Cell wall remodeling under salt stress : Insights into changes in polysaccharides, feruloylation, lignification, and phenolic metabolism in maize

Although cell wall polymers play important roles in the tolerance of plants to abiotic stress, the effects of salinity on cell wall composition and metabolism in grasses remain largely unexplored. Here, we conducted an in-depth study of changes in cell wall composition and phenolic metabolism induced upon salinity in maize seedlings and plants. Cell wall characterization revealed that salt stress modulated the deposition of cellulose, matrix polysaccharides and lignin in seedling roots, plant roots and stems. The extraction and analysis of arabinoxylans by size-exclusion chromatography, 2D-NMR spectroscopy and carbohydrate gel electrophoresis showed a reduction of arabinoxylan content in salt-stressed roots. Saponification and mild acid hydrolysis revealed that salinity also reduced the feruloylation of arabinoxylans in roots of seedlings and plants. Determination of lignin content and composition by nitrobenzene oxidation and 2D-NMR confirmed the increased incorporation of syringyl units in lignin of maize roots. Salt stress also induced the expression of genes and the activity of enzymes enrolled in phenylpropanoid biosynthesis. The UHPLC-MS-based metabolite profiling confirmed the modulation of phenolic profiling by salinity and the accumulation of ferulate and its derivatives 3- and 4-O-feruloyl quinate. In conclusion, we present a model for explaining cell wall remodeling in response to salinity. This article is protected by copyright. All rights reserved

Feruloyl esterases: Biocatalysts to overcome biomass recalcitrance and for the production of bioactive compounds

International audienceFerulic acid and its hydroxycinnamate derivatives represent one of the most abundant forms of low molecular weight phenolic compounds in plant biomass. Feruloyl esterases are part of a microorganism’s plant cell wall-degrading enzymatic arsenal responsible for cleaving insoluble wall-bound hydroxycinnamates and soluble cytosolic conjugates. Stimulated by industrial requirements, accelerating scientific discoveries and knowledge transfer, continuous improvement efforts have been made to identify, create and repurposed biocatalysts dedicated to plant biomass conversion and biosynthesis of high-added value molecules. Here we review the basic knowledge and recent advances in biotechnological characteristics and the gene content encoding for feruloyl esterases. Information about several enzymes is systematically organized according to their function, biochemical properties, substrate specificity, and biotechnological applications. This review contributes to further structural, functional, and biotechnological R&D both for obtaining hydroxycinnamates from agricultural by-products as well as for lignocellulose biomass treatments aiming for production of bioethanol and other derivatives of industrial interest

Inhibiting tricin biosynthesis improves maize lignocellulose saccharification.

Lignin is a technological bottleneck to convert polysaccharides into fermentable sugars, and different strategies of genetic-based metabolic engineering have been applied to improve biomass saccharification. Using maize seedlings grown hydroponically for 24 h, we conducted a quick non-transgenic approach with five enzyme inhibitors of the lignin and tricin pathways. Two compounds [3,4-(methylenedioxy)cinnamic acid: MDCA and 2,4-pyridinedicarboxylic acid: PDCA] revealed interesting findings on root growth, lignin composition, and saccharification. By inhibiting hydroxycinnamoyl-CoA ligase, a key enzyme of phenylpropanoid pathway, MDCA decreased the lignin content and improved saccharification, but it decreased root growth. By inhibiting flavone synthase, a key enzyme of tricin biosynthesis, PDCA decreased total lignin content and improved saccharification without affecting root growth. PDCA was three-fold more effective than MDCA, suggesting that controlling lignin biosynthesis with enzymatic inhibitors may be an attractive strategy to improve biomass saccharification

Designing xylan for improved sustainable biofuel production

Increasing greenhouse gas emissions and diminishing supplies of fossil-derived fuels underline the need for environmentally sustainable energy resources. Lignocellulose, more generically named simply as plant biomass, represent one of the most abundant renewable resources for biofuels. Lignocellulosic biomass from forest residues, agro-wastes and energy grasses is extensively exploited for bioenergy production. This article is protected by copyright. All rights reserved

