Plant vasculature is essential for the transport of water, nutrients, and signaling molecules across organs, while also providing critical mechanical support for growth and development. Disruptions in vascular bundle formation can therefore lead to severe physiological and developmental defects. In maize, ethyl methanesulfonate (EMS)-induced dominant nonallelic Wilty mutants exhibit a pronounced wilting phenotype even under well-watered conditions, indicating underlying defects in vascular function. In this study, we characterized the Wi3 mutant, identified as ZmDrought-Overly-Sensitive1/DOS1, and compared it with the previously described Wi2 mutant to uncover shared mechanisms underlying their phenotypes. We provide evidence, by bulk segregant resequencing linkage disequilibrium of SNPs adjacent to the causal Wilty SNPs in respective ß- and α-tubulin genes, for the personal communication from Gerry Neuffer that Wi2/ß-tub6 provenance is from ACR-related stock, whereas Wi3/α-tub4 allele is from Mo17, not B73 as claimed by the authors who cloned Dos1. Histochemical staining and Fourier-transform infrared (FTIR) spectroscopy of vascular bundles in Wi3 indicated apparent alterations in cellulose and lignin content consistent with those observed in Wi2. Transcriptome analysis of shoot meristems further indicated that similar sets of genes and pathways are differentially expressed in both mutants, suggesting convergence on common biological pathways. Using bulk-segregant whole-genome resequencing, we identified alpha-tubulin4 (TUA4) as the causal gene in Wi3 (ZmDOS1), harboring a C-to-T substitution within the N-terminal GTPase-binding domain. This mutation results in a glutamic acid196-to-lysine substitution. Given that α- and β-tubulin subunits heterodimerize, and in many plants and animal mutant alleles are dominant-negative gains-of-function, we infer Wi2, Wi3, and likely Wi4, based on very similar FTIR biophysical difference spectra, may act as effectors of vascular bundle cell wall deposition, potentially involving vesicle trafficking as recently shown for asymmetric cell divisions in maize stomatal development. Together, these findings highlight the functional interdependence of tubulin subunits and provide a plausible mechanistic framework for the striking biophysical, transcriptomic, and phenotypic similarities observed between Wi2, Wi3/ZmDOS1, and Wi4 mutants.
ABSTRACT Cluster bean ( Cyamopsis tetragonoloba L.) is usually grown for its high galactomannan content as an industrial crop. Young edible cluster bean pods are also eaten as a nutritious vegetable in many areas. There is little research on the effects of drought on pod yield or the nutritional content of edible cluster bean pods, even with the growing importance of producing vegetable cluster bean in dry and semiarid regions. In this study, the effect of silicon nanoparticles (Si NPs) and methyl jasmonate (MeJA) applied concurrently on the growth performance, pod productivity, and nutritional quality of vegetable cluster bean was evaluated at three irrigation rates. A three‐factor experiment was done in a completely randomized block design with three replications under field conditions, with three irrigation rates (100% field capacity, 50% field capacity, and 25% field capacity), three Si NP levels (0, 50, and 100 ppm), and three MeJA levels (0, 20, and 40 μmol L −1 ). Drought stress (with irrigation at 25% FC) significantly reduced the growth of the plant, pod yield, crude protein, crude fat, fiber, dry weight, and mineral composition of the edible cluster bean pods. Pod weight reduction of approximately 50% occurred when they were irrigated at 25% FC, compared to pod yield from well‐watered plants. The addition of Si NPs and MeJA, when applied together, was able to reduce the adverse effects of low and moderate drought on pod yield and improve the productivity and nutritional quality of the edible cluster bean pods. The combined application of 100‐ppm Si‐NPs and 40‐μmol L −1 MeJA was the most effective treatment for improving the productivity and nutritional quality of vegetable cluster bean under deficit irrigation. These findings suggest that this combination for cluster bean cultivation under moderate drought conditions after economic feasibility and field‐scale validation studies.
