Conserved amino acid residues and gene expression patterns associated with the substrate preferences of the competing enzymes FLS and DFR.

BACKGROUND: Flavonoids, an important class of specialized metabolites, are synthesized from phenylalanine and present in almost all plant species. Different branches of flavonoid biosynthesis lead to products like flavones, flavonols, anthocyanins, and proanthocyanidins. Dihydroflavonols form the branching point towards the production of non-colored flavonols via flavonol synthase (FLS) and colored anthocyanins via dihydroflavonol 4-reductase (DFR). Despite the wealth of publicly accessible data, there remains a gap in understanding the mechanisms that mitigate competition between FLS and DFR for the shared substrate, dihydroflavonols. RESULTS: An angiosperm-wide comparison of FLS and DFR sequences revealed the amino acids at positions associated with the substrate specificity in both enzymes. A global analysis of the phylogenetic distribution of these amino acid residues revealed that monocots generally possess FLS with Y132 (FLSY) and DFR with N133 (DFRN). In contrast, dicots generally possess FLSH and DFRN, DFRD, and DFRA. DFRA, which restricts substrate preference to dihydrokaempferol, previously believed to be unique to strawberry species, is found to be more widespread in angiosperms and has evolved independently multiple times. Generally, angiosperm FLS appears to prefer dihydrokaempferol, whereas DFR appears to favor dihydroquercetin or dihydromyricetin. Moreover, in the FLS-DFR competition, the dominance of one over the other is observed, with typically only one gene being expressed at any given time. CONCLUSION: This study illustrates how almost mutually exclusive gene expression and substrate-preference determining residues could mitigate competition between FLS and DFR, delineates the evolution of these enzymes, and provides insights into mechanisms directing the metabolic flux of the flavonoid biosynthesis, with potential implications for ornamental plants and molecular breeding strategies.

Mechanisms of ammonium detoxification in submerged macrophytes under shade conditions.

Excessive ammonium disrupts the biological and physical characteristics of aquatic freshwater ecosystems, causing nutrient imbalances and toxicity. Different macrophytes exhibit varying tolerance levels to ammonium stress, influenced by species-specific adaptations. However, eutrophic water bodies not only have high nutrient loads but also exhibit low light transparency, necessitating an understanding of how submerged macrophytes cope with both high ammonium concentrations and low light conditions. In this study, we explored the tolerance of submerged macrophytes under these challenging conditions by testing various ammonium concentrations and light intensities. Our findings reveal that Myriophyllum spicatum demonstrates high ammonium tolerance under both optimal and low light intensities. Specifically, under optimal light, the primary ammonium assimilation pathway is catalyzed by NADH-GDH (Nicotinamide Adenine Dinucleotide-dependent Glutamate Dehydrogenase), with its activity increasing 4-fold at 50 mg L-1 [NH4+-N] compared to the control. Conversely, under low light intensity, the GS (Glutamine Synthetase)-catalyzed pathway becomes predominant, with GS activity rising 3-fold at 50 mg L-1 [NH4+-N] compared to the control. These results provide new insights into the adaptive mechanisms of M. spicatum, highlighting its flexible strategies for ammonium assimilation and its potential application in water restoration efforts. This study offers valuable information on the enzymatic pathways involved in ammonium detoxification, which is essential for developing effective strategies to manage and restore eutrophic aquatic systems.

Uptake and transport mechanisms for cadmium by Myriophyllum aquaticum in a constructed wetland.

Myriophyllum aquaticum (M. aquaticum), as a Cd-highly enriched and tolerant species, has greater application in phytoremediation of Cd-polluted waters. Mechanisms of Cd uptake and transport of M. aquaticum were comprehensively investigated in this work. Transport direction of Cd was observed both from the roots to the aboveground and vice versa. The aboveground can be harvested during vigorous growth and flowering periods, further improving the efficient phytoremediation of Cd-polluted wastewater. Moreover, analysis of transpiration inhibition, low-temperature treatment and metabolic inhibition indicated that the uptake and transport of Cd by M. aquaticum can be achieved via the coexistence of the free diffusion-dominated apoplast pathway dominated by transpiration and the "cellular pathway" dominated by active absorption, with the active energy-demanding cellular pathway playing a dominant role. The obtained results have important implications in the in-depth exploration of uptake, transport and distribution mechanisms of heavy metals during phytoremediation of aquatic plants.

