Nature Plants

Introductory Paragraph (Nature Plants format)Parthenogenesis or fertilization-independent embryogenesis occurs at low frequency in sexual plants. Expression of BABYBOOM-like (BBML) and PARTHENOGENESIS (PAR) genes in the egg cell of several diploid dicot crops induce parthenogenesis at varying frequency; however, recovery of viable haploid seeds has rarely been reported, perhaps due to a lack of viable endosperm formation. In the legume cowpea (Vigna unguiculata L. Walp), ectopic egg cell expression of the endogenous BBML homolog (VuBBML1) and PAR from Taraxacum officinale induces parthenogenesis; however, seeds abort as endosperm formation is blocked following self-pollination. Expression of VuBBML1 in both the egg cell and central cell, together with central cell fertilization following self-pollination, results in viable seeds that germinate and give rise to haploid plants. VuBBML1 has a functional role in the formation of cowpea embryo and endosperm seed compartments. This finding opens possibilities for establishing double haploid production during homozygous parental breeding, and asexual seed induction for fixing hybrid vigor in cowpea.

Plant cell wall enlargement is fundamental to crop productivity and its sensitivity to drought1. Tip growth and diffuse growth are contrasting wall enlargement patterns often proposed to be limited by different processes: localized secretion and remodeling of pectins for tip growth versus loosening and sliding of cellulosic networks by -expansins (EXPAs) for diffuse growth2,3. Here, we knocked out root-hair specific EXPA7 and EXPA18 in Arabidopsis, abolishing root-hair tip growth which was restored by complementation with genes from some, but not all, expansin clades. Notably, EXPA13 and EXPA20 failed to complement; they belong to two ancient clades lacking a highly conserved Asp considered essential for expansin activity. Mutation of this Asp in EXPA7 confirmed its requirement for wall enlargement. EXPA-mCherry fusions revealed widely contrasting patterns of subcellular trafficking and wall-binding for different EXPAs. The results demonstrate an essential EXPA requirement for root-hair tip growth and uncover a greater diversity of expansin functions than previously recognized.

Meiotic recombination ensures accurate chromosome segregation and promotes genetic diversity by generating crossovers between homologous chromosomes1. While essential in most sexually reproducing organisms, recombination is variably regulated and can be absent in some lineages, a condition known as achiasmy2. However, obligate achiasmy in both sexes of a sexual species has not been previously documented. Here, we investigate the beak-sedge Rhynchospora tenuis, a holocentric plant with the lowest known chromosome number among flowering plants (n = 2) and inverted meiosis3. Using chromosome-scale genome assemblies from nine accessions, molecular cytogenetics, immunocytochemistry, high-throughput single-gamete sequencing and whole-genome sequencing of controlled crosses, we show that R. tenuis undergoes obligate, genome-wide achiasmy in both male and female meiosis. Despite normal early meiotic axis formation, synapsis fails, crossovers are not detected cytologically or genetically, and univalents persist at metaphase I. Extensive haplotype-specific accumulation of transposable elements (TEs) generates segregation distortion (e.g. meiotic drive), favouring the transmission of larger, TE-rich chromosomes. Remarkably, sexual reproduction is retained with fertilisation producing viable seeds only when translocation-compatible gametes meet, indicating strong post-meiotic selection that eliminates incompatible homozygous combinations. As a result, all surviving offspring are genetically identical to the maternal genotype, effectively restoring heterozygosity each generation and mimicking clonal reproduction. We propose that the combined effects of recombination loss, low chromosome number, holocentricity, inverted meiosis, and selective transmission of longer chromosomes enable faithful segregation and clonal-like inheritance despite sexual reproduction. These findings challenge the boundary between sex and clonality, revealing a unique evolutionary strategy linking genome architecture, recombination loss, and transmission bias.

