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bioRxiv2021

INDEHISCENT regulates explosive seed dispersal

How traits evolve to produce novelty or stasis is an open question in biology. We investigate this question in Cardamine hirsuta, a relative of Arabidopsis thaliana that employs explosive fracture to disperse its seeds. This trait evolved through key morphomechanical innovations that distinguish the otherwise very similar, dehiscent fruit of these two species. Using CRISPR/Cas9, we show that dehiscence zone formation is absolutely required for explosive fracture in C. hirsuta, and is controlled by the bHLH transcription factor INDEHISCENT (IND). Using mutant screens, we identified the MADS-box transcription factor FRUITFULL (FUL) as a negative regulator of IND in C. hirsuta. Although FUL function is conserved in C. hirsuta, the consequences of IND mis-expression differ in ful mutants of C. hirsuta versus A. thaliana. In ful mutants of both species, valve tissue is replaced by dehiscence zone tissue, which comprises two distinct cell types: lignified layer and separation layer cells. While A. thaliana ful mutants develop stunted fruit with ectopic lignified layer cells, C. hirsuta ful mutants have elongated fruit with ectopic separation layer cells. We show that IND dose determines the proportion of these two cell types in ectopic dehiscence zones. We also show that the extent of ectopic lignification caused by IND mis-expression determines fruit length. Our findings indicate developmental system drift in the conserved gene network patterning dehiscent fruit in two related Brassicaceae species.

plant biology↗

bioRxiv2025

Genetic determinants of aerial root traits that support biological nitrogen fixation in maize

Modern agriculture depends on chemically synthesized nitrogen fertilizer, which ensures high yields but also can carry significant environmental and economic costs. Biological nitrogen fixation (BNF) already supplies nitrogen to legume crops and several avenues of research are underway to extend it to non-legume crops. In maize (Zea mays), aerial roots have been shown to contribute to BNF in some varieties, and both having many aerial roots and large aerial roots contributes to the fixation trait. However, much of the genetics controlling aerial root number and size is still unknown. Here we validate and quantify BNF in maize varieties from Southern Mexico under controlled conditions and evaluate a population of double haploids derived from the elite inbred PHZ51 crossed with these varieties. We find that most aerial root traits (root number, nodes with roots, root size) are reasonably heritable (h2 0.5-0.75) and generally uncorrelated with each other. QTL mapping identifies 5 QTL each affecting nodes with aerial roots and aerial root number per node; in both cases all but 1 QTL show an increase from the landrace allele. We also identify 11 QTL for aerial root diameter, with most positive QTL coming from PHZ51. Between the two populations, only a few QTL overlap, indicating a presumably high diversity of genes affecting aerial root morphology in landrace populations. Combining the best QTL into elite material may provide a path toward meaningful levels of BNF for maize, and additional work is needed to determine how viable this approach will be in field settings.

plant biology↗

bioRxiv2024

Phytochemical and Therapeutic Evaluation of Ficus nervosa Leaves: Impact on Oxidative Stress, Analgesic, Diarrhea, and Blood Clotting in a Mice Model