ABSTRACT Drought severely limits sunflower ( Helianthus annuus L.) productivity, necessitating sustainable microbial strategies to enhance plant resilience. Endophytic bacteria modulate plant stress responses through diverse plant growth‐promoting (PGP) traits. This study characterized Pseudomonas protegens M4 and Citrobacter braakii M34 and evaluated their individual and combined effects on sunflower drought tolerance. Greenhouse experiments assessed inoculation with M4, M34, and their consortium (M4 + M34) under optimal (100% field capacity, FC) and drought conditions (75%–25% FC). Initial taxonomic identification based on 16S rRNA gene sequencing was further refined using whole‐genome sequencing (WGS) for high‐resolution classification and functional analysis. Genomic annotation and antiSMASH analysis revealed key PGP pathways in both strains, including enzymes involved in ethylene regulation, phosphate solubilization, biosynthesis of auxin‐like compounds, osmoprotectant production, and reactive oxygen species detoxification, alongside shared and strain‐specific secondary metabolite gene clusters. Under drought conditions, M4 primarily enhanced growth‐related traits such as shoot development and leaf area, whereas M34 was associated with physiological stress tolerance indicators. The consortium M4 + M34 exhibited synergistic effects, significantly increasing the chlorophyll content, relative water content, and proline accumulation while reducing electrolyte leakage. Principal component analysis further confirmed the consortium's superior performance. These findings highlight the potential of genomically and functionally characterized endophytic bacterial consortia as effective bioinoculant strategies for improving sunflower productivity under water‐limited conditions.
Meloidogyne graminicola is a destructive and widespread pest in tropical rice-growing systems, whereas resistance in Oryza sativa remains exceptionally rare. A major quantitative trait locus (QTL) associated with resistance has previously been identified at the distal end of chromosome 11 in the resistant varieties, LD 24 (Sri Lanka) and KPM (Thailand). In this study, we combined bioinformatic and molecular approaches to characterize this locus and phenotypically screened a collection of Sri Lankan landraces to investigate the distribution and potential origin of resistance. We identified a rare 7-bp deletion in LOC_Os11g44580 completely associated with the resistance phenotype, along with a notable occurrence of heterozygosity at this locus in some accessions. The deletion is located within a genomic region recently reported to harbor the resistance gene MG1 in the Chinese cultivar Zhonghua 11. Among 121 Sri Lankan landraces screened using the 7-bp deletion marker, eight accessions carried the deletion, whereas 12 were heterozygous; all exhibited resistance to Mg. Notably, the majority of resistant landraces belonged to the distinct Sri Lankan subgroup indica 2 and were predominantly collected from the Northern Province of Sri Lanka, suggesting a potential geographic origin of this resistance-associated allele. This study identifies 20 novel resistant varieties and highlights a regional concentration of resistance, providing valuable resources for breeding and insights into the evolutionary basis of nematode resistance in rice.
ABSTRACT Integration of internal metabolism with environmental cues is essential for successful establishment and survival of seedlings, with light an important mediator of the transition from skotomorphogenesis to photomorphogenesis. However, detailed knowledge of the metabolic events associated with this critical transition in developing tissues of barley germlings/seedlings is currently lacking. We systematically mapped the temporal and tissue‐resolved metabolic reprogramming in seedlings grown under light/dark photoperiod and complete darkness, using nontargeted high‐resolution LC–MS metabolomics to characterize seeds, shoots, and roots at 3–9 days postimbibition. Dark‐grown seedlings displayed allocation of resources to length rather than density. Multivariate analyses revealed distinct, light‐induced metabolic reprogramming in each tissue type. Among 192 annotated metabolites, seed metabolism was dominated by mobilization and pre‐emptive defense priming, with roots developing energy‐centered networks. In darkness, shoots directed metabolism toward rapid elongation and synthesis of defensive hordatine isoforms A and B, alongside linolenic acid and tryptophan derivatives. Light triggered metabolic switches where shoots allocated resources toward photoprotective flavonoids and antifungal hordatine C. This work provides the first spatiotemporal metabolic atlas of metabolic awakening in germinating barley seeds and seedling establishment based on data‐driven insights into metabolic pathways. It redefines skotomorphogenesis as an active escape‐and‐defense strategy and offers insight into photomorphogenesis in support of tissue‐specific metabolomic differentiation.
