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.

Comparative venation costs of monocotyledon and dicotyledon species in the eastern Colorado steppe.

MAIN CONCLUSION: Leaf vein network cost (total vein surface area per leaf volume) for major veins and vascular bundles did not differ between monocot and dicot species in 21 species from the eastern Colorado steppe. Dicots possessed significantly larger minor vein networks than monocots. Across the tree of life, there is evidence that dendritic vascular transport networks are optimized, balancing maximum speed and integrity of resource delivery with minimal resource investment in transport and infrastructure. Monocot venation, however, is not dendritic, and remains parallel down to the smallest vein orders with no space-filling capillary networks. Given this departure from the "optimized" dendritic network, one would assume that monocots are operating at a significant energetic disadvantage. In this study, we investigate whether monocot venation networks bear significantly greater carbon/construction costs per leaf volume than co-occurring dicots in the same ecosystem, and if so, what physiological or ecological advantage the monocot life form possesses to compensate for this deficit. Given that venation networks could also be optimized for leaf mechanical support or provide herbivory defense, we measured the vascular system of both monocot and dicots at three scales to distinguish between leaf investment in mechanical support (macroscopic vein), total transport and capacitance (vascular bundle), or exclusively water transport (xylem) for both parallel and dendritic venation networks. We observed that vein network cost (total vein surface area per leaf volume) for major veins and vascular bundles was not significantly different between monocot species and dicot species. Dicots, however, possess significantly larger minor vein networks than monocots. The 19 species subjected to gas-exchange measurement in the field displayed a broad range of Amax and but demonstrated no significant relationships with any metric of vascular network size in major or minor vein classes. Given that monocots do not seem to display any leaf hydraulic disadvantage relative to dicots, it remains an important research question why parallel venation (truly parallel, down to the smallest vessels) has not arisen more than once in the history of plant evolution.

Convergent evolution of fern nectaries facilitated independent recruitment of ant-bodyguards from flowering plants.

Plant-herbivore interactions reciprocally influence species' evolutionary trajectories. These interactions have led to many physical and chemical defenses across the plant kingdom. Some plants have even evolved indirect defense strategies to outsource their protection to ant bodyguards by bribing them with a sugary reward (nectar). Identifying the evolutionary processes underpinning these indirect defenses provide insight into the evolution of plant-animal interactions. Using a cross-kingdom, phylogenetic approach, we examined the convergent evolution of ant-guarding nectaries across ferns and flowering plants. Here, we discover that nectaries originated in ferns and flowering plants concurrently during the Cretaceous, coinciding with the rise of plant associations in ants. While nectaries in flowering plants evolved steadily through time, ferns showed a pronounced lag of nearly 100 My between their origin and subsequent diversification in the Cenozoic. Importantly, we find that as ferns transitioned from the forest floor into the canopy, they secondarily recruited ant bodyguards from existing ant-angiosperm relationships.

Twenty years of big plant genera.

In 2004, David Frodin published a landmark review of the history and concepts of big plant genera. Two decades of taxonomic activity have taken place since, coinciding with a revolution in phylogenetics and taxonomic bioinformatics. Here we use data from the World Flora Online (WFO) to provide an updated list of big (more than 500 species) and megadiverse (more than 1000 species) flowering plant genera and highlight changes since 2004. The number of big genera has increased from 57 to 86; today one of every four plant species is classified as a member of a big genus, with 14% in just 28 megadiverse genera. Most (71%) of the growth in big genera since 2000 is the result of new species description, not generic re-circumscription. More than 15% of all currently accepted flowering plant species described in the last two decades are in big genera, suggesting that groups previously considered intractable are now being actively studied taxonomically. Despite this rapid growth in big genera, they remain a significant yet understudied proportion of plant diversity. They represent a significant proportion of global plant diversity and should remain a priority not only for taxonomy but for understanding global diversity patterns and plant evolution in general.

Shifts in evolutionary lability underlie independent gains and losses of root-nodule symbiosis in a single clade of plants.

