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Introducing curcumin biosynthesis in Arabidopsis enhances lignocellulosic biomass processing

Nature Plants volume 5, pages 225–237 (2019)Cite this article

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Abstract

Lignin is the main cause of lignocellulosic biomass recalcitrance to industrial enzymatic hydrolysis. By partially replacing the traditional lignin monomers by alternative ones, lignin extractability can be enhanced. To design a lignin that is easier to degrade under alkaline conditions, curcumin (diferuloylmethane) was produced in the model plant Arabidopsis thaliana via simultaneous expression of the turmeric (Curcuma longa) genes DIKETIDE-CoA SYNTHASE (DCS) and CURCUMIN SYNTHASE 2 (CURS2). The transgenic plants produced a plethora of curcumin- and phenylpentanoid-derived compounds with no negative impact on growth. Catalytic hydrogenolysis gave evidence that both curcumin and phenylpentanoids were incorporated into the lignifying cell wall, thereby significantly increasing saccharification efficiency after alkaline pretreatment of the transgenic lines by 14–24% as compared with the wild type. These results demonstrate that non-native monomers can be synthesized and incorporated into the lignin polymer in plants to enhance their biomass processing efficiency.

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Fig. 1: Curcumin biosynthesis and incorporation into the lignin polymer.
Fig. 2: Curcumin couples with traditional monolignols in in vitro coupling assays.
Fig. 3: Structure of curcuminoid- and phenylpentanoid-containing metabolites detected in pCesA4:DCS_CURS2 lines.
Fig. 4: Bright-field and fluorescence microscopy of WT and transgenic transverse inflorescence stem cross-sections and curcumin breakdown in alkaline conditions.
Fig. 5: Cellulose-to-glucose conversion during saccharification of inflorescence stems of the WT and pCesA4:DCS_CURS2 lines.

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The authors declare that all data supporting the findings of this study are available within the paper and its Supplementary Information files.

References

  1. Boerjan, W., Ralph, J. & Baucher, M. Lignin biosynthesis. Annu. Rev. Plant Biol. 54, 519–546 (2003).

    Article  CAS  PubMed  Google Scholar 

  2. Vanholme, R. et al. Metabolic engineering of novel lignin in biomass crops. New Phytol. 196, 978–1000 (2012).

    Article  CAS  PubMed  Google Scholar 

  3. Mottiar, Y., Vanholme, R., Boerjan, W., Ralph, J. & Mansfield, S. D. Designer lignins: harnessing the plasticity of lignification. Curr. Opin. Biotechnol. 37, 190–200 (2016).

    Article  CAS  PubMed  Google Scholar 

  4. Ralph, J. et al. Lignins: natural polymers from oxidative coupling of 4-hydroxyphenylpropanoids. Phytochem. Rev. 3, 29–60 (2004).

    Article  CAS  Google Scholar 

  5. Eudes, A. et al. Biosynthesis and incorporation of side-chain-truncated lignin monomers to reduce lignin polymerization and enhance saccharification. Plant Biotechnol. J. 10, 609–620 (2012).

    Article  CAS  PubMed  Google Scholar 

  6. Kim, K. H. et al. Impact of lignin polymer backbone esters on ionic liquid pretreatment of poplar. Biotechnol. Biofuels 10, 101 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Wilkerson, C. G. et al. Monolignol ferulate transferase introduces chemically labile linkages into the lignin backbone. Science 344, 90–93 (2014).

    Article  CAS  PubMed  Google Scholar 

  8. Zhou, S. et al. Chemical pulping advantages of zip-lignin hybrid poplar. ChemSusChem 10, 3565–3573 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Bhalla, A. et al. Engineered lignin in poplar biomass facilitates Cu-catalyzed alkaline-oxidative pretreatment. ACS Sustain Chem. Eng. 6, 2932–2941 (2018).

    Article  CAS  Google Scholar 

  10. Smith, R. A. et al. Engineering monolignol p-coumarate conjugates into poplar and Arabidopsis lignins. Plant Physiol. 169, 2992–3001 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  11. Sibout, R. et al. Structural redesigning Arabidopsis lignins into alkali-soluble lignins through the expression of p-coumaroyl-CoA:monolignol transferase PMT. Plant Physiol. 170, 1358–1366 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Ragauskas, A. J. et al. Lignin valorization: improving lignin processing in the biorefinery. Science 344, 1246843 (2014).