Herbarium specimens as tools for exploring the evolution of fatty acid-derived natural products in plants.

Plants synthesize natural products via lineage-specific offshoots of their core metabolic pathways, including fatty acid synthesis. Recent studies have shed light on new fatty acid-derived natural products and their biosynthetic pathways in disparate plant species. Inspired by this progress, we set out to develop tools for exploring the evolution of fatty-acid derived products. We sampled multiple species from all major clades of euphyllophytes, including ferns, gymnosperms, and angiosperms (monocots and eudicots), and we show that the compositional profiles (though not necessarily the total amounts) of fatty-acid derived surface waxes from preserved plant specimens are consistent with those obtained from freshly collected tissue in a semi-quantitative and sometimes quantitative manner. We then sampled herbarium specimens representing 57 monocot species to assess the phylogenetic distribution and evolution, of two fatty acid-derived natural products found in that clade: beta-diketones and alkylresorcinols. These chemical data, combined with analyses of 26 monocot genomes, revealed a co-occurrence (though not necessarily a causal relationship) between whole genome duplication and the evolution of diketone synthases from an ancestral alkylresorcinol synthase-like polyketide synthase. Limitations of using herbarium specimen wax profiles as proxies for those of fresh tissue seem likely to include effects from loss of epicuticular wax crystals, effects from preservation techniques, and variation in wax chemical profiles due to genotype or environment. Nevertheless, this work reinforces the widespread utility of herbarium specimens for studying leaf surface waxes (and possibly other chemical classes) and reveals some of the evolutionary history of fatty acid-derived natural products within monocots.

Evolution and post-transcriptional regulation insights of m6A writers, erasers, and readers in plant epitranscriptome.

As a dynamic and reversible post-transcriptional marker, N6-methyladenosine (m6A) plays an important role in the regulation of biological functions, which are mediated by m6A pathway components including writers (MT-A70, FIP37, VIR and HAKAI family), erasers (ALKBH family) and readers (YTH family). There is an urgent need for a comprehensive analysis of m6A pathway components across species at evolutionary levels. In this study, we identified 4062 m6A pathway components from 154 plant species including green algae, utilizing large-scale phylogenetic to explore their origin and evolution. We discovered that the copy number of writers was conserved among different plant lineages, with notable expansions in the ALKBH and YTH families. Synteny network analysis revealed conserved genomic contexts and lineage-specific transpositions. Furthermore, we used Direct RNA Sequencing (DRS) to reveal the Poly(A) length (PAL) and m6A ratio profiles in six angiosperms species, with a particular focus on the m6A pathway components. The ECT1/2-Poeaece4 sub-branches (YTH family) with unique genomic contexts exhibited significantly higher expression level than genes of other ECT1/2 poeaece sub-branches (ECT1/2-Poeaece1-3), accompanied by lower m6A modification and PAL. Besides, conserved m6A sites distributed in CDS and 3'UTR were detected in the ECT1/2-Poaceae4, and the dual-luciferase assay further demonstrated that these conserved m6A sites in the 3'UTR negatively regulated the expression of Firefly luciferase (LUC) gene. Finally, we developed transcription factor regulatory networks for m6A pathway components, using yeast one-hybrid assay demonstrated that PheBPC1 could interact with the PheECT1/2-5 promoter. Overall, this study presents a comprehensive evolutionary and functional analysis of m6A pathway components and their modifications in plants, providing a valuable resource for future functional analysis in this field.

A trait-based root acquisition-defence-decomposition framework in angiosperm tree species.