Root nodule symbiosis (RNS) is a complex trait that enables plants to access atmospheric nitrogen converted into usable forms through a mutualistic relationship with soil bacteria. Pinpointing the evolutionary origins of RNS is critical for understanding its genetic basis, but building this evolutionary context is complicated by data limitations and the intermittent presence of RNS in a single clade of ca. 30,000 species of flowering plants, i.e., the nitrogen-fixing clade (NFC). We developed the most extensive de novo phylogeny for the NFC and an RNS trait database to reconstruct the evolution of RNS. Our analysis identifies evolutionary rate heterogeneity associated with a two-step process: An ancestral precursor state transitioned to a more labile state from which RNS was rapidly gained at multiple points in the NFC. We illustrate how a two-step process could explain multiple independent gains and losses of RNS, contrary to recent hypotheses suggesting one gain and numerous losses, and suggest a broader phylogenetic and genetic scope may be required for genome-phenome mapping.

Improving species distribution forecasts by measuring and communicating uncertainty: An invasive species case study.

Forecasting invasion risk under future climate conditions is critical for the effective management of invasive species, and species distribution models (SDMs) are key tools for doing so. However, SDM-based forecasts are uncertain, especially when correlative statistical models extrapolate to nonanalog environmental domains, such as future climate conditions. Different assumptions about the functional form of the temperature-suitability relationship can impact predicted habitat suitability under novel conditions. Hence, methods to understand the sources of uncertainty are critical when applying SDMs. Here, we use high-resolution predictions of lake water temperatures to project changes in habitat suitability under future climate conditions for an invasive macrophyte (Myriophyllym spicatum). Future suitability was predicted using five global circulation models and three statistical models that assumed different species-temperature functional responses. The suitability of lakes for M. spicatum was overall predicted to increase under future climate conditions, but the magnitude and direction of change in suitability varied greatly among lakes. Variability was most pronounced for lakes under nonanalog temperature conditions, indicating that predictions for these lakes remained highly uncertain. Integrating predictions from SDMs that differ in their species-environment response function, while explicitly quantifying uncertainty across analog and nonanalog domains, can provide a more robust and useful approach to forecasting invasive species distribution under climate change.

Ancient Duplication and Lineage-Specific Transposition Determine Evolutionary Trajectory of ERF Subfamily across Angiosperms.

AP2/ERF transcription factor family plays an important role in plant development and stress responses. Previous studies have shed light on the evolutionary trajectory of the AP2 and DREB subfamilies. However, knowledge about the evolutionary history of the ERF subfamily in angiosperms still remains limited. In this study, we performed a comprehensive analysis of the ERF subfamily from 107 representative angiosperm species by combining phylogenomic and synteny network approaches. We observed that the expansion of the ERF subfamily was driven not only by whole-genome duplication (WGD) but also by tandem duplication (TD) and transposition duplication events. We also found multiple transposition events in Poaceae, Brassicaceae, Poales, Brassicales, and Commelinids. These events may have had notable impacts on copy number variation and subsequent functional divergence of the ERF subfamily. Moreover, we observed a number of ancient tandem duplications occurred in the ERF subfamily across angiosperms, e.g., in Subgroup IX, IXb originated from ancient tandem duplication events within IXa. These findings together provide novel insights into the evolution of this important transcription factor family.

Cretaceous chloranthoids: early prominence, extinct diversity and missing links.

BACKGROUND: The Chloranthaceae comprise four extant genera (Hedyosmum, Ascarina, Chloranthus and Sarcandra), all with simple flowers. Molecular phylogenetics indicates that the Chloranthaceae diverged very early in angiosperm evolution, although how they are related to eudicots, magnoliids, monocots and Ceratophyllum is uncertain. Fossil pollen similar to that of Ascarina and Hedyosmum has long been recognized in the Early Cretaceous, but over the last four decades evidence of extinct Chloranthaceae based on other types of fossils has expanded dramatically and contributes significantly to understanding the evolution of the family. SCOPE: Studies of fossils from the Cretaceous, especially mesofossils of Early Cretaceous age from Portugal and eastern North America, recognized diverse flowers, fruits, seeds, staminate inflorescences and stamens of extinct chloranthoids. These early chloranthoids include forms related to extant Hedyosmum and also to the Ascarina, Chloranthus and Sarcandra clade. In the Late Cretaceous there are several occurrences of distinctive fossil androecia related to extant Chloranthus. The rich and still expanding Cretaceous record of Chloranthaceae contrasts with a very sparse Cenozoic record, emphasizing that the four extant genera are likely to be relictual, although speciation within the genera might have occurred in relatively recent times. In this study, we describe three new genera of Early Cretaceous chloranthoids and summarize current knowledge on the extinct diversity of the group. CONCLUSIONS: The evolutionary lineage that includes extant Chloranthaceae is diverse and abundantly represented in Early Cretaceous mesofossil floras that provide some of the earliest evidence of angiosperm reproductive structures. Extinct chloranthoids, some of which are clearly in the Chloranthaceae crown group, fill some of the morphological gaps that currently separate the extant genera, help to illuminate how some of the unusual features of extant Chloranthaceae evolved and suggest that Chloranthaceae are of disproportionate importance for a more refined understanding of ecology and phylogeny of early angiosperm diversification.

