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At least 235 records · Page 13
All sourcesNASA NTRSDOE OSTI
DOE OSTI2021

Exogenous chalcone synthase expression in developing poplar xylem incorporates naringenin into lignins

Abstract Lignin, a polyphenolic polymer, is a major chemical constituent of the cell walls of terrestrial plants. The biosynthesis of lignin is a highly plastic process, as highlighted by an increasing number of noncanonical monomers that have been successfully identified in an array of plants. Here, we engineered hybrid poplar (Populus alba x grandidentata) to express chalcone synthase 3 (MdCHS3) derived from apple (Malus domestica) in lignifying xylem. Transgenic trees displayed an accumulation of the flavonoid naringenin in xylem methanolic extracts not inherently observed in wild-type trees. Nuclear magnetic resonance analysis revealed the presence of naringenin in the extract-free, cellulase-treated xylem lignin of MdCHS3-poplar, indicating the incorporation of this flavonoid-derived compound into poplar secondary cell wall lignins. The transgenic trees also displayed lower total cell wall lignin content and increased cell wall carbohydrate content and performed significantly better in limited saccharification assays than their wild-type counterparts.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Insights of Molecular Mechanism of Xylem Development in Five Black Poplar Cultivars

Black poplar (Populus deltoides, P. nigra, and their hybrids) is the main poplar cultivars in China. It offers interesting options of large-scale biomass production for bioenergy due to its rapid growth and high yield. Poplar wood properties were associated with chemical components and physical structures during wood formation. In this study, five poplar cultivars, P. euramericana ‘Zhonglin46’ (Pe1), P. euramericana ‘Guariento’ (Pe2), P. nigra ‘N179’ (Pn1), P. deltoides ‘Danhong’ (Pd1), and P. deltoides ‘Nanyang’ (Pd2), were used to explore the molecular mechanism of xylem development. We analyzed the structural differences of developing xylem in the five cultivars and profiled the transcriptome-wide gene expression patterns through RNA sequencing. The cross sections of the developing xylem showed that the cell wall thickness of developed fiber in Pd1 was thickest and the number of xylem vessels of Pn1 was the least. A total of 10,331 differentially expressed genes were identified among 10 pairwise comparisons of the five cultivars, most of them were related to programmed cell death and secondary cell wall thickening. K-means cluster analysis and Gene Ontology enrichment analysis showed that the genes highly expressed in Pd1 were related to nucleotide decomposition, metabolic process, transferase, and microtubule cytoskeleton; whereas the genes highly expressed in Pn1 were involved in cell wall macromolecule decomposition and polysaccharide binding processes. Based on a weighted gene co-expression network analysis, a large number of candidate regulators for xylem development were identified. And their potential regulatory roles to cell wall biosynthesis genes were validated by a transient overexpression system. This study provides a set of promising candidate regulators for genetic engineering to improve feedstock and enhance biofuel conversion in the bioenergy crop Populus.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

The molecular architecture distinctions between compression, opposite and normal wood of Pinus radiata

In gymnosperms compression wood is a specialised type of structural cell wall formed in response to biomechanical stresses. The differences in terms of gross structure, ultrastructure and chemistry are well-known. However, the differences between compression wood, normal wood, and opposite wood regarding the arrangements and interactions of the various polymers and water within their cell walls still needs to be established. The analysis of 13 C-labelled Pinus radiata by solid-state NMR spectroscopy and other complementary techniques revealed several new aspects of compression and opposite wood molecular architecture. Compared to normal wood, compression wood has a lower water content, its overall nanoporosity is reduced, and the water and matrix polymers have a lower molecular mobility. Galactan, which is a specific marker of compression wood, is broadly distributed within the cell wall, disordered, and not aligned with cellulose, and is found to be in close proximity to xylan. Dehydroabietic acid (a resin acid) is immobilised and close to the H-lignin only in compression wood. Although the overall molecular mobility of normal wood and opposite wood are similar, opposite wood has different arabinose conformations, a large increase in the amount of chain ends, contains significantly more galactan and has additional unassigned mobile components highlighting the different molecular arrangement of cell wall polymers in opposite and normal wood.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Architecture of a catalytically active homotrimeric plant cellulose synthase complex

