4.8 Article

H-lignin can be deposited independently of CINNAMYL ALCOHOL DEHYDROGENASE C and D in Arabidopsis

Journal

PLANT PHYSIOLOGY
Volume 189, Issue 4, Pages 2015-2028

Publisher

OXFORD UNIV PRESS INC
DOI: 10.1093/plphys/kiac210

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Funding

  1. Center for Direct Catalytic Conversion of Biomass to Biofuels, an Energy Frontier Research Center - US Department of Energy, Office of Science, Office of Basic Energy Sciences [DE-SC0000997]
  2. US Department of Energy, Office of Science, Office of Basic Energy Sciences, Chemical Sciences, Geosciences, and Biosciences Division [DE-FG02-07ER15905]
  3. Great Lakes Bioenergy Research Center (DOE BER Office of Science) [DE-SC0018409]

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Lignin is a major obstacle for biomass saccharification, but recent research has found that genetically altering lignin by knocking out two subunits of Mediator can alleviate the growth inhibition phenotype. This alteration leads to the synthesis of a lignin polymer derived from p-coumaryl alcohol.
Lignin contributes substantially to the recalcitrance of biomass toward saccharification. To circumvent this problem, researchers have genetically altered lignin, although, in a number of cases, these efforts have resulted in an undesirable yield penalty. Recent findings have shown that by knocking out two subunits (MED5A and MED5B) of the transcriptional regulatory complex Mediator, the stunted growth phenotype of mutants in p-coumaroyl shikimate 3 '-hydroxylase, reduced epidermal fluorescence 8-1 (ref8-1), can be alleviated. Furthermore, these plants synthesize a lignin polymer almost entirely derived from p-coumaryl alcohol. Plants deficient in cinnamyl alcohol dehydrogenase (CAD) are notable in that they primarily incorporate coniferaldehyde and sinapaldehyde into their lignin. We tested the hypothesis that by stacking mutations in the genes encoding for the CAD paralogs C and D on an Arabidopsis (Arabidopsis thaliana) med5a/5b ref8-1 genetic background, the biosynthesis of p-coumaryl alcohol would be blocked, making p-coumaraldehyde available for polymerization into a novel kind of lignin. The med5a/5b ref8-1 cadc cadd plants are viable, but lignin analysis demonstrated that they continue to synthesize p-hydroxyphenyl lignin despite being mutated for the CADs typically considered to be required for monolignol biosynthesis. In addition, enzyme activity tests showed that even in the absence of CADC and CADD, there is high CAD activity in stems. We tested the potential involvement of other CADs in p-coumaraldehyde biosynthesis in the quintuple mutant by mutating them using the CRISPR/Cas9 system. Lignin analysis demonstrated that the resulting hextuple mutant plants continue to deposit p-coumaryl alcohol-derived lignin, demonstrating a route for the synthesis of p-hydroxyphenyl lignin in Arabidopsis independent of four CAD isoforms. The biosynthesis of p-hydroxyphenyl subunits of the cell wall polymer lignin is independent of the canonical cinnamyl alcohol dehydrogenases involved in guaiacyl and syringyl monomers.

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