Two conifer GUX clades are responsible for distinct glucuronic acid patterns on xylan

Abstract

 Wood of coniferous trees (softwood), is a globally significant carbon sink and an important source of biomass. Despite that, little is known about the genetic basis of softwood cell wall biosynthesis. Branching of xylan, one of the main hemicelluloses in softwood secondary cell walls, with glucuronic acid (GlcA) is critical for biomass recalcitrance. Here, we investigate the decoration patterns of xylan by conifer GlucUronic acid substitution of Xylan (GUX) enzymes. Through molecular phylogenetics we identify two distinct conifer GUX clades.Using transcriptional profiling we show that the genes are preferentially expressed in secondary cell wall forming tissues. With in vitro and in planta assays we demonstrate that conifer GUX enzymes from both clades are active glucuronyltransferases. Conifer GUX enzymes from each clade have different specific activities. While members of clade one add evenly spaced GlcA branches, the members of clade two are also capable of glucuronidating two consecutive xyloses. Importantly, these types of xylan patterning are present in softwood. Since xylan patterning might modulate xylan-cellulose and xylan-lignin interactions, our results further the understanding of softwood cell wall biosynthesis and provide breeding or genetic engineering targets that can be used to modify softwood properties.

The biosynthesis, degradation, and function of cell wall β-xylosylated xyloglucan mirrors that of arabinoxyloglucan

“…The promoters of XXT2 and CESA3 and the coding sequence of Sl XST1 were amplified from Arabidopsis or tomato genomic DNA, respectively, by PCR using Q5 High-Fidelity DNA Polymerase (New England Biolabs, Ipswich, MA, USA; primers are listed in Table S3). Constructs and transgenic Arabidopsis lines were then generated as previously described (Lyczakowski et al ., 2021).…”

Importance of Water in Maintaining Softwood Secondary Cell Wall Nanostructure

“…These hemicelluloses are themselves interacting with lignin via electrostatic contacts and α-ether or glucuronylester (of xylan) linkages. ,− Moreover, the even patterning produced by the repeat of the XUXAXX pattern of AGX softwood allows it to bind cellulose in a two-fold screw configuration . A smaller proportion of three-fold screw AGX, possibly with two consecutive GlcA substitutions (XUUXX), may interact with the lignin nanodomain via electrostatic interactions. ,,,− However, so far none of these studies considered the importance of the cell wall hydration status or its importance for the polymer conformations in their native state.…”

Glucuronic acid: not just another brick in the cell wall

“…Xylan glucuronosyl transferases encoded by GUX1 – GUX5 (GlucUronic acid substitution of Xylan 1–5) add glucuronic acid to xylan, a feature that allowed the characterization of novel functions for MeGlcA substitutions. Mutations of the GUX genes support a role for these enzymes in xylan glucuronosylation in Arabidopsis (Mortimer et al ., 2010, 2015; Bromley et al ., 2013) and more recently in conifers (Lyczakowski et al ., 2021). The gux1 and gux2 Arabidopsis single mutants show reduced xylan glucuronosylation, while the gux1gux2 double mutant has a nearly complete depletion of MeGlcA substitutions without any associated reduction in xylan amount, indicating that xylan backbone biosynthesis is independent of its substitutions (Mortimer et al ., 2010).…”

“…Xylan hemicelluloses interact with cellulose microfibrils and are covalently linked to lignins, forming a lignin–carbohydrate complex. Although the role of modifications of the xylan structure remains unclear, recent studies have shed light on new functions of xylan substitutions in the assembly of cell walls and how such substitutions are associated with improved depolymerization of cell wall polysaccharides (Whitehead et al ., 2018; Lyczakowski et al ., 2021; Tryfona et al ., 2023). In this issue of New Phytologist , Derba‐Maceluch et al .…”