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At least 73 records · Page 4

Wall extensibility: its nature, measurement and relationship to plant cell growth

Expansive growth of plant cells is controlled principally by processes that loosen the wall and enable it to expand irreversibly. The central role of wall relaxation for cell expansion is reviewed. The most common methods for assessing the extension properties of plant cell walls ( wall extensibility') are described, categorized and assessed critically. What emerges are three fundamentally different approaches which test growing cells for their ability (a) to enlarge at different values of turgor, (b) to induce wall relaxation, and (c) to deform elastically or plastically in response to an applied tensile force. Analogous methods with isolated walls are similarly reviewed. The results of these different assays are related to the nature of plant cell growth and pertinent biophysical theory. I argue that the extensibilities' measured by these assays are fundamentally different from one another and that some are more pertinent to growth than others.

Review, Academic

Conserved Macromolecular Architecture of Poplar Secondary Cell Walls Revealed by ssNMR and Atomistic Modeling

The macromolecular architecture of plant secondary cell walls governs wood's mechanical and biochemical properties, yet its natural intra-species variability remains poorly characterized. Here, we combined 13C solid-state NMR (ssNMR), multivariate statistical analysis, and molecular modeling to profile nanoscale structure across 13 genetically diverse Populus trichocarpa genotypes grown in 13C-enriched atmospheres. SsNMR-derived phenotypes spanning composition, structure, mobility, and inter-polymer proximities reveal a conserved architecture, with a subtle yet coordinated variation organizing into dominant structural and secondary mobility axes. A representative atomistic model captures these features and reproduces experimental metrics. Molecular dynamics simulations support a weak but consistent positive correlation between cellulose abundance and crystalline-like order, with interior cellulose chains enriched in tg (trans-gauche) conformations without expanding crystalline cores. Together, experiment and simulation reveal a genetically buffered, broadly conserved nanoscale architecture across genotypes, where subtle fine-tuning of cellulose bundling and matrix packing balances mechanical performance with biological function.

09 BIOMASS FUELS

Center for Lignocellulose Structure and Formation (CLSF)

Lignocellulose is the carbon- and energy-rich material synthesized by plants when they make secondary cell walls, e.g. in woody tissues. Plants annually convert gigatons of atmospheric CO 2 into these complex and useful biomaterials that comprise over half of all biological carbon on Earth. This biological conversion helps to reduce atmospheric CO 2 that contributes to climate change while generating renewable plant cell walls that are used commercially on a massive scale in the paper, timber and textile industries. Recent technical developments open new possibilities for use of modified wood and cellulose fibers as large-scale alternatives to steel, plastics, and other nonrenewable materials with high carbon footprints and energy inputs, as well as for generation of biofuels and bio-hydrogen. Despite the utility of plant-based biomaterials, many physical and biological aspects of cell wall structure and formation remain obscure, and this limits the current scope for engineering these renewable resources for greater utility.

09 BIOMASS FUELS

Ultrastructure of potato tubers formed in microgravity under controlled environmental conditions

Previous spaceflight reports attribute changes in plant ultrastructure to microgravity, but it was thought that the changes might result from growth in uncontrolled environments during spaceflight. To test this possibility, potato explants were examined (a leaf, axillary bud, and small stem segment) grown in the ASTROCULTURETM plant growth unit, which provided a controlled environment. During the 16 d flight of space shuttle Columbia (STS-73), the axillary bud of each explant developed into a mature tuber. Upon return to Earth, tuber slices were examined by transmission electron microscopy. Results showed that the cell ultrastructure of flight-grown tubers could not be distinguished from that of tuber cells grown in the same growth unit on the ground. No differences were observed in cellular features such as protein crystals, plastids with starch grains, mitochondria, rough ER, or plasmodesmata. Cell wall structure, including underlying microtubules, was typical of ground-grown plants. Because cell walls of tubers formed in space were not required to provide support against the force due to gravity, it was hypothesized that these walls might exhibit differences in wall components as compared with walls formed in Earth-grown tubers. Wall components were immunolocalized at the TEM level using monoclonal antibodies JIM 5 and JIM 7, which recognize epitopes of pectins, molecules thought to contribute to wall rigidity and cell adhesion. No difference in presence, abundance or distribution of these pectin epitopes was seen between space- and Earth-grown tubers. This evidence indicates that for the parameters studied, microgravity does not affect the cellular structure of plants grown under controlled environmental conditions.