BackgroundFicus nervosa (FN) leaves have been traditionally used in ethnomedicine for the treatment of various ailments, this study aimed to investigate the cold methanol extract of FN for its antioxidant, analgesic, antidiarrheal, and thrombolytic properties, along with preliminary phytochemical screening to identify key bioactive compounds and acute toxicity test to assess the safety profile of the extract. MethodsIn this study, we conducted an initial investigation to identify the major phytochemical groups present in the leaves of Ficus nervosa. Using conventional phytochemical screening methods on cold methanol extracts, we consistently identified phenols and flavonoids as the predominant bioactive compounds. Following this phytochemical characterization, we assessed the biological activities of the extracts across multiple models. The antioxidant capacity and free radical-scavenging ability of the leaves extracts were evaluated using the DPPH (2,2-Diphenyl-1-picrylhydrazyl) assay. Analgesic efficacy was assessed through acetic acid-induced writhing and electrically induced heat nociception tests. Antidiarrheal effects were examined using castor oil-induced diarrhea and an enteropooling assay in mice. To determine thrombolytic activity, we conducted a human blood clot lysis assay, while preclinical toxicity testing was conducted to evaluate the safety profile of the extracts. Statistical analysis was performed using one-way ANOVA with Duncans Multiple Range Test (DMRT) to validate the significance of the results. ResultsThe study of Antioxidant Potential, the ESFs (ethyl soluble fraction) antioxidant capacity, measured at 409.91 {+/-} 0.59 mg of ascorbic acid equivalent (AAE)/g in the phosphomolybdenum assay, signifies a strong free radical scavenging activity. This level of antioxidant action, alongside a potent free radical neutralizing concentration (IC of 9.99 {+/-} 0.41 g/mL), suggests that the ESF could effectively counteract oxidative stress. Analgesic activity at 500 mg/kg, FN extract produced a 68.05% suppression of writhing in mice, which was statistically significant (p<0.001) compared to standard morphines (74.05%) inhibition. The extract also significantly prolonged reaction latency to thermal-induced pain in hotplate model. The antidiarrheal activity showed 52.09% and 59.0% suppression of defecation, compared to standard loperamides 73.1%, with statistically significant p-values (p<0.04 and p<0.01). These effects, combined with the notable decrease in intestinal fluid accumulation and defecation frequency in enteropooling assays, indicate that FN extract might interact with intestinal motility or fluid secretion pathways, providing antidiarrheal benefits. Additionally, the chloroform-soluble fraction of FN exhibited moderate thrombolytic activity, achieving 39.79% clot lysis compared to streptokinase (69.52%). Although this thrombolytic potential is moderate, it indicates that the fraction contains compounds with some fibrinolytic properties, which could be useful as adjunctive agents in preventing thrombosis. ConclusionOverall, the cold methanol extract of FN leaves demonstrates the therapeutic potential in preclinical settings. Future research is warranted to isolate the specific bioactive compounds and explain their mechanisms of action to further support the development of new treatments and contributing to modern medicinal practices based on this plant leaves.

plant biology↗

bioRxiv2025

Spatial analysis of cell patterning to aid genetic and phenotypic understanding of grass stomatal density: a case study in maize

Biological processes involve complex hierarchies where composite traits result from multiple component traits. However, holistically understanding of how sets of component traits interact to underpin genotype-to-phenotype relationships is generally lacking. Stomatal density (SD) is a tractable model system for exploring how high-throughput phenotyping (HTP) data could be exploited by a new spatial analysis approach to better understand a developmentally and functionally important trait. SD is a composite trait, resulting from various components related to cell identity and size, which are themselves governed by a series of spatio-developmental processes. Data from 180 recombinant inbred lines of maize (Zea mays (L.)) were analyzed by a new Stomatal Patterning Phenotype (SPP) to: (1) describe the average spatial probability distribution of the nearest neighboring stomata; (2) derive a core set of component traits related to cell size, cell packing and positional probabilities; (3) build a structural equation model of component traits underlying SD; and (4) identify stomatal patterning quantitative trait loci (QTL). The core set of SPP-derived traits explained 74% of the variation in SD. Analyzing SPP component traits allowed some loci previously identified as generic SD QTL to be recognized as specific to lateral versus longitudinal elements of stomatal patterning. Therefore, this study highlights how novel insights can be gained by decomposing a composite trait (e.g. SD) into a set of component traits that were present in HTP data but not previously exploited.

plant biology↗

bioRxiv2020

A cost-effective maize ear phenotyping platform enables rapid categorization and quantification of kernels

High-throughput phenotyping systems are powerful, dramatically changing our ability to document, measure, and detect biological phenomena. Here, we describe a cost-effective combination of a custom-built imaging platform and deep-learning-based computer vision pipeline. A minimal version of the maize ear scanner was built with low-cost and readily available parts. The scanner rotates a maize ear while a cellphone or digital camera captures a video of the surface of the ear. Videos are then digitally flattened into two-dimensional ear projections. Segregating GFP and anthocyanin kernel phenotype are clearly distinguishable in ear projections, and can be manually annotated using image analysis software. Increased throughput was attained by designing and implementing an automated kernel counting system using transfer learning and a deep learning object detection model. The computer vision model was able to rapidly assess over 390,000 kernels, identifying male-specific transmission defects across a wide range of GFP-marked mutant alleles. This includes a previously undescribed defect putatively associated with mutation of Zm00001d002824, a gene predicted to encode a vacuolar processing enzyme (VPE). We show that by using this system, the quantification of transmission data and other ear phenotypes can be accelerated and scaled to generate large datasets for robust analyses. One sentence summaryA maize ear phenotyping system built from commonly available parts creates images of the surface of ears and identifies kernel phenotypes with a deep-learning-based computer vision pipeline.

plant biology↗

bioRxiv2021

BoolSim, a Graphical Interface for Open Access Boolean Network Simulations and Use in Guard Cell CO2 Signaling