    Article  CAS  PubMed  Google Scholar 

  13. Chen, Y.-R. & Tan, T.-H. Inhibition of the c-Jun N-terminal kinase (JNK) signaling pathway by curcumin. Oncogene 17, 173–178 (1998).

    Article  CAS  PubMed  Google Scholar 

  14. Katsuyama, Y., Kita, T., Funa, N. & Horinouchi, S. Curcuminoid biosynthesis by two type III polyketide synthases in the herb Curcuma longa. J. Biol. Chem. 284, 11160–11170 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Hauser, C. R., Swamer, F. W. & Ringler, B. I. Alkaline cleavage of unsymmetrical β-diketones. Ring opening of acylcylohexanones to form ε-acyl caproic acids. J. Am. Chem. Soc. 70, 4023–4026 (1948).

    Article  CAS  PubMed  Google Scholar 

  16. Wang, Y.-J. et al. Stability of curcumin in buffer solutions and characterization of its degradation products. J. Pharm. Biomed. Anal. 15, 1867–1876 (1997).

    Article  CAS  PubMed  Google Scholar 

  17. Tomren, M. A., Másson, M., Loftsson, T. & Tønnesen, H. H. Studies on curcumin and curcuminoids: XXXI. Symmetric and asymmetric curcuminoids: stability, activity and complexation with cyclodextrin. Int. J. Pharm. 338, 27–34 (2007).

    Article  CAS  PubMed  Google Scholar 

  18. Price, L. C. & Buescher, R. W. Kinetics of alkaline degradation of the food pigments curcumin and curcuminoids. J. Food. Sci. 62, 267–269 (1997).

    Article  CAS  Google Scholar 

  19. Pearson, R. G. & Mayerle, E. A. Mechanism of the hydrolytic cleavage of carbon—carbon bonds. I. Alkaline hydrolysis of β-diketones. J. Am. Chem. Soc. 73, 926–930 (1951).

    Article  CAS  Google Scholar 

  20. Tønnesen, H. H. & Karlsen, J. Studies on curcumin and curcuminoids. V. Alkaline degradation of curcumin. Z. Lebens. Unters. Forsch. 180, 132–134 (1985).

    Article  Google Scholar 

  21. Tsuji, Y. et al. Introduction of chemically labile substructures into Arabidopsis lignin through the use of LigD, the Cα-dehydrogenase from Sphingobium sp. strain SYK-6. Plant Biotechnol. J. 13, 821–832 (2015).

    Article  CAS  PubMed  Google Scholar 

  22. Criss, D. L., Fisher, T. H. & Schultz, T. P. Alkaline hydrolysis of nonphenolic α-carbonyl β-O-4 lignin dimers substituted on the leaving phenoxide ring: comparison with benzylic hydroxyl analogues. Holzforschung 52, 57–60 (1998).

    Article  CAS  Google Scholar 

  23. Mnich, E. et al. Degradation of lignin β‐aryl ether units in Arabidopsis thaliana expressing LigD, LigF and LigG from Sphingomonas paucimobilis SYK‐6. Plant Biotechnol. J. 15, 581–593 (2017).

    Article  CAS  PubMed  Google Scholar 

  24. Imai, A., Yokoyama, T., Matsumoto, Y. & Meshitsukat, G. Significant lability of guaiacylglycerol β-phenacyl ether under alkaline conditions. J. Agric. Food Chem. 55, 9043–9046 (2007).

    Article  CAS  PubMed  Google Scholar 

  25. Freudenberg, K. & Neish, A. C. Constitution and Biosynthesis of Lignin (Springer-Verlag, New York, 1968).

  26. Tobimatsu, Y. et al. Hydroxycinnamate conjugates as potential monolignol replacements: in vitro lignification and cell wall studies with rosmarinic acid. ChemSusChem 5, 676–686 (2012).