The Morelloid clade (black nightshades) is one of the most strongly supported clades within the megadiverse Solanum genus. It comprises 76 globally distributed, non-spiny herbaceous and suffrutescent species. While often erroneously considered poisonous weeds, several species are economically important as orphan crops. The clade is closely related to tomato and potato but, due to a lack of focused breeding efforts, remains a reservoir of genetic diversity for crop improvement. Despite this potential, we lack fundamental knowledge on the evolution of the Morelloid clade. The group includes polyploid species with unknown parental origins--likely reflecting reticulate processes such as hybridization, introgression, and associated backcrossing events. Prior analyses have been unable to disentangle these processes, leaving the mechanisms underlying reticulate evolution in the Morelloid clade poorly understood. Here, we use genome skimming to produce a well-supported maximum likelihood plastid phylogeny from complete circularized plastomes and a coalescent-based species tree from combined Angiosperms353 and conserved ortholog set nuclear markers. Our dataset, composed of previously published data and deep genome skimming from herbarium samples, spans 26 Morelloid species. To investigate patterns of non-treelike evolution, we used a nuclear phylogenetic network, multispecies coalescent simulations, a fused rooted nuclear chloroplast tree, and quantification of nuclear gene tree concordance. We show that incongruence between nuclear and plastid trees is pervasive and cannot be explained by incomplete lineage sorting alone. Instead, our results demonstrate that events consistent with repeated chloroplast capture have shaped the reticulate evolutionary history of the clade, especially among African polyploid and Pan-American diploid lineages.

Arbuscular mycorrhizal fungi (AMF) establish a dynamic and asynchronous symbiosis with a wide range of land plants, involving distinct stages of root colonization and associated cellular responses that co-occur within the same root. Whilst decades of research have significantly advanced our understanding of the plants symbiotic gene repertoire, this spatial and temporal complexity has hindered a detailed dissection of the molecular mechanisms underlying fungal accommodation. Here, we present the first single-nucleus RNA-sequencing (snRNA-seq) dataset of Solanum lycopersicum roots colonized by Rhizophagus irregularis. Unsupervised subclustering of an AM-specific cell population resolves AM-responsive root epidermal cells as well as a developmental gradient of cortical cells across distinct stages of arbuscule formation, unveiling stage-specific transcriptional signatures during AMF colonization. Moreover, using Motif-Informed Network Inference based on single-cell EXpression data (MINI-EX), we put forward candidate transcription factors orchestrating these stage-specific transcriptional programs. Together, our data support novel hypotheses on how diverse plant developmental and physiological processes - including localized cell cycle reactivation and the integration of multiple nutritional cues - are coordinated to facilitate the establishment of a functional symbiosis. As such, this high-resolution dataset serves as a valuable resource for candidate gene prioritization and future reverse genetic studies.

In plant cells, the multi-domain proteins FRIENDLY and REC regulate the cellular organization, distribution and proliferation of mitochondria and plastids, respectively. Both proteins share a similar overall domain architecture and belong to the larger CLUSTERED MITOCHONDRIA (CLU) superfamily. Domains of CLU proteins have been shown to interact with translation related proteins, tRNA synthetases and even mRNA, but their exact modes of operation remain cryptic and how organelle specificity of CLU paralogs in plant cells is achieved unknown. We characterized the single CLU family protein of the liverwort Marchantia polymorpha that we demonstrate to be transcribed either with or without exon 22, which changes the configuration of the TPR domains in the C-terminus. Knockout of MpCLU affects both mitochondria and plastids, and independent rescues show that the splice variant with exon 22 (MpCLU22) serves mitochondrial- and the one lacking exon 22 (MpCLUspl22) plastid biology. The CLU-C domain of the protein is responsible for nuclear localisation and expressed alone induces a phenotype that differs in photosynthesis performance and transcriptome changes from that of the knockout of MpCLU. Our results identify the C-terminal TPR motif to be responsible for organelle specificity in plants and they provide an example of how genome reformatting and gene loss can be compensated for by the alternative splicing of a single exon.