Non-NASA Center

The DYRKP1 kinase regulates cell wall degradation in Chlamydomonas by inducing matrix metalloproteinase expression

Abstract The cell wall of plants and algae is an important cell structure that protects cells from changes in the external physical and chemical environment. This extracellular matrix, composed of polysaccharides and glycoproteins, must be constantly remodeled throughout the life cycle. However, compared to matrix polysaccharides, little is known about the mechanisms regulating the formation and degradation of matrix glycoproteins. We report here that a plant kinase belonging to the dual-specificity tyrosine phosphorylation-regulated kinase (DYRKP1) family present in all eukaryotes regulates cell wall degradation after mitosis of Chlamydomonas reinhardtii by inducing the expression of matrix metalloproteinases. Without DYRKP1, daughter cells cannot disassemble parental cell walls and remain trapped inside for more than 10 days. On the other hand, the dual-specificity tyrosine phosphorylation-regulated kinase complementation lines show normal degradation of the parental cell wall. Transcriptomic and proteomic analyses indicate a marked downregulation of MMP gene expression and accumulation, respectively, in the dyrkp1 mutants. The mutants deficient in matrix metalloproteinases retain palmelloid structures for a longer time than the background strain, like dyrkp1 mutants. Our findings show that dual-specificity tyrosine phosphorylation-regulated kinase, by ensuring timely MMP expression, enables the successful execution of the cell cycle. Altogether, this study provides insight into the life cycle regulation in plants and algae.

Kim, Minjae (ORCID:0000000223561295)

Native Architecture of Wheat Straw Cell Walls: A Unified Model from X-ray Scattering and Solid-State NMR

Plant secondary cell walls constitute the dominant reservoir of renewable biomass, comprising tightly packed cellulose, hemicellulose, and lignin at the nanoscale. Recent advances in solid-state NMR spectroscopy and the availability of small-angle X-ray scattering for biomass characterization have led to an accumulation of experimental data on cell wall organization, yet no explicit structure model has simultaneously satisfied both Xray and NMR observations. Using wheat straw as a model system, we propose a structural framework consistent with current knowledge of cellulose biosynthesis, X-ray scattering data, and one- and two-dimensional 13 C solid-state NMR spectra. In this model, 18-chain elementary fibrils align in parallel and populate the cross-section at random. Arabinose-substituted xylan shows no conformational dependence for cellulose-binding in wheat, and only a minor fraction of 2-fold xylan appears in close proximity to cellulose, unlike in Arabidopsis, where xylan is more tightly attached to the cellulose surface. While NMR data cannot unambiguously resolve the internal arrangement of the 18 glucan chains, X-ray scattering profiles uniquely constrain the fibril size and exclude the possibility of tight bundling in the intact walls. The specific interaction between the matrix polymers and the cellulose elementary fibrils must be reconsidered in light of the small interfibril spaces, which bring the matrix components into spatial proximity with cellulose even in the absence of attractive interactions. These findings provide fundamental molecular-level insight into cellulose fibril architecture and matrix−polymer interactions, resolving longstanding discrepancies between spectroscopic and scattering data and advancing our understanding of biopolymer assembly into structurally and functionally versatile lignocellulosic biomaterials.

