to enhance the titer of taxadiene in Yarrowia lipolytica. Synth Syst Biotechnol, 2023, 8: 331-338

[122] Yanagibashi S, Bamba T, Kirisako T, Kondo A, Hasunuma T. The potency of mitochondria enlargement for mitochondria-mediated terpenoid production in yeast. Appl Microbiol Biotechnol, 2024, 108: 110

[123] Yang X, Nambou K, Wei L, Hua Q. Heterologous production of α-farnesene in metabolically engineered strains of Yarrowia lipolytica. Bioresour Technol, 2016, 216: 1040-1048

[124] Yang YL, Zhang S, Ma K, Xu Y, Tao Q, Chen Y, Chen J, Guo S, Ren J, Wang W, Tao Y, Yin WB, Liu H. Discovery and characterization of a new family of diterpene cyclases in bacteria and fungi. Angew Chem Int Ed Engl, 2017, 56: 4749-4752

[125] Yang H, Zhang K, Shen W, Chen L, Xia Y, Zou W, Cao Y, Chen X. Efficient production of cembratriene-ol in Escherichia coli via systematic optimization. Microb Cell Fact, 2023, 22 17

[126] Yang H, Zhang K, Shen W, Xia Y, Li Y, Chen X. Boosting production of cembratriene-ol in Saccharomyces cerevisiae via systematic optimization. Biotechnol J, 2023

[127] Yin Z, Dickschat JS. Engineering fungal terpene biosynthesis. Nat Prod Rep, 2023, 40: 28-45

[128] Yin Z, Goldfuss B, Dickschat JS. Double bond geometry controls diterpene synthase catalysis. ACS Catal, 2026

[129] Yuan J, Ching C-B. Dynamic control of ERG9 expression for improved amorpha-4,11-diene production in Saccharomyces cerevisiae. Microb Cell Fact, 2015, 14: 38-47

[130] Yuzbasheva EY, Agrimi G, Yuzbashev TV, Scarcia P, Vinogradova EB, Palmieri L, Shutov AV, Kosikhina IM, Palmieri F, Sineoky SP. The mitochondrial citrate carrier in Yarrowia lipolytica: Its identification, characterization and functional significance for the production of citric acid. Metab Eng, 2019, 54: 264-274

[131] Zargar A, Quan DN, Emamian M, Tsao CY, Wu HC, Virgile CR, Bentley WE. Rational design of ‘controller cells’ to manipulate protein and phenotype expression. Metab Eng, 2015, 30: 61-68

[132] Zerbe P, Chiang A, Dullat H, O'Neil-Johnson M, Starks C, Hamberger B, Bohlmann J. Diterpene synthases of the biosynthetic system of medicinally active diterpenoids in Marrubium vulgare. Plant J, 2014, 79: 914-927

[133] Zhang Y, Jiang P, Ye M, Kim SH, Jiang C, Lü J. Tanshinones: sources, pharmacokinetics and anti-cancer activities. Int J Mol Sci, 2012, 13: 13621-13666

[134] Zhang C, Ju H, Lu CZ, Zhao F, Liu J, Guo X, Wu Y, Zhao GR, Lu W. High-titer production of 13R-manoyl oxide in metabolically engineered Saccharomyces cerevisiae. Microb Cell Fact, 2019, 18: 73

[135] Zhang X, Zhu K, Shi H, Wang X, Zhang Y, Wang F, Li X. Engineering Escherichia coli for effective and economic production of cis-abienol by optimizing isopentenol utilization pathway. J Clean Pro, 2022, 351: 131310

[136] Zhang TL, Yu HW, Ye LD. Metabolic engineering of for terpenoid production: tools and strategies. ACS Synth Biol, 2023, 12: 639-656

[137] Zhang Y, Wiese L, Fang H, Alseekh S, Perez de Souza L, Scossa F, Molloy JJ, Christmann M, Fernie AR. Synthetic biology identifies the minimal gene set required for paclitaxel biosynthesis in a plant chassis. Mol Plant, 2023, 16: 1951-1961

[138] Zhang M, Bai F, Shen Y, Cai P, Zhou YJ. Expanding the versatility of the protein master Pichia pastoris towards efficient biosynthesis of diterpenoid sclareol. Trends Biotechnol, 2026

[139] Zhao F, Bai P, Liu T, Li D, Zhang X, Lu W, Yuan Y. Optimization of a cytochrome P450 oxidation system for enhancing protopanaxadiol production in Saccharomyces cerevisiae. Biotechnol Bioeng, 2016, 113: 1787-1795

[140] Zhao C, Kim Y, Zeng Y, Li M, Wang X, Hu C, Gorman C, Dai SY, Ding SY, Yuan JS. Co-compartmentation of terpene biosynthesis and storage via synthetic droplet. ACS Synth Biol, 2018, 7: 774-781

[141] Zhao Y, Wu Z, Li J, Qi Y, Zhang X, Shen C. The key role of cytochrome P450s in the biosynthesis of plant derived natural products. Plant Physiol Biochem, 2025

[142] Zhou YJ, Gao W, Rong Q, Jin G, Chu H, Liu W, Yang W, Zhu Z, Li G, Zhu G, Huang L, Zhao ZK. Modular pathway engineering of diterpenoid synthases and the mevalonic acid pathway for miltiradiene production. J Am Chem Soc, 2012, 134: 3234-3241

[143] Zhu ZT, Du MM, Gao B, Tao XY, Zhao M, Ren YH, Wang FQ, Wei DZ. Metabolic compartmentalization in yeast mitochondria: burden and solution for squalene overproduction. Metab Eng, 2021, 68: 232-245

Funding

National Natural Science Foundation of China(22578214)

Jiangsu Basic Research Center for Synthetic Biology(BK20233003)

State Key Laboratory of Materials-Oriented Chemical Engineering(SKL-MCE-24A10)

Jiangsu Synergetic Innovation Center for Advanced Bio-Manufacture(XTSW4B04)

Jiangsu Funding Program for Excellent Postdoctoral Talent(2025ZB628)

Natural Science Research of Jiangsu Higher Education Institutions of China(25KJB180005)

RIGHTS & PERMISSIONS

The Author(s)