Silent constraints: the hidden challenges faced in plant metabolic engineering.

Metabolic engineering is embraced as a method to sustainably enhance production of valuable phytochemicals with beneficial properties. However, successful production of these compounds in plants is not always predictable even when the pathways are fully known, frequently due to the lack of comprehensive understanding of plant metabolism as a whole, and interconnections between different primary, secondary, and hormone metabolic networks. Here, we highlight critical hidden constraints, including substrate availability, silent metabolism, and metabolic crosstalk, that impair engineering strategies. We explore how these constraints have historically been manifested in engineering attempts and propose how modern advancements will enable future strategies to overcome these impediments.

[1]  N. Dudareva,et al.  A peroxisomal thioesterase plays auxiliary roles in plant β-oxidative benzoic acid metabolism. , 2018, The Plant journal : for cell and molecular biology.

[2]  E. Schijlen,et al.  Novel routes towards bioplastics from plants: elucidation of the methylperillate biosynthesis pathway from Salvia dorisiana trichomes , 2020, Journal of experimental botany.

[3]  W. Schwab,et al.  Expression of Clarkia S-linalool synthase in transgenic petunia plants results in the accumulation of S-linalyl-beta-D-glucopyranoside. , 2001, The Plant journal : for cell and molecular biology.

[4]  C. Chapple,et al.  Glucosinolate and phenylpropanoid biosynthesis are linked by proteasome-dependent degradation of PAL , 2019, bioRxiv.

[5]  A. M. Colón,et al.  A kinetic model describes metabolic response to perturbations and distribution of flux control in the benzenoid network of Petunia hybrida. , 2010, The Plant journal : for cell and molecular biology.

[6]  D. Kliebenstein,et al.  Plant Secondary Metabolites as Defenses, Regulators, and Primary Metabolites: The Blurred Functional Trichotomy. , 2020, Plant physiology.

[7]  M. Schalk,et al.  Redirection of cytosolic or plastidic isoprenoid precursors elevates terpene production in plants , 2006, Nature Biotechnology.

[8]  J. Cooney,et al.  Silencing a phloretin‐specific glycosyltransferase perturbs both general phenylpropanoid biosynthesis and plant development , 2017, The Plant journal : for cell and molecular biology.

[9]  Madeleine Ernst,et al.  Comprehensive mass spectrometry-guided phenotyping of plant specialized metabolites reveals metabolic diversity in the cosmopolitan plant family Rhamnaceae. , 2019, The Plant journal : for cell and molecular biology.

[10]  Anthony L. Schilmiller,et al.  Coexpression Analysis Identifies Two Oxidoreductases Involved in the Biosynthesis of the Monoterpene Acid Moiety of Natural Pyrethrin Insecticides in Tanacetum cinerariifolium1[OPEN] , 2017, Plant Physiology.

[11]  D. Lewis,et al.  Members of an R2R3-MYB transcription factor family in Petunia are developmentally and environmentally regulated to control complex floral and vegetative pigmentation patterning. , 2011, The Plant journal : for cell and molecular biology.

[12]  W. Boerjan,et al.  Bioactivity: phenylpropanoids' best kept secret. , 2019, Current opinion in biotechnology.

[13]  Yoshikazu Tanaka,et al.  Biosynthesis of plant pigments: anthocyanins, betalains and carotenoids. , 2008, The Plant journal : for cell and molecular biology.

[14]  M. Haring,et al.  CCoAOMT Down-Regulation Activates Anthocyanin Biosynthesis in Petunia1 , 2015, Plant Physiology.

[15]  J. Noel,et al.  Contribution of isopentenyl phosphate to plant terpenoid metabolism , 2018, Nature Plants.

[16]  W. Kreis,et al.  Exploiting enzyme promiscuity to shape plant specialized metabolism. , 2019, Journal of experimental botany.

[17]  J. Noel,et al.  Coordination of auxin and ethylene biosynthesis by the aminotransferase VAS1 , 2013, Nature chemical biology.

[18]  M. Ram,et al.  Metabolic engineering for the production of plant therapeutic compounds , 2021 .

[19]  E. Hinchliffe,et al.  Improving the nutritional value of Golden Rice through increased pro-vitamin A content , 2005, Nature Biotechnology.

[20]  M. Tegeder,et al.  Manipulation of sucrose phloem and embryo loading affects pea leaf metabolism, carbon and nitrogen partitioning to sinks as well as seed storage pools. , 2019, The Plant journal : for cell and molecular biology.

[21]  J. Weng,et al.  Exploring Uncharted Territories of Plant Specialized Metabolism in the Postgenomic Era. , 2020, Annual review of plant biology.

[22]  J. Keasling,et al.  Remodeling the isoprenoid pathway in tobacco by expressing the cytoplasmic mevalonate pathway in chloroplasts. , 2012, Metabolic engineering.