Cellulose is an essential plant cell wall component and represents the most abundant biopolymer on Earth. Supramolecular plant cellulose synthase complexes organize multiple linear glucose polymers into microfibrils as load-bearing wall components. We determined the structure of a poplar cellulose synthase CesA homotrimer that suggests a molecular basis for cellulose microfibril formation. This complex, stabilized by cytosolic plant-conserved regions and helical exchange within the transmembrane segments, forms three channels occupied by nascent cellulose polymers. Secretion steers the polymers toward a common exit point, which could facilitate protofibril formation. CesA’s N-terminal domains assemble into a cytosolic stalk that interacts with a microtubule-tethering protein and may thus be involved in CesA localization. Here, our data suggest how cellulose synthase complexes assemble and provide the molecular basis for plant cell wall engineering.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Metatranscriptomics sheds light on the links between the functional traits of fungal guilds and ecological processes in forest soil ecosystems

Soil fungi belonging to different functional guilds, such as saprotrophs, pathogens, and mycorrhizal symbionts, play key roles in forest ecosystems. To date, no study has compared the actual gene expression of these guilds in different forest soils. We used metatranscriptomics to study the competition for organic resources by these fungal groups in boreal, temperate, and Mediterranean forest soils. Using a dedicated mRNA annotation pipeline combined with the JGI MycoCosm database, we compared the transcripts of these three fungal guilds, targeting enzymes involved in C- and N mobilization from plant and microbial cell walls. Genes encoding enzymes involved in the degradation of plant cell walls were expressed at a higher level in saprotrophic fungi than in ectomycorrhizal and pathogenic fungi. However, ectomycorrhizal and saprotrophic fungi showed similarly high expression levels of genes encoding enzymes involved in fungal cell wall degradation. Transcripts for N-related transporters were more highly expressed in ectomycorrhizal fungi than in other groups. Here, we showed that ectomycorrhizal and saprotrophic fungi compete for N in soil organic matter, suggesting that their interactions could decelerate C cycling. Metatranscriptomics provides a unique tool to test controversial ecological hypotheses and to better understand the underlying ecological processes involved in soil functioning and carbon stabilization.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

High temperature high strength austenitic steel fabricated by laser powder-bed fusion

Extremely fast cooling rates during laser powder-bed fusion (LPBF) can result in materials with unique microstructures. For LPBF 316L stainless steel, the formation of sub-grain cellular structures with high dislocation density has been linked to superior tensile properties at room temperature. This cellular structure offers also a new route for the development of high temperature LPBF steels with the nucleation of nano-size strengthening carbides in the cell walls. HK30Nb steel (Fe-25Cr-20Ni-Nb-C) was, therefore, fabricated by LPBF to evaluate its potential for high temperature applications. Optimization of the fabrication parameters yielded material with density greater than 99.7%, with nano Nb-rich precipitates in the cell walls. Annealing at 800 °C for 5h resulted in the nucleation and growth of additional precipitates mainly in the cell walls and at grain boundaries. Furthermore, the high dislocation density led to yield strength at 20–900 °C two to three times higher than yield strength for cast HK30Nb and the nano carbides in the cell walls significantly improved the cellular structure stability at 800 °C.