Carbohydrates

Exploring the binding properties and activities of ancestral expansins

Bacterial expansins are non-lytic proteins capable of loosening cellulose networks, offering promising applications in agriculture, biotechnology, and material science. Their ability to disrupt noncovalent interactions in biopolymer matrices such as cellulose and chitin positions them as valuable tools for upgrading abundant natural materials. However, their industrial use remains limited due to their relatively low wall-loosening activity compared to plant expansins. To address this limitation, we applied Ancestral Sequence Resurrection (ASR) to reconstruct and characterize ancient variants of the Bacillus subtilis expansin BsEXLX1. ASR is a powerful evolutionary tool that enables the inference and synthesis of ancestral proteins, allowing researchers to explore functional traits that may have been lost over time. This approach not only provides insights into protein evolution but also facilitates the design of proteins with enhanced properties, such as improved substrate affinity or structural stability. In this study, we combined biochemical and biophysical assays to evaluate the activity and binding behavior of ancestral expansins. Our results reveal that ancestral variants exhibit increased cellulose affinity, reduced binding to acidic polysaccharides, and greater salt resistance. Furthermore, these traits enhance their wall-loosening activity and demonstrate the utility of ASR in engineering surface-active proteins for industrial applications, particularly in biomass processing and cellulose modification.

09 BIOMASS FUELS

Investigating the Relationship between the Cell Wall Integrity Pathway and Unfolded Protein Response

Plants have made significant contributions to astronaut health in spaceflight missions. To further spaceflight research in optimizing plant viability, this study aims to understand the factors involved in maintaining cell wall integrity, which is vital to plant morphology and structural stability. Spaceflight can negatively impact the cell wall; thus, it is crucial to investigate how to mitigate spaceflight stressors to maintain the integrity of the cell wall. The structural integrity of plants’ cell walls depends on secondary cell wall biogenesis, which enables the repair and architectural support of plants like A. thaliana. This biogenesis is triggered by a signal transduction cascade: first initiated by cell wall stress, the CWI (cell wall integrity) pathway is activated, followed by the UPR (unfolded protein response), then the cell wall’s integrity is maintained through secondary cell wall biogenesis. Through a re-analysis of GeneLab Dataset 321 (GLDS-321), a study from NASA’s Open Science Data Repository that investigates the effects of spaceflight on the UPR, several genes were found to be associated with the cell wall. This proposal postulates a relationship between the UPR and the CWI pathway and their direct effect on secondary cell wall biogenesis by investigating IRX7, a gene associated with secondary cell wall biogenesis. The predicted outcome of overexpressing IRX7 is increased resilience of the cell wall by upregulating both the UPR and the CWI pathway, while silencing IRX7 is predicted to compromise the cell wall integrity by downregulating the UPR and the CWI pathway. This study will give insight into the needed measures to increase cell wall resilience in stressful environments: As spaceflight durations increase and uncertain climate change events progress on Earth, understanding how to optimize cell wall resilience – a fundamental pillar of plant health – can effectively enhance mass crop production and quality and ensure the physical and psychological health of astronauts in long-term space missions.

GL4HS

Isolation and characterization of beta-glucan synthase: A potential biochemical regulator of gravistimulated differential cell wall loosening

In plants, gravity stimulates differential growth in the upper and lower halves of horizontally oriented organs. Auxin regulation of cell wall loosening and elongation is the basis for most models of this phenomenon. Auxin treatment of pea stem tissue rapidly increases the activity of Golgi-localized Beta-1,4-glucan synthase, an enzyme involved in biosynthesis of wall xyloglucan which apparently constitutes the substrate for the wall loosening process. The primary objective is to determine if auxin induces de novo formation of Golgi glucan synthase and increases the level of this glucan synthase mRNA. This shall be accomplished by (a) preparation of a monoclonal antibody to the synthase, (b) isolation, and characterization of the glucan synthase, and (c) examination for cross reactivity between the antibody and translation products of auxin induced mRNAs in pea tissue. The antibody will also be used to localize the glucan synthase in upper and lower halves of pea stem tissue before, during and after the response to gravity.

Kuzmanoff, K. M.

