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At least 73 records · Page 4

Engineered Production of Hydroxycinnamoyl Tyramine Conjugates Limits the Growth of the Pathogen Pseudomonas syringae in Arabidopsis

Hydroxycinnamoyl tyramine conjugates are phenolamides produced by plants in response to pathogen attack and biotic stresses. Their proposed mechanisms of action include cytotoxicity towards pathogens, cell wall reinforcement to restrict pathogen proliferation, and signaling activity to trigger general stress responses. Here, we engineered the production of the tyramine conjugates p-coumaroyltyramine (CT) and feruloyltyramine (FT) in Arabidopsis to gain insight into their mode of action. Co-expression of feedback-insensitive 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase and tyrosine decarboxylase increased tyramine content. Additional expression of tyramine hydroxycinnamoyltransferase led to de-novo production of CT and FT, which were found as soluble and cell-wall-bound forms. FT was associated with lignin in stems. The growth of pathogenic Pseudomonas syringae was reduced in rosettes of the Arabidopsis CT- and FT-producing lines compared to wild type. These lines also exhibited increased transpirational water loss in excised rosettes. Transcriptomic analysis of transgenic lines grown under normal conditions revealed alterations in the expression of genes associated with the biological circadian clock. These changes led to a reduction in flavonoids and an early flowering phenotype. Important changes in the expression of genes related to abiotic stress such as drought, cold, heat, and hypoxia potentially contribute to reduced growth of P. syringae in engineered Arabidopsis.

Phenolamides↗

Mechanistic Modeling of Creep and Aging for Time Extrapolation in LPBF 316 SS - LANL

A polycrystal constitutive model was extended to quantify the effects heterogeneity in the microstructure, porosity, and internal stresses on the mechanical response (creep, tensile) of 316H steel processed by additive manufacturing. The starting point is a recently developed and fully calibrated advanced microstructuresensitive mechanistic constitutive model fit to literature data pertaining to conventionally processed steels. The model is embedded within the full field fast Fourier transform framework (LApx) and several hundred simulations were performed. Synthetic microstructures with different grain aspect ratios were generated using the Neper package to study the role of grain morphology on the mechanical responses. The effect of initial cell interior dislocation density, cell-wall density, precipitate content, porosity, and internal stresses on creep rates are quantified for different polycrystalline microstructures. The study allowed to prospectively quantify the role of distinct features of the microstructure on the mechanical response under tension and creep loading scenarios. Critically it is found that minor anisotropies in the tensile response of samples, emanating from the aspect ratio of grains correlated into significant differences in creep rate as a function of loading directions. The study also allows to identify initial microstructure porosities below which the materials response only moderately degraded by the initial microstructure.

36 MATERIALS SCIENCE↗

Rapid High-Resolution Analysis of Polysaccharide-Lignin Interactions in Secondary Plant Cell Walls Using Proton-Detected Solid-State NMR

The plant secondary cell wall, a complex matrix composed of cellulose, hemicellulose, and lignin, is crucial for the mechanical strength and water-proofing properties of plant tissues, and serves as a primary source of biomass for biorenewable energy and biomaterials. Structural analysis of these polymers and their interactions within the secondary cell wall has been heavily relying on 13 C-based solid-state NMR techniques. In this study, we explore the application of 1 H-detected solid-state NMR techniques for rapid, high-resolution structural characterization of polysaccharides and lignin, demonstrated on the stems of hardwood eucalyptus. We explored the use of synthesized 2D spectra to resolve central 1 H resonances and the combined application of 3D hCCH and hCHH experiments for complete resonance assignment and unambiguous identification of lignin-carbohydrate interactions. Our findings emphasize the central role of acetylated three-fold xylan conformers, rather than two-fold, in stabilizing the carbohydrate-lignin interface, with glucuronic acid sidechains in eucalyptus glucuronoxylan colocalizing with lignin, revised cellulose-lignin interactions involving uncoated microfibril surfaces, and pectin-lignin interactions indicative of early-stage lignification. These results present a novel approach for rapid structural analysis of lignocellulosic biomaterials without the need for solubilization or extraction.