36 MATERIALS SCIENCE↗
DOE OSTI2021

Control of Carbon Allocation in Phenylpropanoid Metabolism

Understanding the molecular mechanisms by which carbon flux is partitioned in plants, and how flux through the phenylpropanoid pathway in particular is regulated and allocated to different branches of the pathway is essential for the eventual rational manipulation of phenylpropanoid flux for bioenergy applications. The phenylpropanoid pathway is responsible for the biosynthesis of a diverse array of metabolites essential for structural integrity, water transport, UV protection, and defense against herbivores and pathogens. The products of the phenylpropanoid pathway, all of which are derived from phenylalanine, range from complex, insoluble polymers such as lignin and suberin, to soluble flavonoids and hydroxycinnamate esters, to volatile compounds used to attract pollinators. By far the most abundant of these is the complex heteropolymer lignin, which accounts for a significant portion of the dry weight of not only woody plants, but also of crop residues. Decreasing or altering lignin structure to provide increased cell wall digestibility could greatly increase the cost effectiveness of converting cell wall polysaccharides to ethanol or second-generation biofuels. On the other hand, it is also possible that plants engineered to accumulate higher levels of lignin could provide improved feedstock for direct combustion and electricity generation as part of a diverse bioenergy portfolio. Consistent with the enormous commitment of energy and carbon it requires, the synthesis of lignin is under tight regulatory control. In order to influence transcription, gene-specific activators and repressors must interact with basal transcription factors and the rest of the core transcriptional machinery. In eukaryotes, this interaction is bridged by the large, multi-subunit complex known as Mediator. Despite the clear importance of Mediator throughout the eukaryotes, the complex has only recently been studied in plants. We have shown that a pair of Mediator subunits, MED5a and MED5b are required for normal regulation of phenylpropanoid pathway product accumulation. Plants lacking these proteins show increased accumulation of phenylpropanoids in multiple tissues indicating that normally they directly or indirectly repress expression of phenylpropanoid pathway genes. Furthermore, we have recently shown that MED5 plays a role in dwarfing in some low lignin plants and feedback between and within metabolic pathways of secondary metabolism in Arabidopsis. Taken together with the established roles of Mediator in transcriptional regulation, and of MED5a and MED5b in the repression of phenylpropanoid metabolism, these observations suggest that MED5a and MED5b are key components of an active, transcriptional process by which phenylpropanoid homeostasis is maintained in wild-type plants. Our research focused on expanding our understanding of the role of Mediator, as well as elucidating the mechanisms by which these proteins influence phenylpropanoid biosynthesis and how metabolite accumulation within the pathway and between metabolic pathways is essential for feedback regulatory control. This research will inform future bioengineering efforts aimed at altering carbon allocation and cell wall engineering by contributing to our understanding of the role of Mediator in regulating lignin biosynthesis, a pathway of plant metabolism that is important on a global scale.

09 BIOMASS FUELS↗
DOE OSTI2022

Biomass formation and sugar release efficiency of Populus modified by altered expression of a NAC transcription factor

Woody biomass is an important feedstock for biofuel production. Manipulation of wood properties that enable efficient conversion of biomass to biofuel reduces cost of biofuel production. Wood cell wall composition is regulated at several levels that involve expression of transcription factors such as wood-/secondary cell wall-associated NAC domains (WND or SND). In Arabidopsis thaliana, SND1 regulates cell wall composition through activation of its down-stream targets such as MYBs. The functional aspects of SND1 homologs in the woody Populus have been studied through transgenic manipulation. In this study, we investigated the role of PdWND1B, Populus SND1 sequence ortholog, in wood formation using transgenic manipulation through over-expression or silencing under the control of a vascular-specific 4-coumarate-CoA ligase (4CL) promoter. As compared with control plants, PdWND1B-RNAi plants were shorter in height, with significantly reduced stem diameter and dry biomass, whereas there were no significant differences in growth and productivity of PdWND1B over-expression plants. Conversely, PdWND1B over-expression lines showed a significant reduction in cellulose and increase in lignin content, whereas there was no significant impact on lignin content of downregulated lines. Stem carbohydrate composition analysis revealed a decrease in glucose, mannose, arabinose, and galactose, but an increase in xylose in the over-expression lines. Transcriptome analysis revealed upregulation of several downstream transcription factors and secondary cell wall related structural genes in the PdWND1B over-expression lines, partly explaining the observed phenotypic changes in cell wall chemistry. Relative to the control, glucose release efficiency and ethanol production from stem biomass was significantly reduced in over-expression lines. Our results show that PdWND1B is an important factor determining biomass productivity, cell wall chemistry and its conversion to biofuels in Populus.