To adapt to the complex belowground environment, plants make trade-offs between root resource acquisition and defence ability. This includes forming partnerships with different types of root associating microorganisms, such as arbuscular mycorrhizal and ectomycorrhizal fungi. These trade-offs, by mediating root chemistry, exert legacy effects on nutrient release during decomposition, which may, in turn, affect the ability of new roots to re-acquire resources, thereby generating a feedback loop. However, the linkages at the basis of this potential feedback loop remain largely unquantified. Here, we propose a trait-based root 'acquisition-defence-decomposition' conceptual framework and test the strength of relevant linkages across 90 angiosperm tree species. We show that, at the plant species level, the root-fungal symbiosis gradient within the root economics space, root chemical defence (condensed tannins), and root decomposition rate are closely linked, providing support to this framework. Beyond the dichotomy between arbuscular mycorrhizal-dominated versus ectomycorrhizal-dominated systems, we suggest a continuous shift in feedback loops, from 'high arbuscular mycorrhizal symbiosis-low defence-fast decomposition-inorganic nutrition' by evolutionarily ancient taxa to 'high ectomycorrhizal symbiosis-high defence-slow decomposition-organic nutrition' by more modern taxa. This 'acquisition-defence-decomposition' framework provides a foundation for testable hypotheses on multidimensional linkages between species' belowground strategies and ecosystem nutrient cycling in an evolutionary context.

Elevated atmospheric CO2 has small, species-specific effects on pollen chemistry and plant growth across flowering plant species.

Elevated atmospheric carbon dioxide (eCO2) can affect plant growth and physiology, which can, in turn, impact herbivorous insects, including by altering pollen or plant tissue nutrition. Previous research suggests that eCO2 can reduce pollen nutrition in some species, but it is unknown whether this effect is consistent across flowering plant species. We experimentally quantified the effects of eCO2 across multiple flowering plant species on plant growth in 9 species and pollen chemistry (%N an estimate for protein content and nutrition in 12 species; secondary chemistry in 5 species) in greenhouses. For pollen nutrition, only buckwheat significantly responded to eCO2, with %N increasing in eCO2; CO2 treatment did not affect pollen amino acid composition but altered secondary metabolites in buckwheat and sunflower. Plant growth under eCO2 exhibited two trends across species: plant height was taller in 44% of species and flower number was affected for 63% of species (3 species with fewer and 2 species with more flowers). The remaining growth metrics (leaf number, above-ground biomass, flower size, and flowering initiation) showed divergent, species-specific responses, if any. Our results indicate that future eCO2 is unlikely to uniformly change pollen chemistry or plant growth across flowering species but may have the potential to alter ecological interactions, or have particularly important effects on specialized pollinators.

Eco-evolutionary processes shaping floral nectar sugar composition.

Floral nectar sugar composition is assumed to reflect the nutritional demands and foraging behaviour of pollinators, but the relative contributions of evolutionary and abiotic factors to nectar sugar composition remain largely unknown across the angiosperms. We compiled a comprehensive dataset on nectar sugar composition for 414 insect-pollinated plant species across central Europe, along with phylogeny, paleoclimate, flower morphology, and pollinator dietary demands, to disentangle their relative effects. We found that phylogeny was strongly related with nectar sucrose content, which increased with the phylogenetic age of plant families, but even more strongly with historic global surface temperature. Nectar sugar composition was also defined by floral morphology, though it was not related to our functional measure of pollinator dietary demands. However, specialist pollinators of current plant-pollinator networks predominantly visited plant species with sucrose-rich nectar. Our results suggest that both physiological mechanisms related to plant water balance and evolutionary effects related to paleoclimatic changes have shaped floral nectar sugar composition during the radiation and specialisation of plants and pollinators. As a consequence, the high velocity of current climate change may affect plant-pollinator interaction networks due to a conflicting combination of immediate physiological responses and phylogenetic conservatism.

Exploring the interplay between angiosperm chlorophyll metabolism and environmental factors.

MAIN CONCLUSION: In this review, we summarize how chlorophyll metabolism in angiosperm is affected by the environmental factors: light, temperature, metal ions, water, oxygen, and altitude. The significance of chlorophyll (Chl) in plant leaf morphogenesis and photosynthesis cannot be overstated. Over time, researchers have made significant advancements in comprehending the biosynthetic pathway of Chl in angiosperms, along with the pivotal enzymes and genes involved in this process, particularly those related to heme synthesis and light-responsive mechanisms. Various environmental factors influence the stability of Chl content in angiosperms by modulating Chl metabolic pathways. Understanding the interplay between plants Chl metabolism and environmental factors has been a prominent research topic. This review mainly focuses on angiosperms, provides an overview of the regulatory mechanisms governing Chl metabolism, and the impact of environmental factors such as light, temperature, metal ions (iron and magnesium), water, oxygen, and altitude on Chl metabolism. Understanding these effects is crucial for comprehending and preserving the homeostasis of Chl metabolism.

