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Title: 4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase

Abstract

Lignin biosynthesis is evolutionarily conserved among higher plants and features a critical 3-hydroxylation reaction involving phenolic esters. Yet, increasing evidence questions the involvement of a single pathway to lignin formation in vascular plants. In this work we describe an enzyme catalyzing the direct 3-hydroxylation of 4-coumarate to caffeate in lignin biosynthesis as a bifunctional peroxidase that oxidizes both ascorbate and 4-coumarate at comparable rates. A combination of biochemical and genetic evidence in the model plants Brachypodium distachyon and Arabidopsis thaliana supports a role for this coumarate 3-hydroxylase (C3H) in the early steps of lignin biosynthesis. The subsequent efficient O-methylation of caffeate to ferulate in grasses is substantiated by in vivo biochemical assays. Our results identify C3H as the only non-membrane bound hydroxylase in the lignin pathway and revise the currently accepted models of lignin biosynthesis, suggesting new gene targets to improve forage and bioenergy crops.

Authors:
ORCiD logo [1];  [1];  [2];  [1];  [2]; ORCiD logo [2]; ORCiD logo [3];  [2]; ORCiD logo [3];  [2];  [2]; ORCiD logo [1]
  1. Univ. of North Texas, Denton, TX (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
  2. Univ. of North Texas, Denton, TX (United States)
  3. Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Publication Date:
Research Org.:
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
OSTI Identifier:
1545188
Grant/Contract Number:  
AC05-00OR22725
Resource Type:
Accepted Manuscript
Journal Name:
Nature Communications
Additional Journal Information:
Journal Volume: 10; Journal Issue: 1; Journal ID: ISSN 2041-1723
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Barros, Jaime, Escamilla-Trevino, Luis, Song, Luhua, Rao, Xiaolan, Serrani-Yarce, Juan Carlos, Palacios, Maite Docampo, Engle, Nancy, Choudhury, Feroza K., Tschaplinski, Timothy J., Venables, Barney J., Mittler, Ron, and Dixon, Richard A. 4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase. United States: N. p., 2019. Web. doi:10.1038/s41467-019-10082-7.
Barros, Jaime, Escamilla-Trevino, Luis, Song, Luhua, Rao, Xiaolan, Serrani-Yarce, Juan Carlos, Palacios, Maite Docampo, Engle, Nancy, Choudhury, Feroza K., Tschaplinski, Timothy J., Venables, Barney J., Mittler, Ron, & Dixon, Richard A. 4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase. United States. doi:10.1038/s41467-019-10082-7.
Barros, Jaime, Escamilla-Trevino, Luis, Song, Luhua, Rao, Xiaolan, Serrani-Yarce, Juan Carlos, Palacios, Maite Docampo, Engle, Nancy, Choudhury, Feroza K., Tschaplinski, Timothy J., Venables, Barney J., Mittler, Ron, and Dixon, Richard A. Tue . "4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase". United States. doi:10.1038/s41467-019-10082-7. https://www.osti.gov/servlets/purl/1545188.
@article{osti_1545188,
title = {4-Coumarate 3-hydroxylase in the lignin biosynthesis pathway is a cytosolic ascorbate peroxidase},
author = {Barros, Jaime and Escamilla-Trevino, Luis and Song, Luhua and Rao, Xiaolan and Serrani-Yarce, Juan Carlos and Palacios, Maite Docampo and Engle, Nancy and Choudhury, Feroza K. and Tschaplinski, Timothy J. and Venables, Barney J. and Mittler, Ron and Dixon, Richard A.},
abstractNote = {Lignin biosynthesis is evolutionarily conserved among higher plants and features a critical 3-hydroxylation reaction involving phenolic esters. Yet, increasing evidence questions the involvement of a single pathway to lignin formation in vascular plants. In this work we describe an enzyme catalyzing the direct 3-hydroxylation of 4-coumarate to caffeate in lignin biosynthesis as a bifunctional peroxidase that oxidizes both ascorbate and 4-coumarate at comparable rates. A combination of biochemical and genetic evidence in the model plants Brachypodium distachyon and Arabidopsis thaliana supports a role for this coumarate 3-hydroxylase (C3H) in the early steps of lignin biosynthesis. The subsequent efficient O-methylation of caffeate to ferulate in grasses is substantiated by in vivo biochemical assays. Our results identify C3H as the only non-membrane bound hydroxylase in the lignin pathway and revise the currently accepted models of lignin biosynthesis, suggesting new gene targets to improve forage and bioenergy crops.},
doi = {10.1038/s41467-019-10082-7},
journal = {Nature Communications},
number = 1,
volume = 10,
place = {United States},
year = {2019},
month = {4}
}