A novel xylosylated fucoglucuronan in Penium reveals structural parallels to rhamnogalacturonan-I and its broad evolutionary footprint in lower plants

Green algae inhabit aquatic environments across the planet and play a crucial role in sustaining the global ecosystem. Ancestors of some Charophytes adapted to terrestrial conditions and eventually evolved into land plants. Extant green algae have inherited traits from their ancestors and evolved into their current morphological and chemical forms, as reflected by their cell walls with distinct shapes and compositions. To illuminate the evolution of plant cell walls and bridge the gap between green algae and land plants, we investigated the charophyte Penium margaritaceum, a close relative of terrestrial plants. We discovered a previously unknown polysaccharide in both its culture medium and cell wall. This polysaccharide, termed xylosylated fucoglucuronan (XFG), possesses a rhamnogalacturonan-I (RG-I)-like backbone composed of repeating [-3-α-Fucp-(1,4)-α-GlcpA-] disaccharides that are extensively xylosylated and acetylated. Surveying approximately 20 non-vascular plants revealed that XFG and RG-I (or related structures) first emerge in certain Chlorophyceae and subsequently co-occur throughout lineages along the evolutionary trajectory to bryophytes, thereby bridging aquatic green algae to early land plants. The striking structural parallels between XFG, RG-I, and ulvan suggest a shared evolutionary origin, offering new insight into how plant cell walls adapted during the transition from marine to freshwater environments and ultimately to land.

Algae

Water uptake by growing cells: an assessment of the controlling roles of wall relaxation, solute uptake, and hydraulic conductance

Growing plant cells increase in volume principally by water uptake into the vacuole. There are only three general mechanisms by which a cell can modulate the process of water uptake: (a) by relaxing wall stress to reduce cell turgor pressure (thereby reducing cell water potential), (b) by modifying the solute content of the cell or its surroundings (likewise affecting water potential), and (c) by changing the hydraulic conductance of the water uptake pathway (this works only for cells remote from water potential equilibrium). Recent studies supporting each of these potential mechanisms are reviewed and critically assessed. The importance of solute uptake and hydraulic conductance is advocated by some recent studies, but the evidence is indirect and conclusions remain controversial. For most growing plant cells with substantial turgor pressure, it appears that reduction in cell turgor pressure, as a consequence of wall relaxation, serves as the major initiator and control point for plant cell enlargement. Two views of wall relaxation as a viscoelastic or a chemorheological process are compared and distinguished.

Non-NASA Center

Effect of microgravity on plant growth

The overall goal of this research is to determine the effect of microgravity proper on plant growth (metabolism and cell wall formation). In addressing this goal, the work conducted during this grant period was divided into three components: analyses of various plant tissues previously grown in space aboard MIR Space Station; analyses of wheat tissues grown on Shuttle flight STS-51; and Phenylpropanoid metabolism and plant cell wall synthesis (earth-based investigations).

Lewis, Norman G.

Enhancing monolignol ferulate conjugate levels in poplar lignin via OsFMT1

The phenolic polymer lignin is one of the primary chemical constituents of the plant secondary cell wall. Due to the inherent plasticity of lignin biosynthesis, several phenolic monomers have been shown to be incorporated into the polymer, as long as the monomer can undergo radicalization so it can participate in coupling reactions. In this study, we significantly enhance the level of incorporation of monolignol ferulate conjugates into the lignin polymer to improve the digestibility of lignocellulosic biomass. Overexpression of a rice Feruloyl-CoA Monolignol Transferase (FMT), OsFMT1, in hybrid poplar (Populus alba x grandidentata) produced transgenic trees clearly displaying increased cell wall-bound ester-linked ferulate, p-hydroxybenzoate, and p-coumarate, all of which are in the lignin cell wall fraction, as shown by NMR and DFRC. We also demonstrate the use of a novel UV–Vis spectroscopic technique to rapidly screen plants for the presence of both ferulate and p-hydroxybenzoate esters. Lastly we show, via saccharification assays, that the OsFMT1 transgenic poplars have significantly improved processing efficiency compared to wild-type and Angelica sinensis-FMT-expressing poplars. The findings demonstrate that OsFMT1 has a broad substrate specificity and a higher catalytic efficiency compared to the previously published FMT from Angelica sinensis (AsFMT). Importantly, enhanced wood processability makes OsFMT1 a promising gene to optimize the composition of lignocellulosic biomass.