09 BIOMASS FUELS↗
DOE OSTI2024

Ectopic Production of 3,4-Dihydroxybenzoate in Planta Affects Cellulose Structure and Organization

Lignocellulosic biomass is a highly sustainable and largely carbon dioxide neutral feedstock for the production of biofuels and advanced biomaterials. Although thermochemical pretreatment is typically used to increase the efficiency of cell wall deconstruction, genetic engineering of the major plant cell wall polymers, especially lignin, has shown promise as an alternative approach to reduce biomass recalcitrance. Poplar trees with reduced lignin content and altered composition were previously developed by overexpressing bacterial 3-dehydroshikimate dehydratase (QsuB) enzyme to divert carbon flux from the shikimate pathway. In this work, three transgenic poplar lines with increasing QsuB expression levels and different lignin contents were studied using small-angle neutron scattering (SANS) and wide-angle X-ray scattering (WAXS). SANS showed that although the cellulose microfibril cross-sectional dimension remained unchanged, the ordered organization of the microfibrils progressively decreased with increased QsuB expression. This was correlated with decreasing total lignin content in the QsuB lines. WAXS showed that the crystallite dimensions of cellulose microfibrils transverse to the growth direction were not affected by the QsuB expression, but the crystallite dimensions parallel to the growth direction were decreased by ~20%. Cellulose crystallinity was also decreased with increased QsuB expression, which could be related to high levels of 3,4-dihydroxybenzoate, the product of QsuB expression, disrupting microfibril crystallization. In addition, the cellulose microfibril orientation angle showed a bimodal distribution at higher QsuB expression levels. Altogether, this study provides new structural insights into the impact of ectopic synthesis of small-molecule metabolites on cellulose organization and structure that can be used for future efforts aimed at reducing biomass recalcitrance.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Mutations in the Pectin Methyltransferase QUASIMODO2 Influence Cellulose Biosynthesis and Wall Integrity in Arabidopsis

Abstract Pectins are abundant in the cell walls of dicotyledonous plants, but how they interact with other wall polymers and influence wall integrity and cell growth has remained mysterious. Here, we verified that QUASIMODO2 (QUA2) is a pectin methyltransferase and determined that QUA2 is required for normal pectin biosynthesis. To gain further insight into how pectin affects wall assembly and integrity maintenance, we investigated cellulose biosynthesis, cellulose organization, cortical microtubules, and wall integrity signaling in two mutant alleles of Arabidopsis (Arabidopsis thaliana) QUA2, qua2 and tsd2. In both mutants, crystalline cellulose content is reduced, cellulose synthase particles move more slowly, and cellulose organization is aberrant. NMR analysis shows higher mobility of cellulose and matrix polysaccharides in the mutants. Microtubules in mutant hypocotyls have aberrant organization and depolymerize more readily upon treatment with oryzalin or external force. The expression of genes related to wall integrity, wall biosynthesis, and microtubule stability is dysregulated in both mutants. These data provide insights into how homogalacturonan is methylesterified upon its synthesis, the mechanisms by which pectin functionally interacts with cellulose, and how these interactions are translated into intracellular regulation to maintain the structural integrity of the cell wall during plant growth and development.