Signaling networks are at the heart of almost all biological processes. Most of these networks contain a large number of components and often the connections between these components are either not known, or the rate equations that govern the dynamics of soluble signaling components are not quantified. This uncertainty in network topology and parameters can make it challenging to formulate detailed mathematical models. Boolean networks, in which all components are either on or off, have emerged as viable alternatives to more detailed mathematical models but can be difficult to implement. Therefore, open source format of such models for community use is desirable. Here we present BoolSim, a freely available graphical user interface (GUI) that allows users to easily construct and analyze Boolean networks. BoolSim can be applied to any Boolean network. We demonstrate BoolSims application using a previously published network for abscisic acid-driven stomatal closure in Arabidopsis. We also show how BoolSim can be used to generate testable predictions by extending the network to include CO2 regulation of stomatal movements. Predictions of the model were experimentally tested and the model was iteratively modified based on experiments showing that ABA closes stomata even at near zero CO2 concentrations (1.5 ppm CO2). One Sentence SummaryThis study presents an open-source, graphical interface for the simulation of Boolean networks and applies it to an abscisic acid signaling network in guard cells, extended to include input from CO2.

plant biology↗

bioRxiv2024

Grape ripening speed slowed down using natural variation

Understanding ripening patterns and governing ripening speed are central aspects of grapevine (Vitis vinifera) berry biology owing to the importance of grape ripeness in winemaking. Despite this, the genetic control of ripening is largely unknown. Here, we report a major quantitative trait locus that controls ripening speed, expressed as speed of sugar accumulation. A haplotype originating from the species Vitis riparia halves maximum speed regardless of crop levels and berry sizes. The sequence of events that are normally completed at the onset of ripening in a two-week period known in viticulture as veraison are taking place at a slower speed, thereby attaining ripeness under milder weather conditions in late summer. V. vinifera cultivars show limited phenotypic variation for ripening speed and no selective sweep in the causal genomic region that could derive from domestication or improvement. Closely related species make up for the lack of standing variation, supplying major effect alleles for adapting grape cultivars to climate change. HIGHLIGHT / SIGNIFICANCE STATEMENTReducing the speed of fruit ripening genetically is a means for adapting the grape berry developmental program to the changing needs of the wine industry and in response to global warming. We identified a haplotype in a wild grape species that slows down the speed of ripening in progenies of Vitis vinifera by limiting the speed of sugar accumulation throughout the duration of ripening, a condition of great importance for winemakers to harvest their grapes at the desired level of technological ripeness.

plant biology↗

bioRxiv2025

Mechanistic insight into the oligomerization of Arabidopsis CRY1 and its inhibition by BIC1

Cryptochromes (CRY) convert blue light signals into biological responses, however, the molecular processes underlying their activation are not fully understood. In this study, we uncover the molecular mechanism underlying the blue-light activation of Arabidopsis CRY1 using time-resolved native mass spectrometry combined with kinetic modelling - an approach that allows us to monitor light-driven complex formation with temporal and molecular resolution. We show that CRY1 activation follows a defined, reversible assembly pathway in which monomers rapidly form dimers that then assemble into tetramers. A quantitative two-step model captures the dynamic interplay between light-driven assembly and thermal disassembly. Strikingly, ATP accelerates tetramer formation and stabilises oligomers by tuning the underlying photochemistry of the FAD chromophore. In contrast, the Blue-light Inhibitor of Cryptochromes 1 (BIC1) acts as a potent antagonist. We found that BIC1 binds to CRY1 even in the dark, with a significant increase in binding strength under blue-light conditions. BIC1 not only blocks CRY1 oligomerisation, but also actively dismantles pre-assembled tetramers. This disassembly process is light-independent and occurs regardless of CRY1s redox state. Together, these findings reveal a finely balanced regulatory system in which ATP and BIC1 act as opposing regulators to control CRY1 activation. This work provides a kinetic and mechanistic framework for reversible cryptochrome signalling and highlights how blue light responses can be precisely modulated at the molecular level.