The genetic control of herkogamy.

Herkogamy is the spatial separation of anthers and stigmas within complete flowers, and is a key floral trait that promotes outcrossing in many angiosperms. The degree of separation between pollen-producing anthers and receptive stigmas has been shown to influence rates of self-pollination amongst plants, with a reduction in herkogamy increasing rates of successful selfing in self-compatible species. Self-pollination is becoming a critical issue in horticultural crops grown in environments where biotic pollinators are limited, absent, or difficult to utilise. In these cases, poor pollination results in reduced yield and misshapen fruit. Whilst there is a growing body of work elucidating the genetic basis of floral organ development, the genetic and environmental control points regulating herkogamy are poorly understood. A better understanding of the developmental and regulatory pathways involved in establishing varying degrees of herkogamy is needed to provide insights into the production of flowers more adept at selfing to produce consistent, high-quality fruit. This review presents our current understanding of herkogamy from a genetics and hormonal perspective.

Parallel evolution of angiosperm-like venation in Peltaspermales: a reinvestigation of Furcula.

Leaf venation is a pivotal trait in the success of vascular plants. Whereas gymnosperms have single or sparsely branched parallel veins, angiosperms developed a hierarchical structure of veins that form a complex reticulum. Its physiological consequences are considered to have enabled angiosperms to dominate terrestrial ecosystems in the Late Cretaceous and Cenozoic. Although a hierarchical-reticulate venation also occurs in some groups of extinct seed plants, it is unclear whether these are stem relatives of angiosperms or have evolved these traits in parallel. Here, we re-examine the morphology of the enigmatic foliage taxon Furcula, a potential early Mesozoic angiosperm relative, and argue that its hierarchical vein network represents convergent evolution (in the Late Triassic) with flowering plants (which developed in the Early Cretaceous) based on details of vein architecture and the absence of angiosperm-like stomata and guard cells. We suggest that its nearest relatives are Peltaspermales similar to Scytophyllum and Vittaephyllum, the latter being a genus that originated during the Late Triassic (Carnian) and shares a hierarchical vein system with Furcula. We further suggest that the evolution of hierarchical venation systems in the early Permian, the Late Triassic, and the Early Cretaceous represent 'natural experiments' that might help resolve the selective pressures enabling this trait to evolve.

Flowering plant reproduction.

Flowering plants, also known as angiosperms, emerged approximately 150 to 200 million years ago. Since then, they have undergone rapid and extensive expansion, now encompassing around 90% of all land plant species. The remarkable diversification of this group has been a subject of in-depth investigations, and several evolutionary innovations have been proposed to account for their success. In this primer, we will specifically focus on one such innovation: the advent of seeds containing endosperm.

Mfind: a tool for DNA barcode analysis in angiosperms and its relationship with microsatellites using a sliding window algorithm.

MAIN CONCLUSION: Mfind is a tool to analyze the impact of microsatellite presence on DNA barcode specificity. We found a significant correlation between barcode entropy and microsatellite count in angiosperm. Genetic barcodes and microsatellites are some of the identification methods in taxonomy and biodiversity research. It is important to establish a relationship between microsatellite quantification and genetic information in barcodes. In order to clarify the association between the genetic information in barcodes (expressed as Shannon's Measure of Information, SMI) and microsatellites count, a total of 330,809 DNA barcodes from the BOLD database (Barcode of Life Data System) were analyzed. A parallel sliding-window algorithm was developed to compute the Shannon entropy of the barcodes, and this was compared with the quantification of microsatellites like (AT)n, (AC)n, and (AG)n. The microsatellite search method utilized an algorithm developed in the Java programming language, which systematically examined the genetic barcodes from an angiosperm database. For this purpose, a computational tool named Mfind was developed, and its search methodology is detailed. This comprehensive study revealed a broad overview of microsatellites within barcodes, unveiling an inverse correlation between the sumz of microsatellites count and barcodes information. The utilization of the Mfind tool demonstrated that the presence of microsatellites impacts the barcode information when considering entropy as a metric. This effect might be attributed to the concise length of DNA barcodes and the repetitive nature of microsatellites, resulting in a direct influence on the entropy of the barcodes.