Insights into dynamic coenocytic endosperm development: Unraveling molecular, cellular, and growth complexity.

The endosperm, a product of double fertilization, is one of the keys to the evolution and success of angiosperms in conquering the land. While there are differences in endosperm development among flowering plants, the most common form is coenocytic growth, where the endosperm initially undergoes nuclear division without cytokinesis and eventually becomes cellularized. This complex process requires interplay among networks of transcription factors such as MADS-box, auxin response factors (ARFs), and phytohormones. The role of cytoskeletal elements in shaping the coenocytic endosperm and influencing seed growth also becomes evident. This review offers a recent understanding of the molecular and cellular dynamics in coenocytic endosperm development and their contributions to the final seed size.

Comparative transcriptomics of seed nourishing tissues: uncovering conserved and divergent pathways in seed plants.

The evolutionary and ecological success of spermatophytes is intrinsically linked to the seed habit, which provides a protective environment for the initial development of the new generation. This environment includes an ephemeral nourishing tissue that supports embryo growth. In gymnosperms this tissue originates from the asexual proliferation of the maternal megagametophyte, while in angiosperms it is a product of fertilization, and is called the endosperm. The emergence of these nourishing tissues is of profound evolutionary value, and they are also food staples for most of the world's population. Here, using Orthofinder to infer orthologue genes among newly generated and previously published datasets, we provide a comparative transcriptomic analysis of seed nourishing tissues from species of several angiosperm clades, including those of early diverging lineages, as well as of one gymnosperm. Our results show that, although the structure and composition of seed nourishing tissues has seen significant divergence along evolution, there are signatures that are conserved throughout the phylogeny. Conversely, we identified processes that are specific to species within the clades studied, and thus illustrate their functional divergence. With this, we aimed to provide a foundation for future studies on the evolutionary history of seed nourishing structures, as well as a resource for gene discovery in future functional studies.

A molecular atlas of plastid and mitochondrial proteins reveals organellar remodeling during plant evolutionary transitions from algae to angiosperms.

Algae and plants carry 2 organelles of endosymbiotic origin that have been co-evolving in their host cells for more than a billion years. The biology of plastids and mitochondria can differ significantly across major lineages and organelle changes likely accompanied the adaptation to new ecological niches such as the terrestrial habitat. Based on organelle proteome data and the genomes of 168 phototrophic (Archaeplastida) versus a broad range of 518 non-phototrophic eukaryotes, we screened for changes in plastid and mitochondrial biology across 1 billion years of evolution. Taking into account 331,571 protein families (or orthogroups), we identify 31,625 protein families that are unique to primary plastid-bearing eukaryotes. The 1,906 and 825 protein families are predicted to operate in plastids and mitochondria, respectively. Tracing the evolutionary history of these protein families through evolutionary time uncovers the significant remodeling the organelles experienced from algae to land plants. The analyses of gained orthogroups identifies molecular changes of organelle biology that connect to the diversification of major lineages and facilitated major transitions from chlorophytes en route to the global greening and origin of angiosperms.

Cytochrome b5 diversity in green lineages preceded the evolution of syringyl lignin biosynthesis.