ABSTRACT Sulfur (S) is an essential macronutrient that plays critical roles in plant growth, redox homeostasis, stress adaptation, and immunity. In rice ( Oryza sativa ), sulfur deficiency is becoming increasingly common due to modern agricultural practices, yet the molecular regulation of sulfur metabolism remains incompletely understood. Recent advances in functional genomics have provided new insights into the mechanisms governing sulfur uptake, transport, assimilation, and downstream metabolic regulation. This review synthesizes current knowledge of sulfur metabolism in rice, with particular emphasis on transgenic and microRNA‐based approaches. We discuss the functions of sulfate transporters, sulfur assimilation enzymes, and sulfur‐containing metabolites such as glutathione in regulating sulfur homeostasis, redox balance, and stress responses. Particular attention is given to the conserved miR395 regulatory module, which coordinates sulfur assimilation and allocation through posttranscriptional regulation of ATP sulfurylase genes. Evidence from functional genomics studies demonstrates that sulfur metabolism is closely integrated with redox signaling, stress tolerance, and plant immunity. Collectively, these findings support a view of sulfur metabolism as a dynamic regulatory network rather than a linear nutrient‐assimilation pathway. We further highlight key knowledge gaps, emerging research opportunities, and future directions for applying functional genomics to improve sulfur use efficiency and resilience in rice. This review provides a framework for advancing both the fundamental understanding and translational application of sulfur metabolism in sustainable rice production.
ABSTRACT Bicellular pollen often enters a desiccated state wherein development is temporarily arrested, enabling long‐term survival while maintaining the capacity for rapid activation upon hydration. Although the generative cell in mature pollen of Lilium species is known to have completed DNA replication, its precise cell‐cycle status and the requirement for de novo transcription during pollen tube growth remain unclear. In this study, we examined the cell‐cycle state and nuclear reactivation dynamics of the generative cell of Lilium longiflorum pollen using detailed confocal microscopy. We found that the generative cell nucleus in mature pollen exhibited prophase‐like chromatin architecture but rapidly underwent transcriptional reactivation and chromatin decondensation upon hydration. This response is inconsistent with M‐phase progression, which is irreversible and does not permit transcriptional reactivation or chromatin decondensation before chromosome segregation. Immunofluorescence analysis of cell‐cycle markers showed that the generative cell lacked M‐phase‐specific features and instead displayed an interphase‐like marker profile. Transcription inhibition experiments further revealed that chromatin decondensation and histone phosphorylation occurred independently of transcription, whereas generative cell division required de novo transcription. Together, these results define a unique nuclear state in which highly compacted chromatin is maintained while cell‐cycle progression remains arrested in G2 and demonstrate that hydration triggers nuclear reactivation through sequential transcription‐independent and transcription‐dependent processes.
Low temperatures are a major environmental stress that limits plant growth and development. Understanding the molecular mechanism of cold tolerance is therefore essential for improving crop performance through molecular breeding. In this study, we focused on CISP, a small, previously uncharacterized protein, specifically induced in barley roots under low temperature conditions, and investigated its potential role in cold tolerance. Heterologous expression of CISP in Escherichia coli enhanced late-phase bacterial growth at low temperatures compared with the wild-type strain. Similarly, overexpression of CISP in Arabidopsis thaliana, which lacks an apparent ortholog, improved seedling growth under low temperature conditions. These results suggest that CISP can promote growth of heterologous organisms used in this study at low temperature environments. To investigate its molecular function, we performed RNA chaperone assays using RNA beacons that tend to form stable secondary structures. CISP reduced the formation of RNA secondary structures and facilitated their destabilization, indicating RNA chaperone-like activity. CISP is a basic, low-molecular-weight protein containing an intrinsically disordered region (IDR) at its N-terminus but lacking canonical RNA-binding domains such as the cold shock domain (CSD) or RNA recognition motif (RRM). Our findings therefore suggest that CISP may represent a previously uncharacterized type of RNA chaperone. This study provides new insights into the structure and function of CISP and its potential contribution to cold tolerance in barley.