Phylogenomics and the rise of the angiosperms.

Angiosperms are the cornerstone of most terrestrial ecosystems and human livelihoods1,2. A robust understanding of angiosperm evolution is required to explain their rise to ecological dominance. So far, the angiosperm tree of life has been determined primarily by means of analyses of the plastid genome3,4. Many studies have drawn on this foundational work, such as classification and first insights into angiosperm diversification since their Mesozoic origins5-7. However, the limited and biased sampling of both taxa and genomes undermines confidence in the tree and its implications. Here, we build the tree of life for almost 8,000 (about 60%) angiosperm genera using a standardized set of 353 nuclear genes8. This 15-fold increase in genus-level sampling relative to comparable nuclear studies9 provides a critical test of earlier results and brings notable change to key groups, especially in rosids, while substantiating many previously predicted relationships. Scaling this tree to time using 200 fossils, we discovered that early angiosperm evolution was characterized by high gene tree conflict and explosive diversification, giving rise to more than 80% of extant angiosperm orders. Steady diversification ensued through the remaining Mesozoic Era until rates resurged in the Cenozoic Era, concurrent with decreasing global temperatures and tightly linked with gene tree conflict. Taken together, our extensive sampling combined with advanced phylogenomic methods shows the deep history and full complexity in the evolution of a megadiverse clade.

Unprecedented variation pattern of plastid genomes and the potential role in adaptive evolution in Poales.

BACKGROUND: The plastid is the photosynthetic organelle in plant cell, and the plastid genomes (plastomes) are generally conserved in evolution. As one of the most economically and ecologically important order of angiosperms, Poales was previously documented to exhibit great plastomic variation as an order of photoautotrophic plants. RESULTS: We acquired 93 plastomes, representing all the 16 families and 5 major clades of Poales to reveal the extent of their variation and evolutionary pattern. Extensive variation including the largest one in monocots with 225,293 bp in size, heterogeneous GC content, and a wide variety of gene duplication and loss were revealed. Moreover, rare occurrences of three inverted repeat (IR) copies in angiosperms and one IR loss were observed, accompanied by short IR (sIR) and small direct repeat (DR). Widespread structural heteroplasmy, diversified inversions, and unusual genomic rearrangements all appeared in Poales, occasionally within a single species. Extensive repeats in the plastomes were found to be positively correlated with the observed inversions and rearrangements. The variation all showed a "small-large-moderate" trend along the evolution of Poales, as well as for the sequence substitution rate. Finally, we found some positively selected genes, mainly in C4 lineages, while the closely related lineages of those experiencing gene loss tended to have undergone more relaxed purifying selection. CONCLUSIONS: The variation of plastomes in Poales may be related to its successful diversification into diverse habitats and multiple photosynthetic pathway transitions. Our order-scale analyses revealed unusual evolutionary scenarios for plastomes in the photoautotrophic order of Poales and provided new insights into the plastome evolution in angiosperms as a whole.

Intensive pasture management alters the composition and structure of plant-pollinator interactions in Sibiu, Romania.

