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

Song L, Zeng W, Wu A, Picard K, Lampugnani ER, Cheetamun R, Beahan C, Cassin A, Lonsdale A, Doblin MS, Bacic A - PLoS ONE (2015)

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin'an, Hangzhou, 311300, P. R. China; ARC Centre of Excellence in Plant Cell Walls, School of Botany, the University of Melbourne, Parkville, VIC 3010, Australia.

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.

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Comparison of xylan XylT activities in microsomes from Arabidopsis, Asparagus and barley (Hordeum vulgare).Microsomes from Arabidopsis stem, Asparagus spear and etiolated barley seedlings were isolated and the XylT activities were analyzed in the absence (-; black) and presence (+; grey) of the exogenous acceptor Xyl6 according to the Materials and Methods.
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pone.0123878.g002: Comparison of xylan XylT activities in microsomes from Arabidopsis, Asparagus and barley (Hordeum vulgare).Microsomes from Arabidopsis stem, Asparagus spear and etiolated barley seedlings were isolated and the XylT activities were analyzed in the absence (-; black) and presence (+; grey) of the exogenous acceptor Xyl6 according to the Materials and Methods.

Mentions: Using a paper chromatography assay measuring incorporation of [14C]-Xyl into a Xyl6 acceptor, Arabidopsis and particularly Asparagus microsomes showed XylT activity (Fig 2). In contrast, no activity could be detected in microsomes prepared from barley coleoptiles. Arabidopsis microsomes incorporated ~2% of total radiolabelled Xyl whereas Asparagus incorporated ~26%, i.e. a 13-fold higher amount than that of Arabidopsis.


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

Song L, Zeng W, Wu A, Picard K, Lampugnani ER, Cheetamun R, Beahan C, Cassin A, Lonsdale A, Doblin MS, Bacic A - PLoS ONE (2015)

Comparison of xylan XylT activities in microsomes from Arabidopsis, Asparagus and barley (Hordeum vulgare).Microsomes from Arabidopsis stem, Asparagus spear and etiolated barley seedlings were isolated and the XylT activities were analyzed in the absence (-; black) and presence (+; grey) of the exogenous acceptor Xyl6 according to the Materials and Methods.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4404143&req=5

pone.0123878.g002: Comparison of xylan XylT activities in microsomes from Arabidopsis, Asparagus and barley (Hordeum vulgare).Microsomes from Arabidopsis stem, Asparagus spear and etiolated barley seedlings were isolated and the XylT activities were analyzed in the absence (-; black) and presence (+; grey) of the exogenous acceptor Xyl6 according to the Materials and Methods.
Mentions: Using a paper chromatography assay measuring incorporation of [14C]-Xyl into a Xyl6 acceptor, Arabidopsis and particularly Asparagus microsomes showed XylT activity (Fig 2). In contrast, no activity could be detected in microsomes prepared from barley coleoptiles. Arabidopsis microsomes incorporated ~2% of total radiolabelled Xyl whereas Asparagus incorporated ~26%, i.e. a 13-fold higher amount than that of Arabidopsis.

Bottom Line: 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.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.

View Article: PubMed Central - PubMed

Affiliation: Nurturing Station for the State Key Laboratory of Subtropical Silviculture, Zhejiang A & F University, Lin'an, Hangzhou, 311300, P. R. China; ARC Centre of Excellence in Plant Cell Walls, School of Botany, the University of Melbourne, Parkville, VIC 3010, Australia.

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.

Show MeSH
Related in: MedlinePlus