ABSTRACT Plasmodiophora brassicae (Phytomyxea, Rhizaria) is the etiological agent of clubroot disease, one of the most important diseases of Brassicaceae crops. Alteration of metabolism and hormone homeostasis leads to the formation of tumor‐like galls in the roots of affected plants. Host plant energy metabolism, defense, and developmental processes are under strong temporal control and the very same processes are affected by clubroot. For the first time, this study uses time‐resolved transcriptome analyses to explore how P. brassicae affects Arabidopsis thaliana in the night during intermediate (14 days after inoculation; DAI) and late (21 DAI) infection. Day–night differences in gene expression were more pronounced in younger rather than older plants in our differential gene expression (DGE) analysis. Consequently, intermediate phases of infection showed more day–night differences than later ones. Clustering of differentially expressed genes (DEGs) in functional categories highlighted how some of the typical processes known to be disrupted by clubroot infection are more significantly affected in the night and also uncovered some disrupted exclusively in the night. RNA modification stood out as the most unambiguously upregulated process in infected Arabidopsis roots in the night. Analysis of the interaction between clubroot infection and diel oscillations in gene expression detected modifications in the rhythmicity of central circadian clock components during the infection. We discuss our findings in the context of manipulation of plant defense and metabolism, identifying targets for experimental validation and highlighting potential new lines of investigation of our time‐resolved datasets to better understand the interaction between P. brassicae and its host.
ABSTRACT Small signaling peptides (SSPs) are important regulators of plant growth, development, and responses to biotic and abiotic stress, yet their role in the C4 grass Sorghum bicolor is largely uncharacterized. To help fill this knowledge gap, 219 sorghum genes that encode SSPs were identified based on SSP sequences previously identified in Arabidopsis thaliana, Zea mays, Oryza sativa, Triticum aestivum, and Brachypodium distachyon. The 219 sorghum SSP‐encoding genes were assigned to 19 gene families, analyzed for the presence of motifs, and aligned with genes that encode SSPs in other plants using phylogenetic analysis. Sorghum genes in 12 of the 19 SSP gene families had not been previously characterized. Expression of the 219 SSP‐encoding genes in sorghum organs, during stem development, and in stem tissues and cell types revealed distinct spatial, temporal, and developmental patterns of expression. Genes associated with the SbCEP and SbRGF families were preferentially expressed in roots, whereas SbEPF genes were expressed in stem epidermal and pith parenchyma cells and panicles. The expression of genes during bioenergy sorghum stem growth and development was investigated because stems account for ~80% of harvested biomass and serve as conduits for water and nutrient transport between leaves and roots. During stem development, 28 SSP genes in several families (CLE, EPF, CEP, GASS, PSY, ES, PSK, CAPE, POE) were expressed at higher levels in zones of cell proliferation. For example, the TDIF homologs SbCLE41 and SbCLE42 were expressed at high levels in nascent stem nodes where they may regulate vascular bundle cambial activity and cell differentiation. A different set of 15 genes in the CIF, POE, CAPE, PSY, CEP, RALF, and CLE families were expressed at higher levels in zones of stem tissue differentiation highlighted by elevated expression of five SbRALFRs in the stem nodal plexus. Cell type–specific expression of many sorghum genes that encode SSPs was observed in fully elongated internodes indicating gene expression is regulated with high spatial resolution. Overall, the results provide a foundation of information for analysis of SSP function in sorghum that can be integrated with knowledge of sorghum gene regulatory networks to modulate traits important for production of sorghum crops.
ABSTRACT Virus‐induced gene silencing (VIGS) is a method that exploits plant antiviral defense mechanisms to downregulate endogenous genes. The technique is versatile, rapid, and widely used for functional genomics studies. Here, we report a method for VIGS in the medicinal plant, Calendula officinalis (pot marigold). This species produces anti‐inflammatory triterpenoids and has also been bred and cultivated as an ornamental plant. We describe a method for the injection of Agrobacterium tumefaciens cultures into leaf midribs and compare visual marker genes for tracking VIGS utilizing constructs that simultaneously target visual marker and target genes. We use these tools to demonstrate that silencing a gene encoding cycloartenol synthase results in changes to leaf phytosterols. This method could be used to further investigate the genetic basis of specialized metabolism in this species and could be adapted to other members of the Asteraceae family, many of which are of economic and chemical value.