Background: Land management change towards intensive grazing has been shown to alter plant and pollinator communities and the structure of plant-pollinator interactions in different ways across the world. Land-use intensification in Eastern Europe is shifting highly diverse, traditionally managed hay meadows towards intensive pastures, but few studies have examined how this influences plant-pollinator networks. We hypothesized that the effects of intensive grazing on networks will depend on how plant communities and their floral traits change. Methods: We investigated plant and pollinator diversity and composition and the structure of plant-pollinator interactions near Sibiu, Romania at sites that were traditionally managed as hay meadows or intensive pastures. We quantified the identity and abundance of flowering plants, and used transect walks to observe pollinator genera interacting with flowering plant species. We evaluated the effects of management on diversity, composition and several indices of network structure. Results: Pollinator but not plant diversity declined in pastures and both plant and pollinator taxonomic composition shifted. Functional diversity and composition remained unchanged, with rather specialized flowers having been found to dominate in both hay meadows and pastures. Apis mellifera was found to be the most abundant pollinator. Its foraging preferences played a crucial role in shaping plant-pollinator network structure. Apis mellifera thus preferred the highly abundant Dorycnium herbaceum in hay meadows, leading to hay meadows networks with lower Shannon diversity and interaction evenness. In pastures, however, it preferred less abundant and more generalized flower resources. With pollinators being overall less abundant and more generalized in pastures, we found that niche overlap between plants was higher. Discussion: With both hay meadows and pastures being dominated by plant species with similar floral traits, shifts in pollinator preferences seem to have driven the observed changes in plant-pollinator interaction networks. We thus conclude that the effects of grazing on pollinators and their interactions are likely to depend on the traits of plant species present in different management types as well as on the effects of grazing on plant community composition. We thereby highlight the need for better understanding how floral abundance shapes pollinator visitation rates and how floral traits may influence this relationship.

A comparative analysis of plastome evolution in autotrophic Piperales.

PREMISE: Many plastomes of autotrophic Piperales have been reported to date, describing a variety of differences. Most studies focused only on a few species or a single genus, and extensive, comparative analyses have not been done. Here, we reviewed publicly available plastome reconstructions for autotrophic Piperales, reanalyzed publicly available raw data, and provided new sequence data for all previously missing genera. Comparative plastome genomics of >100 autotrophic Piperales were performed. METHODS: We performed de novo assemblies to reconstruct the plastomes of newly generated sequence data. We used Sanger sequencing and read mapping to verify the assemblies and to bridge assembly gaps. Furthermore, we reconstructed the phylogenetic relationships as a foundation for comparative plastome genomics. RESULTS: We identified a plethora of assembly and annotation issues in published plastome data, which, if unattended, will lead to an artificial increase of diversity. We were able to detect patterns of missing and incorrect feature annotation and determined that the inverted repeat (IR) boundaries were the major source for erroneous assembly. Accounting for the aforementioned issues, we discovered relatively stable junctions of the IRs and the small single-copy region (SSC), whereas the majority of plastome variations among Piperales stems from fluctuations of the boundaries of the IR and the large single-copy (LSC) region. CONCLUSIONS: This study of all available plastomes of autotrophic Piperales, expanded by new data for previously missing genera, highlights the IR-LSC junctions as a potential marker for discrimination of various taxonomic levels. Our data indicates a pseudogene-like status for cemA and ycf15 in various Piperales. Based on a review of published data, we conclude that incorrect IR-SSC boundary identification is the major source for erroneous plastome assembly. We propose a gold standard for assembly and annotation of high-quality plastomes based on de novo assembly methods and appropriate references for gene annotation.

Isolating Antipathogenic Fungal Coumarins from Coriaria nepalensis and Determining Their Primary Mechanism In Vitro.

Through bioassay-guided isolation, eight undescribed coumarins (1-8), along with six reported coumarins (9-14), were obtained from Coriaria nepalensis. The new structures were determined by using IR, UV, NMR, HRESIMS, and ECD calculations. The results of the biological activity assays showed that compound 9 exhibited broad spectrum antifungal activities against all tested fungi in vitro and a significant inhibitory effect on Phytophthora nicotianae with an EC50 value of 3.00 µg/mL. Notably, compound 9 demonstrated greater curative and protective effects against tobacco balack shank than those of osthol in vivo. Thus, 9 was structurally modified to obtain new promising antifungal agents, and the novel derivatives (17b, 17j, and 17k) exhibited better effects on Sclerotinia sclerotiorum than did lead compound 9. Preliminary mechanistic exploration illustrated that 9 could enhance cell membrane permeability, destroy the morphology and ultrastructure of cells, and reduce the exopolysaccharide content of P. nicotianae mycelia. Furthermore, the cytotoxicity results revealed that compound 9 exhibited relatively low cytotoxicity against HEK293 cell lines with an inhibition rate of 33.54% at 30 µg/mL. This research is promising for the discovery of new fungicides from natural coumarins with satisfactory ecological compatibility.

