Lignin Engineering in Forest Trees

Abstract

Wood is a renewable resource that is mainly composed of lignin and cell wall polysaccharides. The polysaccharide fraction is valuable as it can be converted into pulp and paper, or into fermentable sugars. On the other hand, the lignin fraction is increasingly being considered a valuable source of aromatic building blocks for the chemical industry. The presence of lignin in wood is one of the major recalcitrance factors in woody biomass processing, necessitating the need for harsh chemical treatments to degrade and extract it prior to the valorization of the cell wall polysaccharides, cellulose and hemicellulose. Over the past years, large research efforts have been devoted to engineering lignin amount and composition to reduce biomass recalcitrance toward chemical processing. We review the efforts made in forest trees, and compare results from greenhouse and field trials. Furthermore, we address the value and potential of CRISPR-based gene editing in lignin engineering and its integration in tree breeding programs.

Keywords: CRISPR; field trial; forest trees; genetic engineering; lignin.

FIGURE 1

Lignin biosynthetic pathway. Alternative monomers and heterologously expressed enzymes are shown in bold. PAL, phenylalanine ammonia-lyase; C4H, cinnamate 4-hydroxylase; 4CL, 4-coumarate:CoA ligase; HCT, p-Hydroxycinnamoyl-CoA:quinate/shikimate-p-hydroxycinnamoyltransferase; C3’H, p-coumaroyl quinate/shikimate 3’-hydroxylase; CSE, caffeoyl shikimate esterase; CCoAOMT, caffeoyl-CoA O-methyltransferase; CCR, cinnamoyl-CoA reductase; F5H, ferulate 5-hydroxylase/CAld5H, coniferaldehyde 5-hydroxylase; COMT, caffeic acid O-methyltransferase; CAD, cinnamyl alcohol dehydrogenase; FMT, feruloyl-CoA monolignol transferase; PMT, p-coumaroyl-CoA monolignol transferase.

FIGURE 2

Patchy gene downregulation by RNAi. Patchy red xylem phenotype observed on trunks of CCR-deficient poplars (right) grown in a field trial in Belgium. The red xylem indicates areas of CCR downregulation. Wood from wild-type trees is whitish (left).

FIGURE 3

Genetic improvement of forest trees through a combination of breeding tools. To accelerate the genetic improvement of forest trees for pulp and biorefinery applications, classical and new breeding tools need to be smartly combined. Classical breeding involves phenotypic selection of trees for controlled crosses, followed by phenotypic selection. With the advent of genome sequence information of many forest trees, new strategies such as Genomic Selection, Genome Wide Association Studies (GWAS) and Breeding with Rare Defective Alleles (BRDA) have been developed to speed up the capture and enrichment of DNA polymorphisms associated with beneficial traits. CRISPR-based genome editing allows to modify the genome in a way that mimics natural polymorphisms. Genetic modification involves the stable integration of foreign DNA into the tree to overproduce (an) enzyme(s) or downregulate (a) gene(s). Combining the classical and New Breeding Techniques is needed to provide sufficient highly quality wood for society.