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. However, increasing evidence questions the involvement of a single pathway to lignin formation in vascular plants. Here 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.

Phenylalanine and Tyrosine as Exogenous Precursors of Wheat (Triticum aestivum L.) Secondary Metabolism through PAL-Associated Pathways

“…P-coumarate 3-hydroxylase encoded by the C3H gene catalyzes the direct 3-hydroxylation of 4-coumarate to caffeate in lignin biosynthesis. In fact, the activity of p-coumarate 3-hydroxylase causes the transition from the pathway of biosynthesis of P-lignin units to the biosynthesis of all other lignin units: C, G, SH, S -lignin monomers [39,40]. C3H1 gene was expressed both in the tyrosine-supplemented and phenylalanine-supplemented nutrient media.…”

Compensatory guaiacyl lignin biosynthesis at the expense of syringyl lignin in4CL1-knockout poplar

“…This suggests involvement of CSE1 and CCoAOMT1 in homeostatic regulation of caffeic acid and caffeoyl-CoA in response to 4CL1 -KO. Recently, a cytosolic ascorbate peroxidase (APX) was shown to possess 4-coumarate 3-hydroxylase (C3H) activity and convert 4-coumaric acid to caffeic acid in both monocots and dicots, supporting an alternative route in lignin biosynthesis involving free phenolic acids (Barros et al, 2019). Two orthologs of this dual-function APX-C3H are present in the poplar genome, and one of them (Potri.009G015400, APX-C3H1) was significantly upregulated in the mutants (Figure 4A).…”

“…Plasticity in the lignin biosynthetic pathway as evidenced by multiple possible routes (Barros et al, 2019) (Figure 6) may accommodate developmental, environmental, or species-dependent variations in lignification. 4CLs, for instance, can utilize multiple hydroxycinnamate substrates in vitro , but how substrate utilization is regulated in vivo is largely unknown.…”

“…Caffeic acid is among the most potent antioxidants of the plant-derived phenolics, with radical scavenging and metal chelation activities to avert potential damaging effects of reactive oxygen species (Chen and Ho, 1997; Perron and Brumaghim, 2009). Its hyper-accumulation in 4cl1 mutants coincided with upregulation of not only APX-C3H1 and CSE1 , but also GSTs, PRDs, CSDs, MSRs , and genes encoding enzymes of the glutathione-ascorbate cycle, all of which have been implicated in detoxification of hydrogen peroxide and superoxide (Rouhier et al, 2008; Gao et al, 2010; Molina-Rueda et al, 2013; Rey and Tarrago, 2018; Barros et al, 2019). Coordinated regulation of multiple antioxidant systems may be crucial to maintain redox balance under the presumably highly oxidative cellular environment resulting from disturbed lignin biosynthesis.…”

Elucidating The Multifunctional Role of The Cell Wall Components in The Maize Exploitation

“…Total lignin content was determined by Klason Lignin protocol (33). Monomer composition was determined by thioacidolysis followed by Gas chromatography-mass spectrometry (GC-MS) (34). The thioacidolysis procedure estimates the proportion of lignin subunits linked by the major β-O-4 linkages.…”