Feruloyl esterase activity and its role in regulating the feruloylation of maize cell walls

Cell walls of grasses have ferulic acid (FA) ester-linked to the arabinosyl substitutions of arabinoxylan (AX). Feruloyl esterases (FAE) are carboxylic acid esterases that release FA from cell walls and synthetic substrates. Despite the importance of FA for cell wall recalcitrance and in response to biotic and abiotic stresses, the physiological function of plant FAEs remains unclear. Here, we developed a simple method for the determination of FAE activity (ZmFAE) in maize using the total protein extract and investigated its role in regulating the feruloylation of cell wall. The method includes a single protein extraction and enzymatic reaction with protein concentration as low as 65 mu g at 35 degrees C for 30 min, using methyl ferulate as the substrate. The methodology allowed the determination of the apparent K-m (392.82 mu M) and V-max (79.15 pkat mg(-1) protein). We also found that ZmFAE activity was correlated (r = 0.829) with the levels of FA in seedling roots, plant roots and leaves of maize. Furthermore, the exposure to osmotic stress resulted in a 50% increase in ZmFAE activity in seedling roots. These data suggest that FAE-catalyzed reaction is important for cell wall feruloylation during plant development and in response to abiotic stress. We conclude proposing a model for the feruloylation and deferuloylation of AX, which explains the role of FAE in regulating the levels of ester-linked FA. Our model might orient further studies investigating the role of plant FAEs and assist strategies for genetic engineering of grasses to obtain plants with reduced biomass recalcitrance

Feruloyl esterase from Aspergillus clavatus improves xylan hydrolysis of sugarcane bagasse

Feruloyl esterase is a subclass of carboxylic acid esterases with the capacity to release ferulic acid and other cinnamic acids from plant cell walls and synthetic substrates. Feruloyl esterases act synergistically with xylanases removing ferulic acid residues esterified to arabinoxylans. Feruloyl esterase type D from Aspergillus clavatus (AcFAE) was expressed in Escherichia coli, purified, and applied with a commercial xylanase consortium (Novozymes) for hydrolysis of sugarcane bagasse. Feruloyl esterase plus xylanase increased 5.13-fold the releasing of ferulic acid from sugarcane bagasse. Removal of only 7.7% of ferulic acid content by AcFAE increased 97.3% the sugarcane bagasse hydrolysis by xylanase. These data support the use of AcFAE as an interesting adjuvant enzyme to improve lignocellulose digestion and biotechnological tool for biorefineries

Suppression of a BAHD acyltransferase decreases p‐coumaroyl on arabinoxylan and improves biomass digestibility in the model grass Setaria viridis

Grass cell walls have hydroxycinnamic acids attached to arabinosyl residues of arabinoxylan (AX), and certain BAHD acyltransferases are involved in their addition. In this study, we characterized one of these BAHD genes in the cell wall of the model grass Setaria viridis. RNAi silenced lines of S. viridis (SvBAHD05) presented a decrease of up to 42% of ester-linked p-coumarate (pCA) and 50% of pCA-arabinofuranosyl, across three generations. Biomass from SvBAHD05 silenced plants exhibited up to 32% increase in biomass saccharification after acid pre-treatment, with no change in total lignin. Molecular dynamics simulations suggested that SvBAHD05 is a p-coumaroyl coenzyme A transferase (PAT) mainly involved in the addition of pCA to the arabinofuranosyl residues of AX in Setaria. Thus, our results provide evidence of p-coumaroylation of AX promoted by SvBAHD05 acyltransferase in the cell wall of the model grass S. viridis. Furthermore, SvBAHD05 is a promising biotechnological target to engineer crops for improved biomass digestibility for biofuels, biorefineries and animal feeding