    Article  CAS  PubMed  Google Scholar 

  27. Lan, W. et al. Tricin, a flavonoid monomer in monocot lignification. Plant Physiol. 167, 1284–1295 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Morreel, K. et al. Mass spectrometry-based sequencing of lignin oligomers. Plant Physiol. 153, 1464–1478 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Morreel, K. et al. Mass spectrometry-based fragmentation as an identification tool in lignomics. Anal. Chem. 82, 8095–8105 (2010).

    Article  CAS  PubMed  Google Scholar 

  30. Tobimatsu, Y. et al. Visualization of plant cell wall lignification using fluorescence‐tagged monolignols. Plant J. 76, 357–366 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Tobimatsu, Y. et al. A click chemistry strategy for visualization of plant cell wall lignification. Chem. Commun. 50, 12262–12265 (2014).

    Article  CAS  Google Scholar 

  32. Singh, R., Tonnesen, H. H., Vogensen, S. B., Loftsson, T. & Masson, M. Studies of curcumin and curcuminoids. XXXVI. The stoichiometry and complexation constants of cyclodextrin complexes as determined by the phase-solubility method and UV-Vis titration. J. Incl. Phenom. Macro. 66, 335–348 (2010).

    Article  CAS  Google Scholar 

  33. Katsuyama, Y., Kita, T. & Horinouchi, S. Identification and characterization of multiple curcumin synthases from the herb Curcuma longa. FEBS Lett. 583, 2799–2803 (2009).

    Article  CAS  PubMed  Google Scholar 

  34. Halpin, C., Cooke, S. E., Barakate, A., Amrani, A. E. & Ryan, M. D. Self‐processing 2A‐polyproteins – a system for co‐ordinate expression of multiple proteins in transgenic plants. Plant J. 17, 453–459 (1999).

    Article  CAS  PubMed  Google Scholar 

  35. Morreel, K. et al. Profiling of oligolignols reveals monolignol coupling conditions in lignifying poplar xylem. Plant Physiol. 136, 3537–3549 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. De Meester, B. et al. Vessel-specific reintroduction of CINNAMOYL-COA REDUCTASE1 (CCR1) in dwarfedccr1 mutants restores vessel and xylary fiber integrity and increases biomass. Plant Physiol. 176, 611–633 (2018).

    Article  CAS  PubMed  Google Scholar 

  37. Vanholme, R. et al. A systems biology view of responses to lignin biosynthesis perturbations in Arabidopsis. Plant Cell 24, 3506–3529 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Matsuda, F. et al. AtMetExpress development: a phytochemical atlas of Arabidopsis development. Plant Physiol. 152, 566–578 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Morreel, K. et al. Systematic structural characterization of metabolites in Arabidopsis via candidate substrate–product pair networks. Plant Cell 26, 929–945 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Morrison, W. H., Hartley, R. D. & Himmelsbach, D. S. Synthesis of substituted truxillic acids from p-coumaric and ferulic acid: simulation of photodimerization in plant cell walls. J. Agric. Food Chem. 40, 768–771 (1992).

    Article  CAS  Google Scholar 

  41. D’Auria, M. & Vantaggi, A. Photochemical dimerization of methoxy substituted cinnamic acid methyl esters. Tetrahedron 48, 2523–2528 (1992).

    Article  Google Scholar 

  42. Hanley, A. B., Russell, W. R. & Chesson, A. Formation of substituted truxillic and truxinic acids in plant cell walls – a rationale. Phytochemistry 33, 957–960 (1993).

    Article  CAS  Google Scholar 

  43. Dauwe, R. et al. Molecular phenotyping of lignin-modified tobacco reveals associated changes in cell-wall metabolism, primary metabolism, stress metabolism and photorespiration. Plant J. 52, 263–285 (2007).

    Article  CAS  PubMed  Google Scholar 

  44. Bonawitz, N. D. et al. Disruption of Mediator rescues the stunted growth of a lignin-deficient Arabidopsis mutant. Nature 509, 376–380 (2014).

    Article  CAS  PubMed  Google Scholar 

  45. Van den Bosch, S. et al. Reductive lignocellulose fractionation into soluble lignin-derived phenolic monomers and dimers and processable carbohydrate pulps. Energy Environ. Sci. 8, 1748–1763 (2015).