09 BIOMASS FUELS↗

Growth–defense trade‐offs and yield loss in plants with engineered cell walls

Summary As a major component of plant secondary cell walls, lignin provides structural integrity and rigidity, and contributes to primary defense by providing a physical barrier to pathogen ingress. Genetic modification of lignin biosynthesis has been adopted to reduce the recalcitrance of lignified cell walls to improve biofuel production, tree pulping properties and forage digestibility. However, lignin‐modification is often, but unpredictably, associated with dwarf phenotypes. Hypotheses suggested to explain this include: collapsed vessels leading to defects in water and solute transport; accumulation of molecule(s) that are inhibitory to plant growth or deficiency of metabolites that are critical for plant growth; activation of defense pathways linked to cell wall integrity sensing. However, there is still no commonly accepted underlying mechanism for the growth defects. Here, we discuss recent data on transcriptional reprogramming in plants with modified lignin content and their corresponding suppressor mutants, and evaluate growth‐defense trade‐offs as a factor underlying the growth phenotypes. New approaches will be necessary to estimate how gross changes in transcriptional reprogramming may quantitatively affect growth. Better understanding of the basis for yield drag following cell wall engineering is important for the biotechnological exploitation of plants as factories for fuels and chemicals.

Ha, Chan Man↗

Gram-positive bacterial cell wall components inhibit herpes simplex virus infection

The role of the mucosal microbiome in viral infections remains unclear. Genital herpes, caused by herpes simplex virus 1 and 2 (HSV-1 and HSV-2), is among the most prevalent sexually transmitted infections worldwide. Despite evidence linking vaginal Lactobacillus to protection against sexually transmitted viruses, the specific microbial components and mechanisms that mediate this defense are not well understood. Here, we show that multiple cell wall components from diverse gram-positive bacteria, including lactobacilli, inhibit HSV-1 and HSV-2 infection in cells and in a mouse model of genital herpes infection. Peptidoglycan (PG) and lipoteichoic acid (LTA), both major components of the gram-positive bacterial cell wall, significantly reduced HSV infectivity in vitro and improved survival and disease outcomes in mice. We further showed that Lactobacillus crispatus surface layer proteins SlpA and SlpB bind HSV-1 and inhibit infection. Antiviral effects of cell wall components were dose-dependent, relied on intact PG structure, and, in the case of PG and LTA, were independent of TLR2-mediated host signaling. Collectively, our findings identify a species-independent antiviral function for gram-positive bacterial cell wall components against HSV and suggest that the composition of the mucosal microbiome may play an underappreciated role in suppressing mucosal herpes infection in humans.

Adams↗

Cynipid wasps systematically reprogram host metabolism and restructure cell walls in developing galls

Many insects have evolved the ability to manipulate plant growth to generate extraordinary structures called galls, in which insect larva can develop while being sheltered and feeding on the plant. In particular, cynipid (Hymenoptera: Cynipidae) wasps have evolved to form morphologically complex galls and generate an astonishing array of gall shapes, colors, and sizes. However, the biochemical basis underlying these remarkable cellular and developmental transformations remains poorly understood. A key determinant in plant cellular development is cell wall deposition that dictates the physical form and physiological function of newly developing cells, tissues, and organs. However, it is unclear to what degree cell walls are restructured to initiate and support the formation of new gall tissue. Here, we characterize the molecular alterations underlying gall development using a combination of metabolomic, histological, and biochemical techniques to elucidate how valley oak (Quercus lobata) leaf cells are reprogrammed to form galls. Strikingly, gall development involves an exceptionally coordinated spatial deposition of lignin and xylan to form de novo gall vasculature. Our results highlight how cynipid wasps can radically change the metabolite profile and restructure the cell wall to enable the formation of galls, providing insights into the mechanism of gall induction and the extent to which plants can be entirely reprogrammed to form unique structures and organs.