Peanut reproduction is foundational for crop yield, breeding, and evolution. However, gene regulation underlying peanut flowering pattern and timing has received limited attention. Cultivated peanut (Arachis hypogaea L.) shows two distinct flowering patterns between two subspecies, with ssp. hypogaea lacking flowers on the main stem and ssp. fastigiata having them. Understanding the gene regulatory networks that control peanut flowering will inform the genetic pathways impacting peanut reproduction, phenology, and yield. To this end, we measured whole-transcriptome gene expression of leaves and shoot tips (meristem) at six plant growth stages from Tifrunner, a peanut cultivar belonging to ssp. hypogaea, and GT-C20, a peanut germplasm belonging to ssp. fastigiata. Overall gene expression was distinct between the two genotypes in both tissue types. Flowering regulators including AhFT, AhSOC1, AhAGL42, and AhSPL3 were differentially expressed in both the main and lateral stem at the time of flowering initiation (T3-first bloom). This indicates that positive regulation of these flowering regulators drives the distinct pattern of flowering on the main stem in GT-C20. Meanwhile, the differential expression of two RING-finger E3 ubiquitin ligases was identified between the two genotypes, indicating that the PAF1-complex (PAF1C) may contribute to the lack of flowering on the main stem of Tifrunner. Gene co-expression network analysis indicates that gibberellic acid (GA) and jasmonic acid (JA) pathways are involved in reproductive regulation. These results provide insight into how flowering physiology is differentially controlled between the two peanut subspecies and provide a launching point for additional research in peanut floral development.
ABSTRACT The chemical phenotype (chemotype) of Cannabis sativa is defined by the ratio of cannabidiolic acid (CBDA) to Δ9‐tetrahydrocannabinolic acid (THCA). Although the Mendelian segregation of these traits suggests a single‐locus biallelic system, recent sequencing and phylogenetic evidence indicate they are encoded by two distinct, tightly linked genes. The precise genomic architecture of this region, known as the B locus, has remained poorly defined. In this study, we analyzed recently released high‐quality Cannabis reference genomes to resolve the structure of the B locus. Our results demonstrate that this region functions as a supergene, characterized by suppressed recombination that facilitates Mendelian‐like switching between phenotypic states. Comparative genomic analysis reveals substantial structural polymorphism within the locus, including significant variations in gene copy number and large‐scale insertions/deletions (indels). Furthermore, we functionally characterized three previously unstudied members of the cannabinoid oxidocyclase family. We find that these enzymes primarily catalyze the production of cannabichromenic acid (CBCA), reinforcing the model that cannabinoid profile is dictated specifically by the presence and expression of THCAS or CBDAS. Finally, we mapped the expression profile of the entire berberine bridge enzyme (BBE) family, identifying widespread expression across plant tissues, including in glandular trichomes. Collectively, these findings resolve the genomic architecture of the B locus, clarify the enzymatic basis of cannabinoid profile determination, and establish a framework for understanding the evolutionary maintenance of chemotype diversity in C. sativa.
ABSTRACT Sulfur (S) is an important nutrient that has wide‐ranging effects on plant health and metabolism. Several classes of transcription factors respond to S deprivation, including R2R3‐MYBs. In Arabidopsis, the AtMYB93 transcription factor‐encoding gene is upregulated by S deprivation. AtMYB93 has a non‐redundant function in lateral root development and redundant functions regulating suberin biosynthesis alongside related MYB transcription factors. Whether AtMYB93's potential role in plant S responses relates to lateral root development, and the extent to which either function is conserved outside Arabidopsis, are unknown. We show that a tomato MYB93 homolog (SlMYB93) can inhibit lateral root development in Arabidopsis. Putative MYB93 homologs in many dicots show root enrichment and tomato MYB93‐related genes show upregulation by S stress. The transcriptome of Atmyb93 mutant roots implicates AtMYB93 in responses to S, while elemental analysis demonstrates that the Atmyb93 mutant has elevated shoot S levels while tomato SlMYB93‐overexpressing plants have reduced shoot S. We uncover a stimulatory effect of S deprivation on Arabidopsis adventitious root development. However, Atmyb93 mutants do not show significant changes in sensitivity to S with respect to lateral or adventitious root development, most likely due to some functional redundancy. Moreover, AtMYB93 promoter activity is not spatially regulated by S deprivation. Taken together, our data suggest that MYB93 may have a role in mediating the regulation of S levels alongside other root transcription factors.