plant biology↗

bioRxiv2022

Elucidating the patterns of pleiotropy and its biological relevance in maize

Pleiotropy - when a single gene controls two or more seemingly unrelated traits - has been shown to impact genes with effects on flowering time, leaf architecture, and inflorescence morphology in maize. However, the genome-wide impact of true biological pleiotropy across all maize phenotypes is largely unknown. Here we investigate the extent to which biological pleiotropy impacts phenotypes within maize through GWAS summary statistics reanalyzed from previously published metabolite, field, and expression phenotypes across the Nested Association Mapping population and Goodman Association Panel. Through phenotypic saturation of 120,597 traits, we obtain over 480 million significant quantitative trait nucleotides. We estimate that only 1.56-32.3% of intervals show some degree of pleiotropy. We then assessed the relationship between pleiotropy and various biological features such as gene expression, chromatin accessibility, sequence conservation, and enrichment for gene ontology terms. We find very little relationship between pleiotropy and these variables when compared to permuted pleiotropy. We hypothesize that biological pleiotropy of common alleles is not widespread in maize and is highly impacted by nuisance terms such as population structure and linkage disequilibrium. Natural selection on large standing natural variation in maize populations may target wide- and large-effect variants, leaving the prevalence of detectable pleiotropy relatively low. Author SummaryThe genetic basis of complex traits has been thought to exhibit pleiotropy, which is the notion that a single locus can control two or more unrelated traits. Widespread reports in the human disease literature show genomic signatures of pleiotropic loci across many traits. However, little is known about the prevalence and behavior of pleiotropy in maize across a large number of phenotypes. Using association mapping of common alleles in over one hundred thousand traits, we determine how pleiotropic each region was and use these quantitative scores to functionally characterize each region of the genome. Our results show little evidence that pleiotropy is a common phenomenon in maize. We observed that maize does not exhibit the same pleiotropic characteristics as human diseases in terms of prevalence, gene expression, chromatin accessibility, or sequence conservation. Rather than pervasive pleiotropy, we hypothesize that strong selection on large and wide effect loci and the need for trait independence at the gene level keep the prevalence of pleiotropy low, thus, allowing for the adaptation of maize varieties to novel environments and conditions.

plant biology↗

bioRxiv2026

Gene-metabolite networks reveal physiological trade-offs but not transcriptional co-regulation between carotenoids and dry matter in cassava (Manihot esculenta) roots

Provitamin A biofortification of cassava (Manihot esculenta) is constrained by a negative genetic correlation between total carotenoid (TC) and dry matter (DM) contents, but its underlying biological mechanism remains unclear. We examined this relationship in 24 African, Latin American, and hybrid cassava genotypes by measuring gene expression and metabolites in inner and outer storage root layers that varied spatially in carotenoid accumulation, enabling comparisons across and within genotypes. TC, and particularly upstream cis-carotenes, were negatively associated with DM after accounting for genotype, and this relationship strengthened across developmental timepoints, consistent with a physiological component of the trade-off. Gene-metabolite co-expression networks constructed from genetic, non-genetic, and overall phenotypic trait components showed no significant topological overlap between carotenoid- and starch-related gene sets, indicating that the TC/DM relationship is not driven by transcriptional co-regulation of the main pathways, although individual associations between carotenoids and carbohydrate-related genes suggested possible indirect metabolic links. Instead, TC accumulation was most strongly associated with regulatory, stress-, redox-, and plastid-related genes, enriched for heme-binding and oxidoreductase functions. These findings suggest that the TC/DM trade-off is not a fixed constraint, and identify candidate regulators of carotenoid content for future functional validation to support cassava provitamin A biofortification. HighlightGenetic, spatial, and developmental variation in cassava storage roots implicate cis-carotenes, but not direct transcriptional regulation of the carotenoid and starch pathways, in the trade-off between carotenoids and dry matter.

plant biology↗

bioRxiv2026

Multi-Omics Integration Predicts Cell-Specific Gene Regulatory Response and Rhizosphere Dynamics in Maize Root Fertilizer Treatment

Improving nitrogen use efficiency in maize (Zea mays) requires understanding how distinct root cell types and regulatory networks process fertilizer inputs. Given the current limited understanding of fertilizer-induced, cell-type-resolved maize roots and regulatory networks, computational biology frameworks are needed to model and predict how nutrient inputs are translated into transcriptional responses. Here, we integrated fertilizer-induced maize root bulk RNA-seq with reference atlases of single-cell RNA-seq and scATAC-seq to construct and predict a cell-specific regulome of the maize root under inorganic and mixed amendments. We demonstrate that inorganic fertilization induced stress associated and management pathways. Regulome analysis identified transcription factors (TF) from the AP2/ERF, NAC, HSF, and WRKY superfamilies that were preferentially active across root tissues. Deconvolution of the regulome onto single-cell atlases predicted core TF activity to the vascular cylinder and pith across both regimes, while mature cortex regulatory programs diverged. Construction of a gene regulatory network revealed that shared TF:target edges maintained the same regulatory orientation across fertilizer regimes. However, a small number of stress related TFs, including WRKY24, DREB1A, and NAC61, underwent a directional change between fertilization treatments. In silico knockout analysis predicted the activation targets for six of the seven regulators in their resident vascular/pith tissues, indicating the network behaves as a coherent, perturbable system. Additionally, soil metagenomic analysis showed that host soil microbial functions overlap with differentially expressed genes (DEGs) in shared functional categories, linking host regulome dynamics to rhizosphere processes. These findings and predictions suggest that the maize root regulome is spatially organized and dynamically reprogrammed by master regulators, predicting high-priority candidate nodes for engineering improved nutrient use efficiency.