Chloroplast proteostasis is vital for plant development, yet whether cells can actively reprogram organelle communication to restore plastid function when essential protease components fail remains unclear. Using a forward genetic suppressor screen in Arabidopsis, we identify loss of the mitochondria-associated protein FRIENDLY MITOCHONDRIA (FMT) as a strong suppressor of the virescent and growth-retarded clpc1 mutant, which lacks the major chloroplast Clp chaperone ClpC1. Suppression is highly specific and occurs independently of GUN1-mediated retrograde signaling. Integrated multi-omics analyses reveal that clpc1 and fmtclpc1 represent two distinct organelle signaling states. In clpc1, loss of ClpC1 triggers a plastid stress state characterized by repression of photosynthesis-associated transcription factors, induction of plastid metabolic stress markers, and impaired proteolytic activity. By contrast, loss of FMT shifts the system into a recovery state despite persistent mitochondrial clustering. Mechanistically, FMT negatively regulates CLPC2, a ClpC1 paralog, and fmt-mediated rescue results from CLPC2 derepression. Moderately elevated ClpC2 restores in vivo proteolysis, as evidenced by recovery of PAA2 substrate turnover, normalization of chloroplast ultrastructure, and reactivation of photosynthesis-related gene expression. Transcriptomic and proteomic profiling further reveal coordinated remodeling of nuclear gene expression and chloroplast protein investment in the recovery state, including reduced cytosolic folding stress and selective induction of jasmonic acid- and salicylic acid-associated signaling networks. Genetic analyses establish that REC1 and REC2 are required for full CLPC2 induction and phenotypic recovery. Together, our findings uncover a latent inter-organelle compensatory mechanism in which mitochondrial perturbation reprograms nuclear gene expression to restore chloroplast proteostasis when ClpC1 function is compromised.

Plant terrestrialization necessitated that a barrage of stressors had to be overcome1. Land plants use an integrated response network in adjusting their molecular physiology to terrestrial stressors2--one of the foremost being UV irradiance. The zygnematophytes are the closest streptophyte algal relatives of land plants3-5, are renowned for their resilience to UV stress6-8, and thus allow to glean key information for inferring the UV response toolkit of the earliest land plants9,10. Throughout streptophyte evolution, specialised metabolism radiated into creating diverse compounds used for responses to environmental challenges, such as sun-shielding compounds and antioxidants11-14. This includes UV-shielding compounds like flavonoids and coumarins but also the land plant specific polymer lignin, giving structural support in vascular plants15; homologs of the underpinning core pathway occur in streptophyte algae16. Here, we exposed the zygnematophyte Mesotaenium to UV-B irradiation and profiled its physiology, morphology, transcriptomics as well as metabolomic features. After UV-B exposure, cells showed rapid photophysiological responses and progressively growing terminal vacuoles. Our transcriptome data capture dynamic changes in gene expression of (i) core downstream responses such as genes homologous to phenol metabolic enzymes, photophysiological homeostats, and DNA repair factors; but also (ii) upstream components featuring key homologs of kinase-mediated signalling cascades, as well as light quality and abscisic acid-mediated signalling components. To scrutinize the acclimatory chassis, we created a metabolite feature database specifically for the Mesotaenium metabolome. Upon UV-B exposure, the metabolome displayed pronounced temporal shifts, with several phenolic features that accumulate along the stress-acclimation kinetics. Overall, we capture a chemodiverse response including various phenolics such as purpurogallin-like, methoxypsoralen-like derivatives and coumarins. Our data establish an integrated model for UV responses in the closest algal relatives of land plants, shedding light on the toolkit that allowed the progenitors of land plants to move out of a protective water column.

Mechanical stimuli provoked by wind, soil compaction, rainfall, and biotic interactions strongly influence plant phenotype, growth, and development. Previous studies indicated that weight-treated Arabidopsis thaliana plants increase stem diameter, vascular bundle number, and seed yield, involving auxin, brassinosteroid, and strigolactone-related genes. In this work, we investigated how the mechanically induced increase in phloem area improves source-to-sink partitioning, while the increase in xylem area negatively affects long-term drought tolerance. Transcriptomic profiling confirmed a large-scale reprogramming of drought-responsive genes in treated plants. Moreover, quantification of sucrose and starch content highlighted an enhanced synthesis and carbohydrate transport, which ultimately and positively impacted lipid and protein contents in seeds. Using loss-of-function mutants, we demonstrate that the phloem loader SUC2 and exporters SWEET11, 12, and 16 are essential for the yield gains triggered by mechanical stress. Furthermore, mechanical treatment alters sugar metabolism. Overall, our findings indicate that weight treatment elicits a complex physiological response, in which sucrose transporters and starch metabolism play a crucial role in mediating its positive effects on seed quality and yield. Significance statementMechanical cues are ubiquitous in natural environments, but their impact on plant carbon allocation and yield remains poorly understood. This study reveals that mechanical stress reshapes vascular architecture and carbohydrate transport, enhancing source-to-sink partitioning and seed quality in Arabidopsis. By identifying sucrose transporters and sugar metabolism as key mediators of mechanically induced yield gains, our findings provide mechanistic insight into how physical forces integrate with metabolic regulation influence plant productivity.