Biochemistry & Molecular Biology↗
DOE OSTI2020

Molecular remodeling in Populus PdKOR RNAi roots profiled using LC-MS/MS proteomics

Plant endo-β-1,4-glucanases belonging to the Glycoside Hydrolase Family 9 have functional roles in cell wall biosynthesis and remodeling via endohydrolysis of (1→4)-β-D-glucosidic linkages. Modification of cell wall chemistry via RNAi-mediated downregulation of Populus deltoides KOR1 (PdKOR), a endo-β-1,4-glucanase gene, in Populus deltoides has been shown to have functional consequences for the composition of secondary metabolome and the ability of modified roots to interact with beneficial microbes. The molecular remodeling that underlies the observed differences at metabolic, physiological, and morphological levels in roots is not well understood. Here we used a LC-MS/MS-based proteome profiling approach to survey the molecular remodeling in root tissues of PdKOR and control plants. A total of 14316 peptides were identified and these mapped to 7139 P. deltoides proteins. Based on 90% sequence identity, the measured protein accessions represent 1187 functional protein groups. Analysis of GO categories and specific individual proteins showed differential expression of proteins relevant to plant-microbe interactions, cell wall chemistry, and metabolism. The new proteome dataset serves as a useful resource for deriving new hypotheses and empirical testing pertaining to functional roles of proteins and pathways in differential priming of plant roots to interactions with microbes.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2022

Engineering Curcumin Biosynthesis in Poplar Affects Lignification and Biomass Yield

Lignocellulosic biomass is recalcitrant toward deconstruction into simple sugars mainly due to the presence of lignin. By engineering plants to partially replace traditional lignin monomers with alternative ones, lignin degradability and extractability can be enhanced. Previously, the alternative monomer curcumin has been successfully produced and incorporated into lignified cell walls of Arabidopsis by the heterologous expression of DIKETIDE-CoA SYNTHASE (DCS) and CURCUMIN SYNTHASE2 (CURS2). The resulting transgenic plants did not suffer from yield penalties and had an increased saccharification yield after alkaline pretreatment. Here, we translated this strategy into the bio-energy crop poplar. Via the heterologous expression of DCS and CURS2 under the control of the secondary cell wall CELLULOSE SYNTHASE A8-B promoter (ProCesA8-B), curcumin was also produced and incorporated into the lignified cell walls of poplar. ProCesA8-B:DCS_CURS2 transgenic poplars, however, suffered from shoot-tip necrosis and yield penalties. Compared to that of the wild-type (WT), the wood of transgenic poplars had 21% less cellulose, 28% more matrix polysaccharides, 23% more lignin and a significantly altered lignin composition. More specifically, ProCesA8-B:DCS_CURS2 lignin had a reduced syringyl/guaiacyl unit (S/G) ratio, an increased frequency of p-hydroxyphenyl (H) units, a decreased frequency of p-hydroxybenzoates and a higher fraction of phenylcoumaran units. Without, or with alkaline or hot water pretreatment, the saccharification efficiency of the transgenic lines was equal to that of the WT. These differences in (growth) phenotype illustrate that translational research in crops is essential to assess the value of an engineering strategy for applications. Further fine-tuning of this research strategy (e.g., by using more specific promoters or by translating this strategy to other crops such as maize) might lead to transgenic bio-energy crops with cell walls more amenable to deconstruction without settling in yield.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2022

One fold, many functions—M23 family of peptidoglycan hydrolases

Bacterial cell walls are the guards of cell integrity. They are composed of peptidoglycan that provides rigidity to sustain internal turgor and ensures isolation from the external environment. In addition, they harbor the enzymatic machinery to secure cell wall modulations needed throughout the bacterial lifespan. The main players in this process are peptidoglycan hydrolases, a large group of enzymes with diverse specificities and different mechanisms of action. They are commonly, but not exclusively, found in prokaryotes. Although in most cases, these enzymes share the same molecular function, namely peptidoglycan hydrolysis, they are leveraged to perform a variety of physiological roles. A well-investigated family of peptidoglycan hydrolases is M23 peptidases, which display a very conserved fold, but their spectrum of lytic action is broad and includes both Gram- positive and Gram- negative bacteria. In this review, we summarize the structural, biochemical, and functional studies concerning the M23 family of peptidases based on literature and complement this knowledge by performing large-scale analyses of available protein sequences. This review has led us to gain new insight into the role of surface charge in the activity of this group of enzymes. We present relevant conclusions drawn from the analysis of available structures and indicate the main structural features that play a crucial role in specificity determination and mechanisms of latency. Our work systematizes the knowledge of the M23 family enzymes in the context of their unique antimicrobial potential against drug-resistant pathogens and presents possibilities to modulate and engineer their features to develop perfect antibacterial weapons.