09 BIOMASS FUELS

Gravitropism in plants: Hydraulics and wall growth properties of responding cells

Gravitropism is the asymmetrical alteration of plant growth in response to a change in the gravity vector, with the typical result that stems grow up and roots grow down. The gravity response is important for plants because it enables them to grow their aerial parts in a mechanically stable (upright) position and to develop their roots and leaves to make efficient use of soil nutrients and sunlight. The elucidation of gravitropic responses will tell much about how gravity exerts its morphogenetic effects on plants and how plants regulate their growth at the cellular and molecular levels.

Cosgrove, Daniel J.

Engineering the green algae Chlamydomonas incerta for recombinant protein production

Chlamydomonas incerta , a genetically close relative of the model green alga Chlamydomonas reinhardtii , shows significant potential as a host for recombinant protein expression. Because of the close genetic relationship between C. incerta and C. reinhardtii , this species offers an additional reference point for advancing our understanding of photosynthetic organisms, and also provides a potential new candidate for biotechnological applications. This study investigates C. incerta ’s capacity to express three recombinant proteins: the fluorescent protein mCherry, the hemicellulose-degrading enzyme xylanase, and the plastic-degrading enzyme PHL7. We have also examined the capacity to target protein expression to various cellular compartments in this alga, including the cytosol, secretory pathway, cytoplasmic membrane, and cell wall. When compared directly with C. reinhardtii , C. incerta exhibited a distinct but notable capacity for recombinant protein production. Cellular transformation with a vector encoding mCherry revealed that C. incerta produced approximately 3.5 times higher fluorescence levels and a 3.7-fold increase in immunoblot intensity compared to C. reinhardtii . For xylanase expression and secretion, both C. incerta and C. reinhardtii showed similar secretion capacities and enzymatic activities, with comparable xylan degradation rates, highlighting the industrial applicability of xylanase expression in microalgae. Finally, C. incerta showed comparable PHL7 activity levels to C. reinhardtii , as demonstrated by the in vitro degradation of a polyester polyurethane suspension, Impranil® DLN. Finally, we also explored the potential of cellular fusion for the generation of genetic hybrids between C. incerta and C. reinhardtii as a means to enhance phenotypic diversity and augment genetic variation. We were able to generate genetic fusion that could exchange both the recombinant protein genes, as well as associated selectable marker genes into recombinant offspring. These findings emphasize C. incerta ’s potential as a robust platform for recombinant protein production, and as a powerful tool for gaining a better understanding of microalgal biology.

cell membranes

The pectin puzzle: Decoding the fine structure of rhamnogalacturonan-I (RG-I) in Arabidopsis thaliana uncovers new pectin features

Pectin is generally divided into four distinct structural categories, namely homogalacturonan, xylogalacturonan, rhamnogalacturonan I (RG-I) and rhamnogalacturonan II. While much of the structural diversity of homogalacturonan, xylogalacturonan and rhamnogalacturonan II has been elucidated, the structural features of RG-I are less well understood. In this work, we employed multiple complementary analytical techniques to present a detailed structural analysis of RG-I in the model species Arabidopsis thaliana . Starting with highly purified RG-I from different Arabidopsis tissues, we employed comparative linkage and nuclear magnetic resonance analysis along with mass spectrometry analysis of enzymatically digested RG-I oligosaccharides. Besides the presence of the canonical α-1,5-arabinan, β-1,4-galactan, β-1,6-galactan and arabinogalactan RG-I side chains of varying lengths, we show that a large portion of the β-1,6-galactan is terminated by either 4-O-methyl β-glucuronic acid (GlcA) residues or, to a smaller degree, β-GlcA that lacks the Me-ether group. Importantly, O-acetylation of RG-I GalA residues is a minor modification while 10 % of the backbone Rha residues are 3-O-acetylated, and most of the acetylated Rha is additionally branched with β-galactose substituents. Taken together, the combined results of these different analytical techniques present the most comprehensive structural overview of Arabidopsis thaliana RG-I to date.

25 ENERGY STORAGE