59 BASIC BIOLOGICAL SCIENCES↗

Bioenergy sorghum stem density increases threefold following internode elongation due to continued accumulation of lignified cell walls and complex regulation of genes involved in cell wall biosynthesis

Bioenergy sorghum is a highly productive drought tolerant C4 grass that accumulates ~ 80% of its harvested biomass in ~ 4 m long stems comprised of > 40 internodes that develop sequentially during an extended vegetative growth phase. Following elongation of each internode, internode density increases ~ threefold to fourfold primarily due to the accumulation of cell walls composed of cellulose, glucuronoarabinoxylan and lignin. Lignin accumulates initially on cell walls of sclerenchyma cells surrounding vascular bundles and later on cell walls of the stem rind and stem core pith parenchyma. Many genes involved in cell wall biosynthesis were expressed continuously during the stem internode densification process whereas others showed dynamic patterns of expression (high to low, low to high). Several CESA genes involved in primary cell wall cellulose synthesis were expressed in the stem rind and core throughout the stem densification phase. In contrast, CESA genes involved in secondary cell wall biogenesis were expressed continuously in the stem rind but downregulated in the stem core shortly after completion of internode elongation. Overall, accumulation of cell wall biomass in elongated internodes during stem densification increases stem mechanical strength and biomass bulk density while modifying biomass composition in ways that could impact the amount and release of cellulosic sugars and lignin-derived bioproducts.

09 BIOMASS FUELS↗

Cycling ferulate in monocot cell walls

Cell walls in grasses contain arabinoxylan polysaccharides decorated with ferulate groups but the mechanism by which the ferulate is attached to arabinoxylans has long remained unknown. Here, a new study shows that ferulate is transferred to arabinoxylan from a 3,6-di-O-feruloyl sucrose intermediate formed in a sucrose ferulate cycle.

59 BASIC BIOLOGICAL SCIENCES↗

Structure-Based Design of Dual Bactericidal and Bacteria-Releasing Nanosurfaces

Here, we report synergistic nanostructured surfaces combining bactericidal and bacteria-releasing properties. A polystyrene-block-poly(methyl methacrylate) (PS-block-PMMA) diblock copolymer is used to fabricate vertically oriented cylindrical PS structures (“PS nanopillars”) on silicon substrates. The results demonstrate that the PS nanopillars (with a height of about 10 nm, size of about 50 nm, and spacing of about 70 nm) exhibit highly effective bactericidal and bacteria-releasing properties (“dual properties”) against Escherichia coli for at least 36 h of immersion in an E. coli solution. Interestingly, the PS nanopillars coated with a thin layer (≈3 nm thick) of titanium oxide (TiO 2 ) (“TiO 2 nanopillars”) show much improved dual properties against E. coli (a Gram-negative bacterium) compared to the PS nanopillars. Moreover, the dual properties emerge against Listeria monocytogenes (a Gram-positive bacterium). To understand the mechanisms underlying the multifaceted property of the nanopillars, coarse-grained molecular dynamics (MD) simulations of a lipid bilayer (as a simplified model for E. coli) in contact with a substrate containing hexagonally packed hydrophilic nanopillars were performed. The MD results demonstrate that when the bacterium–substrate interaction is strong, the lipid heads adsorb onto the nanopillar surfaces, conforming the shape of a lipid bilayer to the structure/curvature of nanopillars and generating high stress concentrations within the membrane (i.e., the driving force for rupture) at the edge of the nanopillars. Membrane rupture begins with the formation of pores between nanopillars (i.e., bactericidal activity) and ultimately leads to the membrane withdrawal from the nanopillar surface (i.e., bacteria-releasing activity). In the case of Gram-positive bacteria, the adhesion area to the pillar surface is limited due to the inherent stiffness of the bacteria, creating higher stress concentrations within a bacterial cell wall. In conclusion, the present study provides insight into the mechanism underlying the “adhesion-mediated” multifaceted property of nanosurfaces, which is crucial for the development of next-generation antibacterial surface coatings for relevant medical applications.