Climate change poses a major threat to humanity by driving biodiversity loss and reducing crop yields1,2. To understand the molecular and developmental impacts of rising temperatures, plant science has relied heavily on the model organism Arabidopsis thaliana. Despite decades of research, its development under fully natural conditions remains poorly understood, and only [~]30% of genes have experimental functional annotations, largely because many functions are subtle or manifest only in specific laboratory or ecological contexts3. Here, we address this gap with a landscape transcriptomic approach that integrates intensive phenotyping and transcriptomic profiling of naturally occurring plants in their native habitats4. Across two contrasting field sites and five growing seasons (2021-2025), we phenotyped more than 3,000 A. thaliana plants and generated >1,600 matching transcriptomes. The resulting >30,000 quantitative trait measurements provide a unique opportunity to link climate fluctuations with plant traits and gene expression. Seasons characterized by extreme temperature anomalies directly influenced plant traits, and climatic variables together explained up to 17% of phenotypic variation. In situ transcriptomes carried clear temperature and local environmental signatures, closely matching temperature-response programs known from the laboratory. Leveraging paired per-plant transcriptomes and phenotypes, we applied machine learning to predict regulators of climate-relevant and other plant traits under natural conditions. The models recovered canonical thermomorphogenesis regulators, including PHYTOCHROME INTERACTING FACTOR 4 (PIF4)5,6, providing ecological evidence that temperature signaling pathways defined in controlled environments operate in the wild, and expanded this regulatory landscape by identifying hormonal receptors, signaling components, and previously uncharacterized genes, some of which we functionally validated. Together, this work demonstrates that landscape transcriptomics, by integrating natural field transcriptomes with phenotypes, and thus, capturing environmental and regulatory states, enables the predictive identification of genetic regulators of temperature responses and broader plant traits. This makes landscape transcriptomics a scalable framework for climate-aware functional genomics in plants.

Understanding how transcription factor binding site (TFBS) location influences gene regulation remains a fundamental challenge in plants. Here we integrate conserved TFBS with single-nucleus chromatin accessibility and expression datasets to resolve how TFBS location relates to regulatory function. Although TF binding patterns are conserved and frequently enriched near the TSS, positional enrichment of TFBSs poorly predicts cell type-specific gene expression. Instead, cell type-specific gene expression aligns with conserved TFBSs that are embedded in cell type-restricted chromatin, which show TF family-specific distributions across distal promoter and genic regions. In contrast, TSS-proximal TFBSs are implicated in rapid, tissue-wide transcriptional stress responses, as supported by hormone-induced gene expression analysis. Finally, distal upstream regions contain conserved TF clusters that overlap rare cell type-specific accessible chromatin and are highly enriched for genes controlling embryonic and meristem patterning, including auxin and other hormone pathway components. This positional partitioning of regulatory function indicates that TFBS location encodes distinct regulatory programs for TFs.

Plant immunity is activated by cell-surface and intracellular receptors that detect pathogen-derived molecules. Mutual potentiation between these two receptor types is essential for robust disease resistance, but the mechanisms underpinning integrated dual receptor immunity remain unknown. Here, we found that activation of the intracellular receptor, ZAR1, induces its site-specific modification by non-proteolytic ubiquitin chains. ZAR1-anchored ubiquitin chains promote oligomerization of ZAR1 into a calcium-permeable resistosome pore and are recognized by RH3, a ubiquitin-binding DEAD-box RNA helicase. Remarkably, RH3 recruits both ZAR1 resistosomes and cell-surface receptor components into a dual receptor complex, thereby enhancing calcium-dependent mRNA translation of core defense proteins and establishing robust immunity. These findings identify ubiquitin recognition as the missing link in mutual potentiation of plant immunity by cell-surface and intracellular receptors.

