Biochemical and structural characterization of glycosyltransferase family 61 proteins reveal a key determinant of sugar donor specificity.

Xylan, the most abundant non-cellulosic polymer in plant cell walls, is structurally diverse, especially in grasses where it is heavily substituted with arabinofuranose and further modified by various residues. Common substitutions across species include glucuronic and 4- -methyl-glucuronic acid. Arabinose and xylose sidechains are synthesized by glycosyltransferase family 61 (GT61) proteins, many of which remain uncharacterized in plants, with limited structural and mechanistic understanding.

EcoFAB 3.0: a sterile system for studying sorghum that replicates previous field and greenhouse observations.

Introduction: Studying plant-microbe interactions is one of the key elements in understanding the path to sustainable agricultural practices. These interactions play a crucial role in ensuring survival of healthy plants, soil and microbial communities. Many platforms have been developed over the years to isolate these highly complex interactions however, these are designed for small model plants.

Engineered reduction of S-adenosylmethionine alters lignin in sorghum.

Background: Lignin is an aromatic polymer deposited in secondary cell walls of higher plants to provide strength, rigidity, and hydrophobicity to vascular tissues. Due to its interconnections with cell wall polysaccharides, lignin plays important roles during plant growth and defense, but also has a negative impact on industrial processes aimed at obtaining monosaccharides from plant biomass. Engineering lignin offers a solution to this issue.

Unraveling plant-microbe symbioses using single-cell and spatial transcriptomics.

Plant-microbe symbioses require intense interaction and genetic coordination to successfully establish in specific cell types of the host and symbiont. Traditional RNA-seq methodologies lack the cellular resolution to fully capture these complexities, but single-cell and spatial transcriptomics (ST) are now allowing scientists to probe symbiotic interactions at an unprecedented level of detail. Here, we discuss the advantages that novel spatial and single-cell transcriptomic technologies provide in studying plant-microbe endosymbioses and highlight key recent studies.

Defined synthetic microbial communities colonize and benefit field-grown sorghum.

The rhizosphere constitutes a dynamic interface between plant hosts and their associated microbial communities. Despite the acknowledged potential for enhancing plant fitness by manipulating the rhizosphere, the engineering of the rhizosphere microbiome through inoculation has posed significant challenges. These challenges are thought to arise from the competitive microbial ecosystem where introduced microbes must survive, and the absence of adaptation to the specific metabolic and environmental demands of the rhizosphere.

Advances in Engineering Nucleotide Sugar Metabolism for Natural Product Glycosylation in .

Glycosylation is a ubiquitous modification present across all of biology, affecting many things such as physicochemical properties, cellular recognition, subcellular localization, and immunogenicity. Nucleotide sugars are important precursors needed to study glycosylation and produce glycosylated products. is a potentially powerful platform for producing glycosylated biomolecules, but it lacks nucleotide sugar diversity.

Expression of dehydroshikimate dehydratase in poplar induces transcriptional and metabolic changes in the phenylpropanoid pathway.

Modification of lignin in feedstocks via genetic engineering aims to reduce biomass recalcitrance to facilitate efficient conversion processes. These improvements can be achieved by expressing exogenous enzymes that interfere with native biosynthetic pathways responsible for the production of the lignin precursors. In planta expression of a bacterial 3-dehydroshikimate dehydratase in poplar trees reduced lignin content and altered the monomer composition, which enabled higher yields of sugars after cell wall polysaccharide hydrolysis.

Putative rhamnogalacturonan-II glycosyltransferase identified through callus gene editing which bypasses embryo lethality.

Rhamnogalacturonan II (RG-II) is a structurally complex and conserved domain of the pectin present in the primary cell walls of vascular plants. Borate cross-linking of RG-II is required for plants to grow and develop normally. Mutations that alter RG-II structure also affect cross-linking and are lethal or severely impair growth.

Spatial co-transcriptomics reveals discrete stages of the arbuscular mycorrhizal symbiosis.

The symbiotic interaction of plants with arbuscular mycorrhizal (AM) fungi is ancient and widespread. Plants provide AM fungi with carbon in exchange for nutrients and water, making this interaction a prime target for crop improvement. However, plant-fungal interactions are restricted to a small subset of root cells, precluding the application of most conventional functional genomic techniques to study the molecular bases of these interactions.

Biosynthesis of Strained Amino Acids by a PLP-Dependent Enzyme through Cryptic Halogenation.

Amino acids (AAs) are modular building blocks which nature uses to synthesize both macromolecules, such as proteins, and small molecule natural products, such as alkaloids and non-ribosomal peptides. While the 20 main proteinogenic AAs display relatively limited side chain diversity, a wide range of non-canonical amino acids (ncAAs) exist that are not used by the ribosome for protein synthesis, but contain a broad array of structural features and functional groups. In this communication, we report the discovery of the biosynthetic pathway for a new ncAA, pazamine, which contains a cyclopropane ring formed in two steps.

Engineered as a Biosynthetic Platform of Nucleotide Sugars.

Glycosylation of biomolecules can greatly alter their physicochemical properties, cellular recognition, subcellular localization, and immunogenicity. Glycosylation reactions rely on the stepwise addition of sugars using nucleotide diphosphate (NDP)-sugars. Making these substrates readily available will greatly accelerate the characterization of new glycosylation reactions, elucidation of their underlying regulation mechanisms, and production of glycosylated molecules.

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.

Biosynthesis of Strained Amino Acids Through a PLP-Dependent Enzyme via Cryptic Halogenation.