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Cited by: 6 works
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Works referenced in this record:

Lignin Valorization: Improving Lignin Processing in the Biorefinery
journal, May 2014

  • Ragauskas, A. J.; Beckham, G. T.; Biddy, M. J.
  • Science, Vol. 344, Issue 6185, p. 1246843-1246843
  • DOI: 10.1126/science.1246843

Cytosolic APx knockdown indicates an ambiguous redox responses in rice
journal, April 2010


Generation and Characterization of the Western Regional Research Center Brachypodium T-DNA Insertional Mutant Collection
journal, September 2012


Arabidopsis GRI is involved in the regulation of cell death induced by extracellular ROS
journal, March 2009

  • Wrzaczek, M.; Brosche, M.; Kollist, H.
  • Proceedings of the National Academy of Sciences, Vol. 106, Issue 13
  • DOI: 10.1073/pnas.0808980106

The origin and evolution of lignin biosynthesis: Tansley review
journal, June 2010


The scaffold proteins of lignin biosynthetic cytochrome P450 enzymes
journal, April 2018


S -Nitrosylation Positively Regulates Ascorbate Peroxidase Activity during Plant Stress Responses
journal, February 2015


Membrane protein complexes catalyze both 4- and 3-hydroxylation of cinnamic acid derivatives in monolignol biosynthesis
journal, December 2011

  • Chen, H. -C.; Li, Q.; Shuford, C. M.
  • Proceedings of the National Academy of Sciences, Vol. 108, Issue 52
  • DOI: 10.1073/pnas.1116416109

Ascorbate Peroxidase 1 Plays a Key Role in the Response of Arabidopsis thaliana to Stress Combination
journal, October 2008

  • Koussevitzky, Shai; Suzuki, Nobuhiro; Huntington, Serena
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Crystal structure of the ascorbate peroxidase–ascorbate complex
journal, March 2003

  • Sharp, Katherine H.; Mewies, Martin; Moody, Peter C. E.
  • Nature Structural & Molecular Biology, Vol. 10, Issue 4
  • DOI: 10.1038/nsb913

Caffeoyl Shikimate Esterase (CSE) Is an Enzyme in the Lignin Biosynthetic Pathway in Arabidopsis
journal, August 2013


Heterologous Expression and Characterization of Soybean Cytosolic Ascorbate Peroxidase
journal, April 1996

  • Dalton, David A.; del Castillo, Leonor Diaz; Kahn, Michael L.
  • Archives of Biochemistry and Biophysics, Vol. 328, Issue 1
  • DOI: 10.1006/abbi.1996.0135

Stress-Induced Phenylpropanoid Metabolism.
journal, July 1995


Downregulation of p‐ COUMAROYL ESTER 3‐ HYDROXYLASE in rice leads to altered cell wall structures and improves biomass saccharification
journal, July 2018

  • Takeda, Yuri; Tobimatsu, Yuki; Karlen, Steven D.
  • The Plant Journal, Vol. 95, Issue 5
  • DOI: 10.1111/tpj.13988

Dynamic changes in transcriptome and cell wall composition underlying brassinosteroid-mediated lignification of switchgrass suspension cells
journal, November 2017


Low frequency of T-DNA based activation tagging in Arabidopsis is correlated with methylation of CaMV 35S enhancer sequences
journal, November 2003


Purification and Characterization of Pea Cytosolic Ascorbate Peroxidase
journal, November 1991