Lignin production marked a milestone in vascular plant evolution, and the emergence of syringyl (S) lignin is lineage specific. S-lignin biosynthesis in angiosperms, mediated by ferulate 5-hydroxylase (F5H, CYP84A1), has been considered a recent evolutionary event. F5H uniquely requires the cytochrome b5 protein CB5D as an obligatory redox partner for catalysis. However, it remains unclear how CB5D functionality originated and whether it coevolved with F5H. We reveal here the ancient evolution of CB5D-type function supporting F5H-catalyzed S-lignin biosynthesis. CB5D emerged in charophyte algae, the closest relatives of land plants, and is conserved and proliferated in embryophytes, especially in angiosperms, suggesting functional diversification of the CB5 family before terrestrialization. A sequence motif containing acidic amino residues in Helix 5 of the CB5 heme-binding domain contributes to the retention of CB5D function in land plants but not in algae. Notably, CB5s in the S-lignin-producing lycophyte Selaginella lack these residues, resulting in no CB5D-type function. An independently evolved S-lignin biosynthetic F5H (CYP788A1) in Selaginella relies on NADPH-dependent cytochrome P450 reductase as sole redox partner, distinct from angiosperms. These results suggest that angiosperm F5Hs coopted the ancient CB5D, forming a modern cytochrome P450 monooxygenase system for aromatic ring meta-hydroxylation, enabling the reemergence of S-lignin biosynthesis in angiosperms.

Chemical diversity in angiosperms - monoterpene synthases control complex reactions that provide the precursors for ecologically and commercially important monoterpenoids.

Monoterpene synthases (MTSs) catalyze the first committed step in the biosynthesis of monoterpenoids, a class of specialized metabolites with particularly high chemical diversity in angiosperms. In addition to accomplishing a rate enhancement, these enzymes manage the formation and turnover of highly reactive carbocation intermediates formed from a prenyl diphosphate substrate. At each step along the reaction path, a cationic intermediate can be subject to cyclization, migration of a proton, hydride, or alkyl group, or quenching to terminate the sequence. However, enzymatic control of ligand folding, stabilization of specific intermediates, and defined quenching chemistry can maintain the specificity for forming a signature product. This review article will discuss our current understanding of how angiosperm MTSs control the reaction environment. Such knowledge allows inferences about the origin and regulation of chemical diversity, which is pertinent for appreciating the role of monoterpenoids in plant ecology but also for aiding commercial efforts that harness the accumulation of these specialized metabolites for the food, cosmetic, and pharmaceutical industries.

Evolution of the WRKY Family in Angiosperms and Functional Diversity under Environmental Stress.

The transcription factor is an essential factor for regulating the responses of plants to external stimuli. The WRKY protein is a superfamily of plant transcription factors involved in response to various stresses (e.g., cold, heat, salt, drought, ions, pathogens, and insects). During angiosperm evolution, the number and function of WRKY transcription factors constantly change. After suffering from long-term environmental battering, plants of different evolutionary statuses ultimately retained different numbers of WRKY family members. The WRKY family of proteins is generally divided into three large categories of angiosperms, owing to their conserved domain and three-dimensional structures. The WRKY transcription factors mediate plant adaptation to various environments via participating in various biological pathways, such as ROS (reactive oxygen species) and hormone signaling pathways, further regulating plant enzyme systems, stomatal closure, and leaf shrinkage physiological responses. This article analyzed the evolution of the WRKY family in angiosperms and its functions in responding to various external environments, especially the function and evolution in Magnoliaceae plants. It helps to gain a deeper understanding of the evolution and functional diversity of the WRKY family and provides theoretical and experimental references for studying the molecular mechanisms of environmental stress.

Isolation of the Inner and Outer Membranes of the Chloroplast Envelope from Angiosperms.

The outer and the inner membranes of the chloroplast envelope, also called OEM and IEM, have distinct lipid and protein compositions. They host molecular systems involved in the biogenesis of the organelle, its cellular function, and its communication with other compartments. Here we describe a method for the isolation of these two membranes starting from intact chloroplast preparations, with two alternative procedures based on the starting material. One was developed from spinach leaves, the other from pea leaves. The two procedures differ in the method used to isolate and rupture chloroplasts and separate each membrane.

Epigenetic variation in early and late flowering plants of the rubber-producing Russian dandelion Taraxacum koksaghyz provides insights into the regulation of flowering time.

