Asparagus Spears as a Model to Study Heteroxylan Biosynthesis during Secondary Wall Development.

Lili Song,Wei Zeng,Cherie Beahan,Andrew Lonsdale,Edwin R Lampugnani,Kelsey Picard,Roshan Cheetamun,Antony Bacic,Monika S Doblin,Andrew Cassin,Aimin Wu,Olga A Zabotina

doi:10.1371/journal.pone.0123878

Lili Song, Wei Zeng + Show 10 more

Open Access

https://doi.org/10.1371/journal.pone.0123878

Copy DOI

Abstract

Garden asparagus (Asparagus officinalis L.) is a commercially important crop species utilized for its excellent source of vitamins, minerals and dietary fiber. However, after harvest the tissue hardens and its quality rapidly deteriorates because spear cell walls become rigidified due to lignification and substantial increases in heteroxylan content. This latter observation prompted us to investigate the in vitro xylan xylosyltransferase (XylT) activity in asparagus. The current model system for studying heteroxylan biosynthesis, Arabidopsis, whilst a powerful genetic system, displays relatively low xylan XylT activity in in vitro microsomal preparations compared with garden asparagus therefore hampering our ability to study the molecular mechanism(s) of heteroxylan assembly. Here, we analyzed physiological and biochemical changes of garden asparagus spears stored at 4 °C after harvest and detected a high level of xylan XylT activity that accounts for this increased heteroxylan. The xylan XylT catalytic activity is at least thirteen-fold higher than that reported for previously published species, including Arabidopsis and grasses. A biochemical assay was optimized and up to seven successive Xyl residues were incorporated to extend the xylotetraose (Xyl4) acceptor backbone. To further elucidate the xylan biosynthesis mechanism, we used RNA-seq to generate an Asparagus reference transcriptome and identified five putative xylan biosynthetic genes (AoIRX9, AoIRX9-L, AoIRX10, AoIRX14_A, AoIRX14_B) with AoIRX9 having an expression profile that is distinct from the other genes. We propose that Asparagus provides an ideal biochemical system to investigate the biochemical aspects of heteroxylan biosynthesis and also offers the additional benefit of being able to study the lignification process during plant stem maturation.

Highlights

Heteroxylans represent a major family of non-cellulosic polysaccharides in dicot secondary walls and monocot primary walls [1]
They are composed of a linear backbone of 1,4-linked β-D-xylose (Xyl) residues substituted with variable side branches that are mostly composed of α-D-glucuronic acid (GlcA), 4-O-methyl-α-D-glucuronic acid (MeGlcA) and/or α-Larabinofuranose (Araf) depending on the species and tissue types [2]
Together our results show that Asparagus is an ideal non-commelinoid monocot model system to study heteroxylan biosynthesis, as it offers a tractable system to purify the in vivo protein complex responsible for the biochemical activity that will be essential in elucidating our understanding of the molecular mechanism of xylan synthesis

Summary

Introduction

Heteroxylans represent a major family of non-cellulosic polysaccharides in dicot secondary walls and monocot primary walls [1]. They are composed of a linear backbone of 1,4-linked β-D-xylose (Xyl) residues substituted with variable side branches that are mostly composed of α-D-glucuronic acid (GlcA), 4-O-methyl-α-D-glucuronic acid (MeGlcA) and/or α-Larabinofuranose (Araf) depending on the species and tissue types [2]. In Arabidopsis, several GTs involved in heteroxylan biosynthesis have been identified via mutant analysis These include members of the CAZy GT family 47 (IRX10 and FRA8/IRX7) [7,8,9], GT43 (IRX9 and IRX14) [10,11] and GT8 (IRX8, PARVUS and GUX) [12,13,14]. Several gene families responsible for the glycosyl residue substitution by non-glycosyl residues have been reported and include DUF579 for transfer of a methyl group to GlcA on the xylan backbone [18], BAHD involved in ferulic and p-coumaric acid addition [19] and TBL29 catalyzing Xyl acetylation [20,21]

Methods

Results

Discussion

Conclusion