  • Mittler, Ron; Zilinskas, Barbara A.
  • Plant Physiology, Vol. 97, Issue 3
  • DOI: 10.1104/pp.97.3.962

Reactive oxygen gene network of plants
journal, October 2004


Synthesis of Cinnamic Acid Derivatives Using Ethanol as Solvent or Microwave Assisted Method.
journal, October 2000

  • Pellón, Rolando F.; Mamposo, Taimirys; González, Erick
  • Synthetic Communications, Vol. 30, Issue 20
  • DOI: 10.1080/00397910008087005

Substrate Binding and Catalytic Mechanism in Ascorbate Peroxidase:  Evidence for Two Ascorbate Binding Sites
journal, November 2002

  • Lad, Latesh; Mewies, Martin; Raven, Emma Lloyd
  • Biochemistry, Vol. 41, Issue 46
  • DOI: 10.1021/bi0261591

Cinnamoyl-CoA reductase, a key enzyme in lignin biosynthesis, is an effector of small GTPase Rac in defense signaling in rice
journal, December 2005

  • Kawasaki, T.; Koita, H.; Nakatsubo, T.
  • Proceedings of the National Academy of Sciences, Vol. 103, Issue 1, p. 230-235
  • DOI: 10.1073/pnas.0509875103

Abolishing activity against ascorbate in a cytosolic ascorbate peroxidase from switchgrass
journal, October 2013


CYP98A3 from Arabidopsis thaliana Is a 3′-Hydroxylase of Phenolic Esters, a Missing Link in the Phenylpropanoid Pathway
journal, June 2001

  • Schoch, Guillaume; Goepfert, Simon; Morant, Marc
  • Journal of Biological Chemistry, Vol. 276, Issue 39
  • DOI: 10.1074/jbc.M104047200

4-Hydroxycinnamic Acid Hydroxylase and Polyphenolase Activities in Sorghum vulgare
journal, April 1972


A 13C isotope labeling method for the measurement of lignin metabolic flux in Arabidopsis stems
journal, June 2018


Expression atlas and comparative coexpression network analyses reveal important genes involved in the formation of lignified cell wall in Brachypodium distachyon
journal, June 2017

  • Sibout, Richard; Proost, Sebastian; Hansen, Bjoern Oest
  • New Phytologist, Vol. 215, Issue 3
  • DOI: 10.1111/nph.14635

Detection and Characterization of p-Coumaric Acid Hydroxylase in Mung Bean, Vigna mungo, Seedlings1
journal, February 1989


An essential role of caffeoyl shikimate esterase in monolignol biosynthesis in Medicago truncatula
journal, June 2016

  • Ha, Chan Man; Escamilla-Trevino, Luis; Yarce, Juan Carlos Serrani
  • The Plant Journal, Vol. 86, Issue 5
  • DOI: 10.1111/tpj.13177

A Genomics Approach to Deciphering Lignin Biosynthesis in Switchgrass
journal, November 2013


Lack of involvement of polyphenol oxidase in ortho-hydroxylation of phenolic compounds in mung bean seedlings
journal, April 1982


Identification of Two Electron-Transfer Sites in Ascorbate Peroxidase Using Chemical Modification, Enzyme Kinetics, and Crystallography
journal, December 1998

  • Mandelman, David; Jamal, Joumana; Poulos, Thomas L.
  • Biochemistry, Vol. 37, Issue 50
  • DOI: 10.1021/bi981958y

Cytosolic Ascorbate Peroxidase 1 Is a Central Component of the Reactive Oxygen Gene Network of Arabidopsis
journal, December 2004

  • Davletova, Sholpan; Rizhsky, Ludmila; Liang, Hongjian
  • The Plant Cell, Vol. 17, Issue 1
  • DOI: 10.1105/tpc.104.026971

Down-regulation of the caffeic acid O-methyltransferase gene in switchgrass reveals a novel monolignol analog
journal, January 2012

  • Tschaplinski, Timothy J.; Standaert, Robert F.; Engle, Nancy L.
  • Biotechnology for Biofuels, Vol. 5, Issue 1
  • DOI: 10.1186/1754-6834-5-71