    Article  CAS  Google Scholar 

  46. Galkin, M. V. & Samec, J. S. M. Selective route to 2-propenyl aryls directly from wood by a tandem organosolv and palladium-catalysed transfer hydrogenolysis. ChemSusChem 7, 2154–2158 (2014).

    Article  CAS  PubMed  Google Scholar 

  47. Galkin, M. V. & Samec, J. S. M. Lignin valorization through catalytic lignocellulose fractionation: a fundamental platform for the future biorefinery. ChemSusChem 9, 1544–1558 (2016).

    Article  CAS  PubMed  Google Scholar 

  48. Renders, T. et al. Synergetic effects of alcohol/water mixing on the catalytic reductive fractionation of poplar wood. ACS Sustain. Chem. Eng. 4, 6894–6904 (2016).

    Article  CAS  Google Scholar 

  49. Hendrickson, J. B., Cram, D. J. & Hammond, G. S. Organic Chemistry 3rd edn (McGraw-Hill, Tokyo, 1970).

  50. Rahimi, A., Ulbrich, A., Coon, J. J. & Stahl, S. S. Formic-acid-induced depolymerization of oxidized lignin to aromatics. Nature 515, 249–252 (2014).

    Article  CAS  PubMed  Google Scholar 

  51. Van Acker, R. et al. Lignin biosynthesis perturbations affect secondary cell wall composition and saccharification yield in Arabidopsis thaliana. Biotechnol. Biofuels 6, 46 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Lee, S., Mo, H., Kim, J. I. & Chapple, C. Genetic engineering of Arabidopsis to overproduce disinapoyl esters, potential lignin modification molecules. Biotechnol. Biofuels 10, 40 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Ralph, J. et al. Identification of the structure and origin of a thioacidolysis marker compound for ferulic acid incorporation into angiosperm lignins (and an indicator for cinnamoyl-CoA reductase deficiency). Plant J. 53, 368–379 (2008).

    Article  CAS  PubMed  Google Scholar 

  54. Sibout, R. & Höfte, H. Plant cell biology: the ABC of monolignol transport. Curr. Biol. 22, R533–R535 (2012).

    Article  CAS  PubMed  Google Scholar 

  55. Karlen, S. D. et al. Monolignol ferulate conjugates are naturally incorporated into plant lignins. Sci. Adv. 2, e1600393 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Leplé, J.-C. et al. Downregulation of cinnamoyl-coenzyme A reductase in poplar: multiple-level phenotyping reveals effects on cell wall polymer metabolism and structure. Plant Cell 19, 3669–3691 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Van Acker, R. et al. Improved saccharification and ethanol yield from field-grown transgenic poplar deficient in cinnamoyl-CoA reductase. Proc. Natl Acad. Sci. USA 111, 845–850 (2014).

    Article  CAS  PubMed  Google Scholar 

  58. Vanholme, R. et al. Caffeoyl shikimate esterase (CSE) is an enzyme in the lignin biosynthetic pathway in Arabidopsis. Science 341, 1103–1106 (2013).

    Article  CAS  PubMed  Google Scholar 

  59. Clough, S. J. & Bent, A. F. Floral dip: a simplified method for Agrobacterium-mediated transformation of Arabidopsis thaliana. Plant J. 16, 735–743 (1998).

    Article  CAS  PubMed  Google Scholar 

  60. Shimada, T. L., Shimada, T. & Hara‐Nishimura, I. A rapid and non‐destructive screenable marker, FAST, for identifying transformed seeds of Arabidopsis thaliana. Plant J. 61, 519–528 (2010).

    Article  CAS  PubMed  Google Scholar 

  61. Sundin, L. et al. Mutation of the inducible ARABIDOPSIS THALIANA CYTOCHROME P450 REDUCTASE2 alters lignin composition and improves saccharification. Plant Physiol. 166, 1956–1971 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Foster, C. E., Martin, T. M. & Pauly, M. Comprehensive compositional analysis of plant cell walls (lignocellulosic biomass). Part I: Lignin. J. Vis. Exp 37, e1745 (2010).