ABSTRACT Herbicide target site resistance in polyploid species is more complex than in diploids due to potential subgenome interactions. This study characterized mutations in the ALS gene across distinct subgenomes of hexaploid Echinochloa crus‐galli and evaluated the cross‐resistance patterns conferred by each mutation to various ALS‐inhibiting herbicides. E. crus‐galli populations were screened, and dose–response curves were performed with ALS inhibitors from different chemical groups. The ALS gene copies of each subgenome (A, B, and C) were sequenced. Copy number variation, global relative expression, and the specific relative expression of ALS gene from each subgenome were performed. The mutations Ala122Thr, Ala205Asn, and Ser653Asn conferred resistance only to imazethapyr, whereas Trp574Leu to imazethapyr, penoxsulam, bispyribac‐sodium, and nicosulfuron, when considered the label rate. ALS mutations were more frequent in subgenome A, but ALS from subgenome C had the highest expression. Biotypes with the same mutation showed different resistance level to herbicides. The biotype SAOJER‐01 had Trp574Leu mutation in subgenome C and was 22 times more resistant to imazethapyr and penoxsulam than CAMAQ‐01, which had the same Trp574Leu mutation in subgenome A. Both SAOJER‐01 and CAMAQ‐01 biotypes showed CYP450 metabolism mediating penoxsulam resistance in addition to the target site mutation. In conclusion, the mutations Ala122Thr, Ala205Asn, Trp574Leu, and Ser653Asn confer resistance to imazethapyr, but only Trp574Leu confers resistance to the other chemical groups. The herbicides penoxsulam, bispyribac‐sodium, and nicosulfuron are effective in controlling three out of four mutations. CYP450‐mediated metabolism coexists in biotypes carrying the Trp574Leu mutation. The subgenome location of the ALS mutation may result in variable levels of resistance.
Plant diseases are the cause of heavy losses of crop production and, therefore, a big contributor to food shortages. Identifying these diseases as early as possible is important to limit the negative effects that these diseases have on the yields, as slow response time will lead to the spread of diseases and further loss. Traditionally, trained staff will go into the fields, multiple times during the growth period, and inspect the plants in samples through field disease monitoring. These traditional processes are time-consuming and costly, and can be error-prone, if the staff is not properly educated or if the staff simply makes mistakes due to oversight, for example. To aid farmers with the process of correctly identifying diseases, artificial intelligence deep learning methods have been employed in recent years. However, to train such deep learning models, one needs to obtain sufficiently large and high-quality datasets and a model architecture that is capable of extracting relevant features to accurately classify the plant leaves. Datasets are still a limitation in the field of plant leaf disease classification. As such, domain adaptation methods such as transfer learning are often employed to overcome this data shortage. However, in current research, these domain adaptation methods almost exclusively rely on ImageNet as the pretraining dataset, a dataset that is domain unrelated to plant leaf disease detection, and models are often left unmodified and un-optimized as a result. In this work, we propose the pretraining of an improved attention-based and SiLU-activated DenseNet201 architecture called PLDC-Net that is pretrained on a large-scale plant leaf disease dataset constructed by the authors to create a domain-specific base model for better domain adaptation to new plants and diseases, validating the improved results through transfer learning, fine-tuning, one-shot learning, and few-shot learning. PLDC-Net has managed up to just over 24% improvements in F1-Score over the baseline in domain adaptation results.