From Flora to Pharmaceuticals: 100 new additions to angiosperms of Gafargaon subdistrict in Bangladesh and unraveling antidiabetic drug candidates targeting DPP4 through in silico approach.

Addition to the angiosperm flora provides essential insights into the biodiversity of a region, contributing to ecological understanding and conservation planning. Gafargaon subdistrict under Mymensingh district in Bangladesh represents a diverse population of angiosperms with a multifaceted ecosystem that demands re-evaluation of the existing angiosperm diversity of Gafargaon to update the status of angiosperm taxa and facilitate their conservation efforts. With this endeavor, a total of 100 angiosperm taxa belonging to 90 genera and 46 families were uncovered as additional occurrence in Gafargaon. The species in the area showcased a variety of life forms, including 63 herbs, 14 shrubs, 14 trees, and 9 climbers. Among the recorded taxa, Chamaecostus cuspidatus (Nees & Mart.) C.D. Specht & D.W. Stev. was selected for antidiabetic drug design endeavor based on citation frequency and ethnomedicinal evidence. A total of 41 phytochemicals of C. cuspidatus were screened virtually, targeting the Dipeptidyl peptidase 4 protein through structure-based drug design approach, which unveiled two lead compounds, such as Tigogenin (-9.0 kcal/mol) and Diosgenin (-8.5 kcal/mol). The lead candidates demonstrated favorable pharmacokinetic and pharmacodynamic properties with no major side effects. Molecular dynamics simulation revealed notable stability and structural compactness of the lead compounds. Principal component analysis and Gibbs free energy landscape further supported the results of molecular dynamics simulation. Molecular mechanics-based MM/GBSA approach unraveled higher free binding energies of Diosgenin (-47.36 kcal/mol) and Tigogenin (-46.70 kcal/mol) over Alogliptin (-46.32 kcal/mol). The outcome of the present investigation would enrich angiosperm flora of Gafargaon and shed light on the role of C. cuspidatus to develop novel antidiabetic therapeutics to combat diabetes.

Genome-Wide Identification of the 14-3-3 Gene Family and Its Involvement in Salt Stress Response through Interaction with NsVP1 in Nitraria sibirica Pall.

14-3-3 proteins are widely distributed in eukaryotic cells and play an important role in plant growth, development, and stress tolerance. This study revealed nine 14-3-3 genes from the genome of Nitraria sibirica Pall., a halophyte with strong salt tolerance. The physicochemical properties, multiple sequence alignment, gene structure and motif analysis, and chromosomal distributions were analyzed, and phylogenetic analysis, cis-regulatory elements analysis, and gene transcription and expression analysis of Ns14-3-3s were conducted. The results revealed that the Ns14-3-3 gene family consists of nine members, which are divided into two groups: ε (four members) and non-ε (five members). These members are acidic hydrophilic proteins. The genes are distributed randomly on chromosomes, and the number of introns varies widely among the two groups. However, all genes have similar conserved domains and three-dimensional protein structures. The main differences are found at the N-terminus and C-terminus. The promoter region of Ns14-3-3s contains multiple cis-acting elements related to light, plant hormones, and abiotic stress responses. Transcriptional profiling and gene expression pattern analysis revealed that Ns14-3-3s were expressed in all tissues, although with varying patterns. Under salt stress conditions, Ns14-3-3 1a, Ns14-3-3 1b, Ns14-3-3 5a, and Ns14-3-3 7a showed significant changes in gene expression. Ns14-3-3 1a expression decreased in all tissues, Ns14-3-3 7a expression decreased by 60% to 71% in roots, and Ns14-3-3 1b expression increased by 209% to 251% in stems. The most significant change was observed in Ns14-3-3 5a, with its expression in stems increasing by 213% to 681%. The yeast two-hybrid experiments demonstrated that Ns14-3-3 5a interacts with NsVP1 (vacuolar H+-pyrophosphatase). This result indicates that Ns14-3-3 5a may respond to salt stress by promoting ionic vacuole compartmentalization in stems and leaves through interactions with NsVP1. In addition, N. sibirica has a high number of stems, allowing it to compartmentalize more ions through its stem and leaf. This may be a contributing factor to its superior salt tolerance compared to other plants.

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.