    Google Scholar 

  63. Van Acker, R. et al. Different routes for conifer- and sinapaldehyde and higher saccharification upon deficiency in the dehydrogenase CAD1. Plant Physiol. 175, 1018–1039 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Brendel, O., P.P.M., I. & Stewart, D. A rapid and simple method to isolate pure alpha-cellulose. Phytochem. Anal. 11, 4 (2000).

    Article  Google Scholar 

  65. Van Acker, R., Vanholme, R., Piens, K. & Boerjan, W. Saccharification protocol for small-scale lignocellulosic biomass samples to test processing of cellulose into glucose. Bio-Protocol 6, e1701 (2016).

    Article  Google Scholar 

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Acknowledgements

P.O. was funded by the National Commission for Scientific and Technological Research (Chile) for a predoctoral fellowship and by SBO-FISH through the ARBOREF project for a postdoctoral fellowship, B.D.M. and L.d.V. were funded by the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Vlaanderen) for predoctoral fellowships, F.F. was funded by the Science Without Borders program from CNPq for a postdoctoral fellowship 206329/2014–8, A.P. was funded by the Research Foundation-Flanders (FWO, grant G0C1914N) and Y.T. was funded by Stanford University’s Global Climate and Energy Program (GCEP). S.V.d.B. was funded by PDM KULeuven and by BIOWOOD (FWO-SBO), J.R. was funded by the DOE Great Lakes Bioenergy Research Center (DOE BER Office of Science DE-FC02-07ER64494 and DE-SC0018409) and R.V. was funded by the FWO for a postdoctoral fellowship. The authors thank A. Bleys for help in preparing the manuscript, E. Parthoens (VIB BioImaging Core) for assitance with the fluorescence microscopy and D. Loqué for providing the vector containing pCesA4. The work was also funded by a CLEM grant from I. Lieten to the VIB BioImaging Core for the acquisition of a Zeiss LSM780 microscope.

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Author notes

  1. These authors contributed equally: Paula Oyarce, Barbara De Meester.

  2. These authors jointly supervised this work: Ruben Vanholme, Wout Boerjan.

Authors and Affiliations

  1. Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium

    Paula Oyarce, Barbara De Meester, Fernando Fonseca, Lisanne de Vries, Geert Goeminne, Andreas Pallidis, Riet De Rycke, Ruben Vanholme & Wout Boerjan

  2. VIB Center for Plant Systems Biology, Ghent, Belgium

    Paula Oyarce, Barbara De Meester, Fernando Fonseca, Lisanne de Vries, Geert Goeminne, Andreas Pallidis, Riet De Rycke, Ruben Vanholme & Wout Boerjan

  3. VIB Metabolomics Core, Ghent, Belgium

    Geert Goeminne, Riet De Rycke, Ruben Vanholme & Wout Boerjan

  4. Ghent University Expertise Centre for Transmission Electron Microscopy and VIB BioImaging Core, Ghent, Belgium

    Riet De Rycke

  5. Department of Biochemistry, University of Wisconsin, Madison, WI, USA

    Yukiko Tsuji, Yanding Li & John Ralph

  6. US Department of Energy, Great Lakes Bioenergy Research Center, Wisconsin Energy Institute, Madison, WI, USA

    Yukiko Tsuji, Yanding Li & John Ralph

  7. Center for Surface Chemistry and Catalysis, KU Leuven, Heverlee, Belgium

    Sander Van den Bosch & Bert Sels

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Contributions

P.O., B.D.M., J.R., R.V. and W.B. designed the experiments. P.O., B.D.M., F.F., L.d.V., G.G., R.D.R., Y.T., Y.L. and S.V.d.B. performed the experiments. P.O., B.D.M., G.G., A.P., Y.T., B.S., J.R. and R.V. collected and analysed data. P.O., B.D.M., R.V. and W.B. wrote the article with contributions from all authors.

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Correspondence to Wout Boerjan.

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Oyarce, P., De Meester, B., Fonseca, F. et al. Introducing curcumin biosynthesis in Arabidopsis enhances lignocellulosic biomass processing. Nature Plants 5, 225–237 (2019). https://doi.org/10.1038/s41477-018-0350-3

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