A systems biology approach was used to characterize Monotropa uniflora, a nonhotosynthetic, myco-heterotrophic plant. In contrast to autotrophic plants, myco-heterotrophic plants obtain carbon and other nutrients by connecting roots to fungi, which establish the physical connection to a photosynthetic host, and thereby source required nutrients. Although a large proportion of plants form mycorrhizal associations, heterotrophic plants that are completely dependent on the mycorrhizal association for nutrients are rare and less well studied. The potential of developing M. uniflora as a model for myco-heterotrophic plants is demonstrated by datasets collected within this study, which include cellular ultrastructural morphologies, metabolomes and transcriptomes from tissue-types that are at different stages of growth or development. The morphological comparisons indicate that cells of the lower stem are older than those of the upper stem. The molecular -omics datasets reveal greater differences as the result of development rather than growth. Despite the obvious absence of photosynthetic functions in M. uniflora, as in photosynthetic plants the most abundant metabolites are sugars and organic acids that are used to translocate carbon in the latter. KEGG pathway enrichment analysis of the transcriptomes indicates that catabolic processes are highly active, which is consistent with the hypothesis that these processes generate intermediary energy metabolites (i.e., ATP and NAD(P)H) that can support anabolic processes associated with growth or development. Correlative analyses of the abundance of cutin monomers or cuticular waxes relative to the expressed transcriptomes identified co-expressing genes, revealing coordinated lipid metabolism network(s) that support cuticle formation in the absence of photosynthetic energy production. This model systems biology study of a myco-heterotrophic plant offers baseline insights into the functioning and resilience of ecological niches that are increasingly threatened by anthropogenic pressures.
Upstream open reading frames (uORFs) are regulatory elements present in the 5' leaders of mRNA that can significantly impact downstream gene expression in eukaryotes. In crop engineering, editing of uORFs can provide an avenue to upregulate expression of native genes without the need to add persistent transgenic copies. Even with genome-wide methods to identify translated uORFs such as ribosome profiling, their functional characterization depends on validation through reporter gene assays and mutagenesis studies. Current screening methods for plants use luciferases or protoplasts to measure differential gene expression between wild-type and mutated transcript leaders, which requires tissue processing and/or substrate addition. Here, we present a time- and cost-efficient alternative to investigate transcript leaders by co-expression of two fluorescent proteins in Nicotiana benthamiana leaf tissue and test our assay on genes involved in photoprotection, editing of which could provide a pathway to increase CO2 assimilation during sun-shade transitions.
ABSTRACT The growing demand for rare‐earth elements (REEs), particularly dysprosium (Dy), underscores the need for sustainable extraction methods. Recovery of Dy, particularly from geographically distributed waste sources, is challenging. This gap positions phytomining, a technique using plants to accumulate metals, as a promising alternative. However, plant species differ in their ability to accumulate metals in high concentrations, necessitating efficient screening methods. In this study, we developed a high‐throughput fluorescence‐based assay to detect and quantify Dy uptake in plant tissues. The Dy detection method described in the present work exploits Dy's unique spectroscopic properties for sensitive and efficient analysis, enabling the detection of concentrations as low as 0.07 μM, with a detection limit of 0.2 μM in a plant matrix. By incorporating sodium tungstate (Na2WO4) as a fluorescence enhancer, we achieved robust emission intensities at 480 and 580 nm, facilitating Dy quantification in complex plant matrices. Additionally, the use of time‐resolved fluorescence techniques reduces background autofluorescence from plant tissues, enhancing signal specificity. Validation of the fluorescence method with inductively coupled plasma mass spectrometry (ICP‐MS) demonstrated a strong correlation in Dy levels. Greenhouse trials confirmed the method's utility for screening Dy accumulation in living plants and highlighted the potential for rapid stand‐off detection. This fluorescence‐based approach offers a scalable, efficient tool for identifying Dy‐accumulating plants and advances phytomining as a sustainable strategy for REE recovery.
In terrestrial plants, drought stress activates abscisic acid (ABA) signaling in guard cells, prompting stomatal closure to reduce water loss. Stomatal closure is accompanied by reorganization of the microfilaments. However, the mechanism by which ABA signaling regulates microfilaments disassembly remains unclear, and the actin-binding proteins (ABPs) involved in this process have yet to be fully identified. In this study, we demonstrated that FIMBRIN2 (FIM2), an actin-bundling protein, has its expression in guard cells upregulated by ABA, and FIM2 is involved in ABA-induced stomatal closure. The fim2 mutant exhibits a drought-sensitive phenotype and delayed stomatal closure. Further confocal microscopy observations confirmed that this delay results from impaired actin bundling and reduced actin turnover activity. These findings reveal FIM2's role in ABA-induced stomatal closure and deepen our understanding of the functions of ABPs and the actin cytoskeleton in plant stress adaptation.