Riboflavin (vitamin B2; RF) and its derivatives flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) are indispensable cofactors for redox reactions in plants. While higher plants possess a conserved pathway for de novo riboflavin biosynthesis, how flavins are transported and spatially distributed between tissues remains unresolved. In particular, no genetically defined plasma membrane-localized flavin transporter has been identified in plants. Here, we identify Arabidopsis PURINE PERMEASE 5 (AtPUP5) as the first genetically defined plasma membrane-localized flavin transporter in plants. Using a riboflavin-auxotrophic yeast mutant, we show that AtPUP5 enhances cellular uptake of RF and, to a lesser extent, FMN, whereas FAD uptake is inefficient. In planta, AtPUP5 overexpression increases RF accumulation following external application. In contrast, loss-of-function mutants do not display defects in bulk RF uptake at the whole-plant level, indicating that AtPUP5 is not essential for global riboflavin acquisition. Notably, AtPUP5 deficiency results in RF overaccumulation in reproductive organs, including inflorescences, siliques, and seeds, irrespective of external RF supply. This organ-specific phenotype is fully suppressed by genetic complementation and coincides spatially with strong AtPUP5 promoter activity in reproductive tissues. These findings demonstrate that AtPUP5 functions at the plasma membrane to regulate localized riboflavin distribution in reproductive tissues. Together, our study establishes the first molecular framework for plasma membrane-mediated flavin distribution in plants and positions spatial regulation as a central component of riboflavin homeostasis in plants. Significance statementFlavins are essential vitamins that support plant metabolism, yet how they are transported and spatially distributed within plants has remained unclear. By identifying AtPUP5 as a plasma membrane-localized protein that mediates localized riboflavin distribution in reproductive organs, this study provides the first genetic insight into carrier-mediated flavin distribution in plants and reveals an unexpected layer of spatial control in plant vitamin metabolism.

The chloroplast division machinery, known as the division ring, is a supramolecular complex composed of bacterial- and host-derived proteins1-5. However, how the division ring generates the force required to sever a chloroplast remains poorly understood. Here, we established an in vitro assay in which chloroplasts isolated from Cyanidioschyzon merolae6,7 undergo GTP hydrolysis-dependent division. Using this assay, we show that Dnm2, rather than FtsZ, acts as the motor driving GTPase-powered contraction of the division ring, thereby physically dividing the chloroplast. We further demonstrate that the division ring is assembled through coiling of a glycosyltransferase-mediated filament and is crosslinked by dimerization of the Dnm2 GTPase domain. Following GTP hydrolysis-dependent force generation, Dnm2 retains its dimeric form in GDP-bound and nucleotide-free states, providing a locking step that suppresses back-slippage of the coiling ring during division. Thus, this mechanical design enables progressive, ratchet-like constriction of the division ring through coiling, overcoming the mechanical load posed by the chloroplast and generating the force required for fission, consistent with quantitative simulations. These findings suggest that a specialized division-ring mechanism, distinct from vesicle fission systems, evolved to mediate endosymbiont fission, allowing host control of endosymbiont proliferation and promoting faithful inheritance of the emerging organelle.

Root segmentation is a fundamental yet challenging task in image-based plant phenotyping. We present the first systematic comparison of Transformer and Convolutional Neural Network (ConvNet) architectures for root segmentation, evaluating 21 architectures across nine diverse datasets and comparing pre-trained models to training from scratch. Transformer-based models significantly outperform ConvNets for segmentation accuracy and root-diameter agreement. Pre-training significantly improves mean Dice from 0.623 to 0.666 (p = 3.3 x 10-10). We also find that Transformers benefit more from pre-training than ConvNets, with Dice improvements of +0.072 versus +0.022 (p = 3.7 x 10-4), supporting the hypothesis that fine-tuned Transformers transfer more effectively across large domain gaps. Among evaluated models, MobileSAM achieved the highest Dice score while maintaining computational efficiency. Dataset choice explained far more performance variance (70.9%) than model architecture (6.7%), suggesting that data curation matters more than model selection.