Amino acids (AAs) are modular and modifiable building blocks which nature uses to synthesize both macromolecules, such as proteins, and small molecule natural products, such as alkaloids and non-ribosomal peptides (NRPs). While the 20 main proteinogenic AAs display relatively limited side-chain diversity, a wide range of non-canonical amino acids (ncAAs) exist that are not used by the ribosome for protein synthesis but contain a broad array of structural features and functional groups not found in proteinogenic AAs. In this communication, we report the discovery of the biosynthetic pathway for a new ncAA, pazamine, which contains a cyclopropane ring formed in two steps.

Deciphering triterpenoid saponin biosynthesis by leveraging transcriptome response to methyl jasmonate elicitation in Saponaria vaccaria.

Methyl jasmonate (MeJA) is a known elicitor of plant specialized metabolism, including triterpenoid saponins. Saponaria vaccaria is an annual herb used in traditional Chinese medicine, containing large quantities of oleanane-type triterpenoid saponins with anticancer properties and structural similarities to the vaccine adjuvant QS-21. Leveraging the MeJA-elicited saponin biosynthesis, we identify multiple enzymes catalyzing the oxidation and glycosylation of triterpenoids in S.

Field performance of switchgrass plants engineered for reduced recalcitrance.

Switchgrass ( L.) is a promising perennial bioenergy crop that achieves high yields with relatively low nutrient and energy inputs. Modification of cell wall composition for reduced recalcitrance can lower the costs of deconstructing biomass to fermentable sugars and other intermediates.

A Suite of Constitutive Promoters for Tuning Gene Expression in Plants.

The need for convenient tools to express transgenes over a large dynamic range is pervasive throughout plant synthetic biology; however, current efforts are largely limited by the heavy reliance on a small set of strong promoters, precluding more nuanced and refined engineering endeavors . To address this technical gap, we characterize a suite of constitutive promoters that span a wide range of transcriptional levels and develop a GoldenGate-based plasmid toolkit named PCONS, optimized for versatile cloning and rapid testing of transgene expression at varying strengths. We demonstrate how easy access to a stepwise gradient of expression levels can be used for optimizing synthetic transcriptional systems and the production of small molecules .

Elongated galactan side chains mediate cellulose-pectin interactions in engineered Arabidopsis secondary cell walls.

The plant secondary cell wall is a thickened matrix of polysaccharides and lignin deposited at the cessation of growth in some cells. It forms the majority of carbon in lignocellulosic biomass, and it is an abundant and renewable source for forage, fiber, materials, fuels, and bioproducts. The complex structure and arrangement of the cell wall polymers mean that the carbon is difficult to access in an economical and sustainable way.

Structural and biochemical insight into a modular β-1,4-galactan synthase in plants.

Rhamnogalacturonan I (RGI) is a structurally complex pectic polysaccharide with a backbone of alternating rhamnose and galacturonic acid residues substituted with arabinan and galactan side chains. Galactan synthase 1 (GalS1) transfers galactose and arabinose to either extend or cap the β-1,4-galactan side chains of RGI, respectively. Here we report the structure of GalS1 from Populus trichocarpa, showing a modular protein consisting of an N-terminal domain that represents the founding member of a new family of carbohydrate-binding module, CBM95, and a C-terminal glycosyltransferase family 92 (GT92) catalytic domain that adopts a GT-A fold.

Medicinal terpenoid UDP-glycosyltransferases in plants: recent advances and research strategies.

Terpenoid glycosides have significant curative effects on many kinds of diseases. Most of these compounds are derived from medicinal plants. Glycosylation is a key step in the biosynthesis of medicinal terpenoids.

Evaluation of engineered low-lignin poplar for conversion into advanced bioproducts.

Background: Lignocellulosic resources are promising feedstocks for the manufacture of bio-based products and bioenergy. However, the inherent recalcitrance of biomass to conversion into simple sugars currently hinders the deployment of advanced bioproducts at large scale. Lignin is a primary contributor to biomass recalcitrance as it protects cell wall polysaccharides from degradation and can inhibit hydrolytic enzymes via non-productive adsorption.

Correction to "Expression of Dehydroshikimate Dehydratase in Sorghum Improves Biomass Yield, Accumulation of Protocatechuate, and Biorefinery Economics".

[This corrects the article DOI: 10.1021/acssuschemeng.2c01160.

Cell wall ester modifications and volatile emission signatures of plant response to abiotic stress.

Growth suppression and defence signalling are simultaneous strategies that plants invoke to respond to abiotic stress. Here, we show that the drought stress response of poplar trees (Populus trichocarpa) is initiated by a suppression in cell wall derived methanol (MeOH) emissions and activation of acetic acid (AA) fermentation defences. Temperature sensitive emissions dominated by MeOH (AA/MeOH <30%) were observed from physiologically active leaves, branches, detached stems, leaf cell wall isolations and whole ecosystems.

Engineering sorghum for higher 4-hydroxybenzoic acid content.

Engineering bioenergy crops to accumulate coproducts can increase the value of lignocellulosic biomass and enable a sustainable bioeconomy. In this study, we engineered sorghum with a bacterial gene encoding a chorismate pyruvate-lyase () to reroute the plastidial pool of chorismate from the shikimate pathway into the valuable compound 4-hydroxybenzoic acid (4-HBA). A gene encoding a feedback-resistant version of 3-deoxy-d-arabino-heptulonate-7-phosphate synthase () was also introduced in an attempt to increase the carbon flux through the shikimate pathway.