36 MATERIALS SCIENCE↗

Oxidative Deboronation of Boronic Acids by Hydrogen Peroxide in Planta Generates Borate for Cross-Linking of Rhamnogalacturonan II

Vascular plants require boron to cross-link the rhamnogalacturonan-II (RG-II) domain of pectin to form functional cell walls. Boronic acids, which form reversible esters with cis-diols like borate, have been proposed to influence RG-II cross-linking, though the mechanism remains unclear. We used suspension-cultured rose cells adapted to grow without boron to investigate the effect of boronic acids on RG-II dimerization. When grown with phenylboronic acid (PBA) as the sole boron source, nearly all RG-II was crosslinked, whereas methylboronic acid (MBA) only partially restored cross-linking. In contrast, in vitro assays showed that homogeneous RG-II monomers did not dimerize with alkyl or aryl boronic acids unless supplemented with hydrogen peroxide (H2O2), which oxidatively converts boronic acids to boric acid. Real-time NMR spectroscopy and density functional theory calculations provided insight into the reaction mechanism and energetics of oxidation respectively. Together, our data show that exogenous boronic acids are a source of boric acid for plants, and that the deboronation reaction generates aryl or alkyl alcohol byproducts that can undergo further chemical modification in planta. The fate and potential roles of these byproducts in planta remain to be determined.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Dynamic Construction, Perception, and Remodeling of Plant Cell Walls

Plant cell walls are dynamic structures that are synthesized by plants to provide durable coverings for the delicate cells they encase. They are made of polysaccharides, proteins, and other biomolecules and have evolved to withstand large amounts of physical force and to resist external attack by herbivores and pathogens but can in many cases expand, contract, and undergo controlled degradation and reconstruction to facilitate developmental transitions and regulate plant physiology and reproduction. Recent advances in genetics, microscopy, biochemistry, structural biology, and physical characterization methods have revealed a diverse set of mechanisms by which plant cells dynamically monitor and regulate the composition and architecture of their cell walls, but much remains to be discovered about how the nanoscale assembly of these remarkable structures underpins the majestic forms and vital ecological functions achieved by plants.

biofuels (including algae and biomass), bio-inspir↗

Outstanding questions on xylan biosynthesis

Xylan is the second most abundant polysaccharide in plant biomass. It is a crucial component of cell wall structure as well as a significant factor contributing to biomass recalcitrance. Xylan consists of a linear chain of β-1,4-linked xylosyl residues that are often substituted with glycosyl side chains, such as glucuronosyl/methylglucuronosyl and arabinofuranosyl residues, and acetylated at O-2 and/or O-3. Xylan from gymnosperms and dicots contains a unique reducing end tetrasaccharide sequence that is not detected in xylan from grasses, bryophytes and seedless vascular plants. Grass xylan is heavily decorated at O-3 with arabinofuranosyl residues that are frequently esterified with hydroxycinnamates. Further, genetic and biochemical studies have uncovered a number of genes involved in xylan backbone elongation and acetylation, xylan glycosyl substitutions and their modifications, and the synthesis of the unique xylan reducing end tetrasaccharide sequence, but some outstanding issues on the biosynthesis of xylan still remain unanswered. Here, we provide a brief overview of xylan structure and focus on discussion of the current understanding and open questions on xylan biosynthesis. Further elucidation of the biochemical mechanisms underlying xylan biosynthesis will not only shed new insights into cell wall biology but also provide molecular tools for genetic modification of biomass composition tailored for diverse end uses.