[23]  D. Tholl.  Biosynthesis and biological functions of terpenoids in plants. , 2015, Advances in biochemical engineering/biotechnology.

[24]  K. Cornish,et al.  Compartmentalized Metabolic Engineering for Artemisinin Biosynthesis and Effective Malaria Treatment by Oral Delivery of Plant Cells. , 2016, Molecular plant.

[25]  Aaron Birchfield,et al.  Metabolic engineering and synthetic biology of plant natural products – A minireview , 2020, Current Plant Biology.

[26]  C. Chapple,et al.  Indole Glucosinolate Biosynthesis Limits Phenylpropanoid Accumulation in Arabidopsis thaliana , 2015, Plant Cell.

[27]  Noa T. Smith,et al.  The production of plant natural products beneficial to humanity by metabolic engineering , 2020 .

[28]  E. Pichersky,et al.  Cytosolic monoterpene biosynthesis is supported by plastid-generated geranyl diphosphate substrate in transgenic tomato fruits. , 2013, The Plant journal : for cell and molecular biology.

[29]  Tariq A. Akhtar,et al.  Multifaceted plant responses to circumvent Phe hyperaccumulation by downregulation of flux through the shikimate pathway and by vacuolar Phe sequestration , 2017, The Plant journal : for cell and molecular biology.

[30]  Thomas Hartmann,et al.  From waste products to ecochemicals: fifty years research of plant secondary metabolism. , 2007, Phytochemistry.

[31]  A. Fernie,et al.  Manipulation of β‐carotene levels in tomato fruits results in increased ABA content and extended shelf life , 2019, Plant biotechnology journal.

[32]  M. Gijzen,et al.  Phytochemical diversity: The sounds of silent metabolism , 2009 .

[33]  Enhanced pyruvate metabolism in plastids by overexpression of putative plastidial pyruvate transporter in Phaeodactylum tricornutum , 2020, Biotechnology for Biofuels.

[34]  B. Spitzer-Rimon,et al.  Generation of the potent anti-malarial drug artemisinin in tobacco , 2011, Nature Biotechnology.

[35]  Jia-bao Huang,et al.  RIBOSE PHOSPHATE ISOMERSASE 1 Influences Root Development by Acting on Cell Wall Biosynthesis, Actin Organization, and Auxin Transport in Arabidopsis , 2020, Frontiers in Plant Science.

[36]  X. Fang,et al.  Characterization of gossypol biosynthetic pathway , 2018, Proceedings of the National Academy of Sciences.

[37]  Joshua S. Yuan,et al.  Terpene metabolic engineering via nuclear or chloroplast genomes profoundly and globally impacts off‐target pathways through metabolite signalling , 2016, Plant biotechnology journal.

[38]  J. Thimmapuram,et al.  Identification of a plastidial phenylalanine exporter that influences flux distribution through the phenylalanine biosynthetic network , 2015, Nature Communications.

[39]  Robert Williams,et al.  Engineering triterpene metabolism in tobacco , 2012, Planta.

[40]  H. Bouwmeester,et al.  Monoterpene biosynthesis potential of plant subcellular compartments. , 2016, The New phytologist.

[41]  C. Chapple,et al.  Linking phenylpropanoid metabolism, lignin deposition, and plant growth inhibition. , 2019, Current opinion in biotechnology.

[42]  P. Verma,et al.  Genetic engineering approach using early Vinca alkaloid biosynthesis genes led to increased tryptamine and terpenoid indole alkaloids biosynthesis in differentiating cultures of Catharanthus roseus , 2017, Protoplasma.

[43]  I. Maoz,et al.  Modulation of auxin formation by the cytosolic phenylalanine biosynthetic pathway , 2020, Nature Chemical Biology.

[44]  J. Noel,et al.  Orthologs of the archaeal isopentenyl phosphate kinase regulate terpenoid production in plants , 2015, Proceedings of the National Academy of Sciences.

[45]  John A. Morgan,et al.  Metabolic flux analysis of secondary metabolism in plants , 2020, Metabolic engineering communications.

[46]  Joshua R. Widhalm,et al.  Biosynthesis and molecular actions of specialized 1,4-naphthoquinone natural products produced by horticultural plants , 2016, Horticulture Research.

[47]  Natalia Dudareva,et al.  The shikimate pathway and aromatic amino Acid biosynthesis in plants. , 2012, Annual review of plant biology.

[48]  P. Beyer,et al.  Engineering the provitamin A (beta-carotene) biosynthetic pathway into (carotenoid-free) rice endosperm. , 2000, Science.

[49]  S. Potlakayala,et al.  Metabolic engineering of chloroplasts for artemisinic acid biosynthesis and impact on plant growth , 2014, Journal of Biosciences.

[50]  Elazar Fallik,et al.  Enrichment of tomato flavor by diversion of the early plastidial terpenoid pathway , 2007, Nature Biotechnology.