Rapid Accumulation of Glutathione During Light Stress in Arabidopsis
journal, May 2018

  • Choudhury, Feroza K.; Devireddy, Amith R.; Azad, Rajeev K.
  • Plant and Cell Physiology, Vol. 59, Issue 9
  • DOI: 10.1093/pcp/pcy101

    Works referencing / citing this record:

    Heterologous Expression and Characterization of Soybean Cytosolic Ascorbate Peroxidase
    journal, April 1996

    • Dalton, David A.; del Castillo, Leonor Diaz; Kahn, Michael L.
    • Archives of Biochemistry and Biophysics, Vol. 328, Issue 1
    • DOI: 10.1006/abbi.1996.0135

    Cytosolic APx knockdown indicates an ambiguous redox responses in rice
    journal, April 2010


    Abolishing activity against ascorbate in a cytosolic ascorbate peroxidase from switchgrass
    journal, October 2013


    Reactive oxygen gene network of plants
    journal, October 2004


    Substrate Binding and Catalytic Mechanism in Ascorbate Peroxidase:  Evidence for Two Ascorbate Binding Sites
    journal, November 2002

    • Lad, Latesh; Mewies, Martin; Raven, Emma Lloyd
    • Biochemistry, Vol. 41, Issue 46
    • DOI: 10.1021/bi0261591

    Identification of Two Electron-Transfer Sites in Ascorbate Peroxidase Using Chemical Modification, Enzyme Kinetics, and Crystallography
    journal, December 1998

    • Mandelman, David; Jamal, Joumana; Poulos, Thomas L.
    • Biochemistry, Vol. 37, Issue 50
    • DOI: 10.1021/bi981958y

    Crystal structure of the ascorbate peroxidase–ascorbate complex
    journal, March 2003

    • Sharp, Katherine H.; Mewies, Martin; Moody, Peter C. E.
    • Nature Structural & Molecular Biology, Vol. 10, Issue 4
    • DOI: 10.1038/nsb913

    The scaffold proteins of lignin biosynthetic cytochrome P450 enzymes
    journal, April 2018


    Cinnamoyl-CoA reductase, a key enzyme in lignin biosynthesis, is an effector of small GTPase Rac in defense signaling in rice
    journal, December 2005

    • Kawasaki, T.; Koita, H.; Nakatsubo, T.
    • Proceedings of the National Academy of Sciences, Vol. 103, Issue 1, p. 230-235
    • DOI: 10.1073/pnas.0509875103

    Arabidopsis GRI is involved in the regulation of cell death induced by extracellular ROS
    journal, March 2009

    • Wrzaczek, M.; Brosche, M.; Kollist, H.
    • Proceedings of the National Academy of Sciences, Vol. 106, Issue 13
    • DOI: 10.1073/pnas.0808980106

    Membrane protein complexes catalyze both 4- and 3-hydroxylation of cinnamic acid derivatives in monolignol biosynthesis
    journal, December 2011

    • Chen, H. -C.; Li, Q.; Shuford, C. M.
    • Proceedings of the National Academy of Sciences, Vol. 108, Issue 52
    • DOI: 10.1073/pnas.1116416109

    Synthesis of Cinnamic Acid Derivatives Using Ethanol as Solvent or Microwave Assisted Method.
    journal, October 2000

    • Pellón, Rolando F.; Mamposo, Taimirys; González, Erick
    • Synthetic Communications, Vol. 30, Issue 20
    • DOI: 10.1080/00397910008087005

    Detection and Characterization of p-Coumaric Acid Hydroxylase in Mung Bean, Vigna mungo, Seedlings1
    journal, February 1989


    Rapid Accumulation of Glutathione During Light Stress in Arabidopsis
    journal, May 2018

    • Choudhury, Feroza K.; Devireddy, Amith R.; Azad, Rajeev K.
    • Plant and Cell Physiology, Vol. 59, Issue 9
    • DOI: 10.1093/pcp/pcy101