59 BASIC BIOLOGICAL SCIENCES↗

The Inducible Accumulation of Cell Wall-Bound p -Hydroxybenzoates Is Involved in the Regulation of Gravitropic Response of Poplar

Lignin in Populus species is acylated with p-hydroxybenzoate. Monolignol p-hydroxybenzoyltransferase 1 (PHBMT1) mediates p-hydroxybenzoylation of sinapyl alcohol, eventually leading to the modification of syringyl lignin subunits. Angiosperm trees upon gravistimulation undergo the re-orientation of their growth along with the production of specialized secondary xylem, i.e., tension wood (TW), that generates tensile force to pull the inclined stem or leaning branch upward. Sporadic evidence suggests that angiosperm TW contains relatively a high percentage of syringyl lignin and lignin-bound p-hydroxybenzoate. However, whether such lignin modification plays a role in gravitropic response remains unclear. By imposing mechanical bending and/or gravitropic stimuli to the hybrid aspens in the wild type (WT), lignin p-hydroxybenzoate deficient, and p-hydroxybenzoate overproduction plants, we examined the responses of plants to gravitropic/mechanical stress and their cell wall composition changes. We revealed that mechanical bending or gravitropic stimulation not only induced the overproduction of crystalline cellulose fibers and increased the relative abundance of syringyl lignin, but also significantly induced the expression of PHBMT1 and the increased accumulation of p-hydroxybenzoates in TW. Furthermore, we found that although disturbing lignin-bound p-hydroxybenzoate accumulation in the PHBMT1 knockout and overexpression (OE) poplars did not affect the major chemical composition shifts of the cell walls in their TW as occurred in the WT plants, depletion of p-hydroxybenzoates intensified the gravitropic curving of the plantlets in response to gravistimulation, evident with the enhanced stem secant bending angle. By contrast, hyperaccumulation of p-hydroxybenzoates mitigated gravitropic response. These data suggest that PHBMT1-mediated lignin modification is involved in the regulation of poplar gravitropic response and, likely by compromising gravitropism and/or enhancing autotropism, negatively coordinates the action of TW cellulose fibers to control the poplar wood deformation and plant growth.

59 BASIC BIOLOGICAL SCIENCES↗

Structures of the mycobacterial membrane protein MmpL3 reveal its mechanism of lipid transport

The mycobacterial membrane protein large 3 (MmpL3) transporter is essential and required for shuttling the lipid trehalose monomycolate (TMM), a precursor of mycolic acid (MA)-containing trehalose dimycolate (TDM) and mycolyl arabinogalactan peptidoglycan (mAGP), in Mycobacterium species, including Mycobacterium tuberculosis and Mycobacterium smegmatis. However, the mechanism that MmpL3 uses to facilitate the transport of fatty acids and lipidic elements to the mycobacterial cell wall remains elusive. Here, we report 7 structures of the M. smegmatis MmpL3 transporter in its unbound state and in complex with trehalose 6-decanoate (T6D) or TMM using single-particle cryo-electron microscopy (cryo-EM) and X-ray crystallography. Combined with calculated results from molecular dynamics (MD) and target MD simulations, we reveal a lipid transport mechanism that involves a coupled movement of the periplasmic domain and transmembrane helices of the MmpL3 transporter that facilitates the shuttling of lipids to the mycobacterial cell wall.

60 APPLIED LIFE SCIENCES↗

Localized growth and remodelling drives spongy mesophyll morphogenesis

The spongy mesophyll is a complex, porous tissue found in plant leaves that enables carbon capture and provides mechanical stability. Unlike many other biological tissues, which remain confluent throughout development, the spongy mesophyll must develop from an initially confluent tissue into a tortuous network of cells with a large proportion of intercellular airspace. How the airspace in the spongy mesophyll develops while the tissue remains mechanically stable is unknown. Here, we use computer simulations of deformable polygons to develop a purely mechanical model for the development of the spongy mesophyll tissue. By stipulating that cell wall growth and remodelling occurs only near void space, our computational model is able to recapitulate spongy mesophyll development observed in Arabidopsis thaliana leaves. We find that robust generation of pore space in the spongy mesophyll requires a balance of cell growth, adhesion, stiffness and tissue pressure to ensure cell networks become porous yet maintain mechanical stability. The success of this mechanical model of morphogenesis suggests that simple physical principles can coordinate and drive the development of complex plant tissues like the spongy mesophyll.