Razew, Alicja↗
DOE OSTI2021

A robust method of nuclei isolation for single-cell RNA sequencing of solid tissues from the plant genus Populus

Single-cell transcriptome analysis has been extensively applied in humans and animal models to uncover gene expression heterogeneity between the different cell types of a tissue or an organ. It demonstrated its capability to discover key regulatory elements that determine cell fate during developmental programs. Single-cell analysis requires the isolation and labeling of the messenger RNA (mRNA) derived from each cell. These challenges were primarily addressed in mammals by developing microfluidic-based approaches. For plant species whose cells contain cell walls, these approaches have generally required the generation of isolated protoplasts. Many plant tissues’ secondary cell wall hinders enzymatic digestion required for individual protoplast isolation, resulting in an unequal representation of cell types in a protoplast population. This limitation is especially critical for cell types located in the inner layers of a tissue or the inner tissues of an organ. Consequently, single-cell RNA sequencing (scRNA-seq) studies using microfluidic approaches in plants have mainly been restricted to Arabidopsis roots, for which well-established procedures of protoplast isolation are available. Here we present a simple alternative approach to generating high-quality protoplasts from plant tissue by characterizing the mRNA extracted from individual nuclei instead of whole cells. We developed the protocol using two different plant materials with varying cellular complexity levels and cell wall structure, Populus shoot apices, and more lignified stems. Using the 10× Genomics Chromium technology, we show that this procedure results in intact mRNA isolation and limited leakage, with a broad representation of individual cell transcriptomes.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2021

Comparison of methodologies used to determine aromatic lignin unit ratios in lignocellulosic biomass

Abstract Background Multiple analytical methods have been developed to determine the ratios of aromatic lignin units, particularly the syringyl/guaiacyl (S/G) ratio, of lignin biopolymers in plant cell walls. Chemical degradation methods such as thioacidolysis produce aromatic lignin units that are released from certain linkages and may induce chemical changes rendering it difficult to distinguish and determine the source of specific aromatic lignin units released, as is the case with nitrobenzene oxidation methodology. NMR methods provide powerful tools used to analyze cell walls for lignin composition and linkage information. Pyrolysis-mass spectrometry methods are also widely used, particularly as high-throughput methodologies. However, the different techniques used to analyze aromatic lignin unit ratios frequently yield different results within and across particular studies, making it difficult to interpret and compare results. This also makes it difficult to obtain meaningful insights relating these measurements to other characteristics of plant cell walls that may impact biomass sustainability and conversion metrics for the production of bio-derived fuels and chemicals. Results The authors compared the S/G lignin unit ratios obtained from thioacidolysis, pyrolysis-molecular beam mass spectrometry (py-MBMS), HSQC liquid-state NMR and solid-state (ss) NMR methodologies of pine, several genotypes of poplar, and corn stover biomass. An underutilized approach to deconvolute ssNMR spectra was implemented to derive S/G ratios. The S/G ratios obtained for the samples did not agree across the different methods, but trends were similar with the most agreement among the py-MBMS, HSQC NMR and deconvoluted ssNMR methods. The relationship between S/G, thioacidolysis yields, and linkage analysis determined by HSQC is also addressed. Conclusions This work demonstrates that different methods using chemical, thermal, and non-destructive NMR techniques to determine native lignin S/G ratios in plant cell walls may yield different results depending on species and linkage abundances. Spectral deconvolution can be applied to many hardwoods with lignin dominated by S and G units, but the results may not be reliable for some woody and grassy species of more diverse lignin composition. HSQC may be a better method for analyzing lignin in those species given the wealth of information provided on additional aromatic moieties and bond linkages. Additionally, trends or correlations in lignin characteristics such as S/G ratios and lignin linkages within the same species such as poplar may not necessarily exhibit the same trends or correlations made across different biomass types. Careful consideration is required when choosing a method to measure S/G ratios and the benefits and shortcomings of each method discussed here are summarized.