Halotropism--the directional growth of roots away from saline environments--requires coordinated integration of tropic cues. We show that halotropic bending in Arabidopsis thaliana roots is fine-tuned by a spatially confined, symmetric reactive oxygen species (ROS) domain generated by the NADPH oxidase RBOHC in elongation-zone epidermal cells. This domain, visualized by dihydrorhodamine-123 staining and confocal microscopy, emerges during the first hours of halostimulation and limits excessive curvature. Reducing ROS, either chemically with ascorbate or diphenyleneiodonium, or genetically in rbohC mutants, enhances halotropic bending, whereas miz2, defective in the ARF-GEF GNOM, exhibits negative halotropism due to an expanded and mislocalized ROS domain that disrupts spatial restriction. The miz2 rbohC double mutant shows a much weaker halotropic response than rbohC alone and similarly lacks the halotropic ROS signals in the elongation zone, indicating that GNOM acts upstream of RBOHC-mediated ROS production. Comparisons with hydrotropism--a moisture-seeking response also involving defined ROS distribution--suggest that GNOM-dependent regulation of RBOHC constitutes a shared module for adjusting root orientation to environmental gradients. Understanding these molecular mechanisms is essential for enhancing crop resilience to soil salinity, particularly in the context of increasing soil salinization driven by climate change.

Arising from Stander et al. Communications Biology https://doi.org/10.1038/s42003-023-05574-8 (2023) Rauvolfia tetraphylla belongs to the Apocynaceae family, one of the largest plant families distributed across Mexico, Tropical America and Southeast Asia. They are known for producing pharmacologically important monoterpene indole alkaloids (MIAs), such as ajmaline, reserpiline, yohimbine and heteroyohimbanes1. Due to their pharmaceutical properties, the biosynthetic modalities of several of these secondary metabolites have been explored2, however, the enzymatic pathways underlying yohimbine production have remained fairly understudied. Elucidating these pathways is therefore critical to enhance our understanding of yohimbane MIA metabolism and aid in their industrial production and drug discovery. This research gap was adequately addressed in a recent edition of Communication Biology, where Stander et al. [STAN23]3 presented a high-quality assembly for R. tetraphylla using a multi-platform high-coverage dataset. Notably, the yohimbane biosynthesis pathway was uncovered for the first time. Based on their metabolomics, proteomics and transcriptomic analysis followed by in vitro biochemical assays, two major findings emerged: (1) A medium chain dehydrogenase/reductase (MDR), yohimbane synthase (YOS) was found that produces a mixture of four diastereomers of yohimbanes (2) Three MDR transcripts (MDRT), MSTRG.5530, MSTRG.5531, and MSTRG.5534 were identified, which, in conjunction with geissoschizine synthase (GS, MSTRG.5528), produce a mixture of yohimbane isomers. However, the well-foundedness of the results is limited by a methodological oversight: the study does not take into account an independent whole-genome triplication event (WGT) in R. tetraphylla other than the one shared across Eudicotyledons, thus limiting the candidates identified in yohimbane biosynthesis and undermining the complexity of the biosynthetic modalities4,5.