The Russian dandelion (Taraxacum koksaghyz) grows in temperate zones and produces large amounts of poly(cis-1,4-isoprene) in its roots, making it an attractive alternative source of natural rubber. Most T. koksaghyz plants require vernalization to trigger flower development, whereas early flowering varieties that have lost their vernalization dependence are more suitable for breeding and domestication. To provide insight into the regulation of flowering time in T. koksaghyz, we induced epigenetic variation by in vitro cultivation and applied epigenomic and transcriptomic analysis to the resulting early flowering plants and late flowering controls, allowing us to identify differences in methylation patterns and gene expression that correlated with flowering. This led to the identification of candidate genes homologous to vernalization and photoperiodism response genes in other plants, as well as epigenetic modifications that may contribute to the control of flower development. Some of the candidate genes were homologous to known floral regulators, including those that directly or indirectly regulate the major flowering control gene FT. Our atlas of genes can be used as a starting point to investigate mechanisms that control flowering time in T. koksaghyz in greater detail and to develop new breeding varieties that are more suited to domestication.

Therapeutic Potential of Pectin and Its Derivatives in Chronic Diseases.

Non-communicable diseases (NCDs) are described as a collection of chronic diseases that do not typically develop from an acute infection, have long-term health effects, and frequently require ongoing care and therapy. These diseases include heart disease, stroke, cancer, chronic lung disease, neurological diseases, osteoporosis, mental health disorders, etc. Known synthetic drugs for the treatment or prevention of NCDs become increasingly dangerous over time and pose high risks due to side effects such as hallucination, heart attack, liver failure, etc. As a result, scientists have had to look for other alternatives that are natural products and that are known to be less detrimental and contain useful bioactive compounds. The increasing understanding of the biological and pharmacological significance of carbohydrates has helped to raise awareness of their importance in living systems and medicine, given they play numerous biological roles. For example, pectin has been identified as a class of secondary metabolites found in medicinal plants that may play a significant role in the treatment and management of a variety of NCDs. Pectin is mainly made of homogalacturonan, which is a linear polymer composed primarily of D-galacturonic acid units (at least 65%) linked in a chain by α-(1,4)-glycosidic linkages. There are also modified pectins or derivatives that improve pectin's bioavailability. Pectin is found in the cell walls of higher plants (pteridophytes, angiosperms, and gymnosperms), particularly in the middle lamella of the plant material. Citrus pectin is used in various industries. This article compiles information that has been available for years about the therapeutic importance of pectin in chronic diseases, different modes of pectin extraction, the chemistry of pectin, and the potency of pectin and its derivatives.

The annonalide diterpene extracted from Casimirella ampla (Miers) reduces inflammatory and antinociceptive events in general models of inflammation.

ETHNOPHARMACOLOGICAL RELEVANCE: The plants of the genus Casimirella ampla (Miers) (C. ampla) are extensively used in folk medicine. For a long time, rural communities have been using extracts from its roots for food and therapeutic purposes. The extract is rich in diterpenoid annonalide (Annona), which has antiophidic, anti-inflammatory and antinociceptive properties. Inflammation is the body's primary defense mechanism against cell damage and invasion by pathogens, which can trigger acute and chronic inflammatory processes. The first line of treatment for this condition consists of the use of non-steroidal anti-inflammatory drugs, but these have numerous associated collateral damages, based on scientific knowledge about diterpenoids from C. ampla, as well as their already reported antinociceptive and anti-inflammatory properties. AIMS OF THE STUDY: Evaluate the effect of Annona in classic models of inflammation and pain. MATERIALS AND METHODS: Animals were pretreated with Annona (0.1, 1.0 and 10 mg/kg), or Tween 80 (2%), or indomethacin (Indo) (10 mg/kg) orally in the paw edema tests induced by carrageenan (Cg), serotonin (5-HT), histamine, bradykinin, 48/80 and, prostaglandin E2 (PGE2), evaluating microscopic lesion scores, migration of leukocytes to the peritoneal cavity, concentration of myeloperoxide (MPO), malonyldialdehyde (MDA) and glutathione (GSH), abdominal contortion test by acetic acid and formalin test. RESULTS: Treatment with Annona compound at a dose of 0.1 mg/kg was more effective in reducing inflammatory, oxidant and nociceptive parameters, as it reduced paw edema induced by carrageenan, through different mediators and migration of inflammatory cells. Furthermore, it worked by reducing the concentration of MPO, MDA, preserving GSH levels and reducing nociception caused by formalin and acetic acid.