09 BIOMASS FUELS↗
DOE OSTI2025

UDP-glucuronic acid decarboxylase in alfalfa: a target to improve ruminal digestibility of stems

Alfalfa (Medicago sativa) has a high nutritional value, but poor digestibility of the stems limits its value as an energy source in ruminant diets. Xylan and lignin negatively affect cell wall digestibility, whereas pectins have high digestibility in the rumen. In plants, UDP-xylose synthase (UXS) catalyses the decarboxylation of UDP-glucuronic acid to form UDP-xylose in an irreversible step that is key for xylan synthesis. Here, we functionally characterized two UXS genes in alfalfa, namely MsaUXS2 and MsaUXS4, and investigated their impact on ruminal digestibility. Both genes are more highly expressed in stems than leaves, and the enzymes have UDP-glucuronic acid decarboxylase activity in vitro. Silencing of MsaUXS2 and MsaUXS4 via RNAi altered plant growth and resulted in a 40% decrease in xylose, a 115% increase in arabinose, and a 60% increase in galacturonic acid in the polysaccharide matrix as well as a 20% decrease in lignin in the cell wall. Together, our results show a major role for UXS2 and UXS4 in xylan synthesis and secondary cell wall deposition in alfalfa. Additionally, in vitro rumen digestibility assays for the silenced lines had on average 30% increased gas production at 24 h, demonstrating the potential of targeting UXS genes to increase stem digestibility.

UDP-xylose synthase↗
DOE OSTI2020

Chemical and Structural Changes in Corn Stover After Ensiling: Influence on Bioconversion

Production of biofuels, bioproducts, and bioenergy requires a well-characterized, stable, and reasonably uniform biomass supply and well-established supply chains for shipping biomass from farm fields to biorefineries, while achieving year-round production targets. Preserving and stabilizing biomass feedstock during storage is a necessity for cost-effective and sustainable biofuel production. Ensiling is a common storage method used to preserve and even improve forage quality; however, the impact of ensiling on biomass physical and chemical properties that influence bioconversion processes has been variable. Our objective in this work was to determine the effects of ensiling on lignocellulosic feedstock physicochemical properties and how that influences bioconversion requirements. We observed statistically significant decreases (p < 0.05) in the content of two major structural carbohydrates (glucan and xylan) of 5 and 8%, respectively, between the ensiled and non-ensiled materials. We were unable to detect differences in sugar yields from structural carbohydrates after pretreatment and enzymatic hydrolysis of the ensiled materials compared to non-ensiled controls. Based on this work, we conclude that ensiling the corn stover did not change the bioconversion requirements compared to the control samples and incurred losses of structural carbohydrates. At the light microscopy level, ensiled corn stover exhibited little structural change or relocation of cell wall components as detected by immunocytochemistry. However, more subtle structural changes were revealed by electron microscopy, as ensiled cell walls exhibit ultrastructural characteristics such as wall delimitation intermediate between non-ensiled and dilute-acid-pretreated cell walls. These findings suggest that alternative methods of conversion, such as deacetylation and mechanical refining, could take advantage of lamellar defects and may be more effective than dilute acid or hot water pretreatment for biomass conversion of ensiled materials.

09 BIOMASS FUELS↗