Lignin biosynthesis in grasses exhibits unique metabolic flexibility, yet the precursor-specific routing of carbon into lignin polymers remains poorly resolved in planta. Here, we combine 13C-isotope labeling with solid-state NMR under sensitivity-enhancement by dynamic nuclear polarization (DNP), to directly track phenylalanine- and tyrosine-derived carbon incorporation into the lignin polymer in Brachypodium distachyon. Precursor-specific 13C labeling reveals that phenylalanine is the dominant contributor to canonical guaiacyl and syringyl lignins, whereas tyrosine preferentially enriches hydroxyphenyl lignin and hydroxycinnamates, including ferulates characteristic of grass cell walls. Two-dimensional 13C-13C correlation NMR resolves distinct lignin moieties arising from each precursor. Disruption of p-coumarate 3-hydroxylase (C3H) selectively impairs phenylalanine-derived lignification, while tyrosine-derived lignin remains comparatively unchanged, maintaining polymer assembly through alternative metabolic routes. These findings show precursor-dependent control of lignin composition and reveal tyrosine-mediated lignification as a compensatory pathway in grasses. This work also establishes precursor-resolved solid-state NMR and DNP as a powerful framework for dissecting lignin biosynthesis and metabolic plasticity in plant cell walls. SIGNIFICANCE STATEMENTLignin is a complex plant polymer that strengthens cell walls but also limits the efficiency of biomass processing for agriculture and bioenergy. Grasses possess a unique lignin biosynthetic flexibility that is not well understood. By combining stable isotope labeling with solid-state NMR spectroscopy, we directly traced how the aromatic amino acids, phenylalanine and tyrosine, contribute differently to lignin formation in intact grass cell walls. We show that phenylalanine primarily builds conventional lignin structures, whereas tyrosine supplies alternative phenolic components and maintains lignin synthesis even when a key biosynthetic enzyme is disrupted. This metabolic flexibility helps explain the unique structural aspects of grass cell walls and identifies precursor-level control as a promising strategy for engineering lignin composition to improve biomass utilization.

Plants acquire essential mineral nutrients from the soil, yet these elements must first traverse the extracellular matrix of the root before reaching the cell surface. How the physical properties of this extracellular compartment influence nutrient distribution and availability remains poorly understood. In plants, this extracellular matrix is formed by the cell wall, which carries a dynamically regulated negative charge that can change during development and in response to environmental cues. Here we demonstrate that cell wall charge functions as a tunable electrostatic gate that determines how iron is partitioned between retention and bioavailability. This decoupling between iron abundance and availability reveals a fundamental tradeoff imposed by extracellular electrostatics. A mechanistic diffusion-binding model shows that increasing wall charge inherently enhances iron sequestration while limiting its mobility at the cell surface. Genetic perturbation of pectin de-methylesterification validates this principle in vivo. Moreover, iron limitation itself triggers active remodeling of cell wall charge, dynamically shifting the balance toward increased iron accessibility. Together, these findings identify the plant cell wall as an active regulator of nutrient homeostasis rather than a passive barrier. By dynamically modulating extracellular electrostatics, roots control iron partitioning and bioavailability, uncovering a new physical layer of regulation in plant mineral nutrition. One-Sentence SummaryThe plant cell wall operates as a tunable electrostatic gate that buffers and releases iron through spatially and environmentally regulated charge dynamics.

Functional genetic redundancy (FGR) within gene families limits the discovery of gene function in plants because single-gene perturbations often fail to produce informative phenotypes. Artificial microRNAs (amiRNAs) provide a strategy to silence multiple related genes simultaneously. However, the existing amiRNA-based libraries used for genetic gene function discovery in plants do not account for the subcellular localization of gene products, which can lead to pleiotropic or difficult-to-interpret phenotypes. Plastids are essential plant cell organelles that integrate central metabolic and signaling processes, including photosynthesis, hormone biosynthesis, and environmental responses. Here we introduce pamiR, a plastid-targeted amiRNA library designed to enable organelle-specific gene function discovery in Arabidopsis thaliana. Using plastid proteomic datasets, we identified high-confidence plastid-localized proteins and designed amiRNAs to target their gene(s) (families) minimizing FGR. This amiRNA library was introduced in a vector with fluorescence-accumulating seed technology enabling rapid, herbicide-free selection and screening in the first generation. Validation by next-generation sequencing, confirmed high representation and uniform distribution of amiRNAs within pamiR. Proof-of-concept screens recovered mutants affecting known and additional candidate genes involved in photosynthesis and abscisic acid biosynthesis. Therefore, the pamiR library provides a fast platform for plastid-focused genetic screens that is compatible with existing mutant collections. One-sentence summaryThe plastid amiRNA (pamiR) library enables organelle-specific forward genetics without functional genetic redundancy.