    S -Nitrosylation Positively Regulates Ascorbate Peroxidase Activity during Plant Stress Responses
    journal, February 2015


    4-Hydroxycinnamic Acid Hydroxylase and Polyphenolase Activities in Sorghum vulgare
    journal, April 1972


    Purification and Characterization of Pea Cytosolic Ascorbate Peroxidase
    journal, November 1991

    • Mittler, Ron; Zilinskas, Barbara A.
    • Plant Physiology, Vol. 97, Issue 3
    • DOI: 10.1104/pp.97.3.962

    Cytosolic Ascorbate Peroxidase 1 Is a Central Component of the Reactive Oxygen Gene Network of Arabidopsis
    journal, December 2004

    • Davletova, Sholpan; Rizhsky, Ludmila; Liang, Hongjian
    • The Plant Cell, Vol. 17, Issue 1
    • DOI: 10.1105/tpc.104.026971

    A Genomics Approach to Deciphering Lignin Biosynthesis in Switchgrass
    journal, November 2013


    Stress-Induced Phenylpropanoid Metabolism.
    journal, July 1995


    Lack of involvement of polyphenol oxidase in ortho-hydroxylation of phenolic compounds in mung bean seedlings
    journal, April 1982


    The origin and evolution of lignin biosynthesis: Tansley review
    journal, June 2010


    Expression atlas and comparative coexpression network analyses reveal important genes involved in the formation of lignified cell wall in Brachypodium distachyon
    journal, June 2017

    • Sibout, Richard; Proost, Sebastian; Hansen, Bjoern Oest
    • New Phytologist, Vol. 215, Issue 3
    • DOI: 10.1111/nph.14635

    An essential role of caffeoyl shikimate esterase in monolignol biosynthesis in Medicago truncatula
    journal, June 2016

    • Ha, Chan Man; Escamilla-Trevino, Luis; Yarce, Juan Carlos Serrani
    • The Plant Journal, Vol. 86, Issue 5
    • DOI: 10.1111/tpj.13177

    Downregulation of p‐ COUMAROYL ESTER 3‐ HYDROXYLASE in rice leads to altered cell wall structures and improves biomass saccharification
    journal, July 2018

    • Takeda, Yuri; Tobimatsu, Yuki; Karlen, Steven D.
    • The Plant Journal, Vol. 95, Issue 5
    • DOI: 10.1111/tpj.13988

    Caffeoyl Shikimate Esterase (CSE) Is an Enzyme in the Lignin Biosynthetic Pathway in Arabidopsis
    journal, August 2013


    Lignin Valorization: Improving Lignin Processing in the Biorefinery
    journal, May 2014

    • Ragauskas, A. J.; Beckham, G. T.; Biddy, M. J.
    • Science, Vol. 344, Issue 6185, p. 1246843-1246843
    • DOI: 10.1126/science.1246843

    Down-regulation of the caffeic acid O-methyltransferase gene in switchgrass reveals a novel monolignol analog
    journal, January 2012

    • Tschaplinski, Timothy J.; Standaert, Robert F.; Engle, Nancy L.
    • Biotechnology for Biofuels, Vol. 5, Issue 1
    • DOI: 10.1186/1754-6834-5-71

    A 13C isotope labeling method for the measurement of lignin metabolic flux in Arabidopsis stems
    journal, June 2018


    Dynamic changes in transcriptome and cell wall composition underlying brassinosteroid-mediated lignification of switchgrass suspension cells
    journal, November 2017


    Generation and Characterization of the Western Regional Research Center Brachypodium T-DNA Insertional Mutant Collection
    journal, September 2012


    The lignin toolbox of the model grass Setaria viridis
    journal, June 2019

    • Ferreira, Sávio Siqueira; Simões, Marcella Siqueira; Carvalho, Gabriel Garon
    • Plant Molecular Biology, Vol. 101, Issue 3
    • DOI: 10.1007/s11103-019-00897-9

    PdWND3A, a wood-associated NAC domain-containing protein, affects lignin biosynthesis and composition in Populus
    journal, November 2019