Science & Technology - Other Topics↗

The Effect of Acetylation on Iron Uptake and Diffusion in Water Saturated Wood Cell Walls and Implications for Decay

Acetylation is widely used as a wood modification process that protects wood from fungal decay. The mechanisms by which acetylation protects wood are not fully understood. With these experiments, we expand upon the literature and test whether previously observed differences in iron uptake by wood were a result of decreased iron binding capacity or slower diffusion. We measured the concentration of iron in 2 mm thick wood sections at 0, 10, and 20% acetylation as a function of time after exposure to iron solutions. The iron was introduced either strongly chelated with oxalate or weakly chelated with acetate. The concentrations of iron and oxalate in solution were chosen to be similar to those found during brown rot decay, while the concentration of iron and acetate matched previous work. The iron content of oxalate-exposed wood increased only slightly and was complete within an hour, suggesting little absorption and fast diffusion, or only slight surface adsorption. The increase in iron concentration from acetate solutions with time was consistent with Fickian diffusion, with a diffusion coefficient on the order of 10 -16 m 2 s -1 . The rather slow diffusion rate was likely due to significant binding of iron within the wood cell wall. The diffusion coefficient did not depend on the acetylation level; however, the capacity for iron absorption from acetate solution was greatly reduced in the acetylated wood, likely due to the loss of OH groups. We explored several hypotheses that might explain why the diffusion rate appears to be independent of the acetylation level and found none of them convincing. Implications for brown rot decay mechanisms and future research are discussed.

54 ENVIRONMENTAL SCIENCES↗

Protocols for isolating and characterizing polysaccharides from plant cell walls: a case study using rhamnogalacturonan-II

Abstract Background In plants, a large diversity of polysaccharides comprise the cell wall. Each major type of plant cell wall polysaccharide, including cellulose, hemicellulose, and pectin, has distinct structures and functions that contribute to wall mechanics and influence plant morphogenesis. In recent years, pectin valorization has attracted much attention due to its expanding roles in biomass deconstruction, food and material science, and environmental remediation. However, pectin utilization has been limited by our incomplete knowledge of its structure. Herein, we present a workflow of principles relevant for the characterization of polysaccharide primary structure using nature’s most complex polysaccharide, rhamnogalacturonan-II (RG-II), as a model. Results We outline how to isolate RG-II from celery and duckweed cell walls and from red wine using chemical or enzymatic treatments coupled with size-exclusion chromatography. From there, we applied mass spectrometry (MS)-based techniques to determine the glycosyl residue and linkage compositions of the intact RG-II and derived oligosaccharides including special considerations for labile monosaccharides. In doing so, we demonstrated that in the duckweed Wolffiella repanda the arabinopyranosyl (Ara p ) residue of side chain B is substituted at O- 2 with rhamnose. We used electrospray-MS techniques to identify non-glycosyl modifications including methyl-ethers, methyl-esters, and acetyl-esters on RG-II-derived oligosaccharides. We then showed the utility of proton nuclear magnetic resonance spectroscopy ( 1 H-NMR) to investigate the structure of intact RG-II and to complement the RG-II dimerization studies performed using size-exclusion chromatography. Conclusions The complexity of pectic polysaccharide structures has hampered efforts aimed at their valorization. In this work, we used RG-II as a model to demonstrate the steps necessary to isolate and characterize polysaccharides using chromatographic, MS, and NMR techniques. The principles can be applied to the characterization of other saccharide structures and will help inform researchers on how saccharide structure relates to functional properties in the future.

09 BIOMASS FUELS↗

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↗