plant biology↗

bioRxiv2016

Reproductive biology of three Ziziphus species (Rhamnaceae)

The reproductive biology of three Ziziphus species: Z. jujube, Z. mauritiana and Z. spina-christi, with respect to seasonal growth stages, floral biology, pollen performance, fruit set and seed viability were compared. To elucidate the reasons for variations in reproductive success we also estimated the ploidy levels among the genotypes of these tree species. Four genotypes of Z. mauritiana and Z. jujube each, and two natural selections of Z. spina-christi were grown in the Negev highlands at the Sde Boqer experimental plot. Field observations were conducted to determine seasonal growth stages and floral biology; laboratory studies included florescent microscopy, differential interference contrast microscopy, in vitro pollen germination and histological techniques. Seasonal growth stages were similar in Z. spina-christi and Z. mauritiana but distinct in Z. jujube, which was the earliest species. Flower opening stages were similar in all species and genotypes. Anthesis timing was genotype specific and did not depend on species. Anthesis types were subdivided into early and late morning as well as afternoon. We found a wide range of values for the number of flowers per inflorescence between species and genotypes, but all genotypes set one fruit per inflorescence with the exception of Z. jujube genotype Tamar, which set two fruit per inflorescence. Significant variation in fruit weight was observed between species as well as genotypes. Fruit of Z. mauritiana and Z jujube weighed between 9 to 37 g while fruit weight of Z. spina-christi was between 0.5 to 0.7 g. Flowers of all species and genotypes had two ovules but viable seed set per fruit differed dramatically between species under open pollination. Viable seed set was about 18% for Z. mauritiana and Z jujube, while Z. spina-christi, had the highest viable seed set at about 84%. Histological analysis revealed post fertilization embryo abortion which may be responsible for the relatively low reproductive success in Z. mauritiana and Z. jujube. The results obtained in this study provide an important basis for selecting elite complementary genotypes toward the optimization of Ziziphus fruit production in semi-arid regions.

Plant Biology↗

bioRxiv2021

Measuring hidden phenotype: Quantifying the shape of barley seeds using the Euler Characteristic Transform

Shape plays a fundamental role in biology. Traditional phenotypic analysis methods measure some features but fail to measure the information embedded in shape comprehensively. To extract, compare, and analyze this information embedded in a robust and concise way, we turn to Topological Data Analysis (TDA), specifically the Euler Characteristic Transform. TDA measures shape comprehensively using mathematical representations based on algebraic topology features. To study its use, we compute both traditional and topological shape descriptors to quantify the morphology of 3121 barley seeds scanned with X-ray Computed Tomography (CT) technology at 127 micron resolution. The Euler Characteristic Transform measures shape by analyzing topological features of an object at thresholds across a number of directional axes. A Kruskal-Wallis analysis of the information encoded by the topological signature reveals that the Euler Characteristic Transform picks up successfully the shape of the crease and bottom of the seeds. Moreover, while traditional shape descriptors can cluster the seeds based on their accession, topological shape descriptors can cluster them further based on their panicle. We then successfully train a support vector machine (SVM) to classify 28 different accessions of barley based exclusively on the shape of their grains. We observe that combining both traditional and topological descriptors classifies barley seeds better than using just traditional descriptors alone. This improvement suggests that TDA is thus a powerful complement to traditional morphometrics to comprehensively describe a multitude of "hidden" shape nuances which are otherwise not detected.

plant biology↗

bioRxiv2026

Comparative genomics reveals potential mechanisms of invasion in Phragmites australis (common reed)

Biological invasions are transforming ecosystems worldwide, yet the genomic bases enabling certain species to dominate new environments remain poorly understood. Phragmites australis, a widespread wetland grass with invasive and native subspecies co-occurring in North America, provides a powerful system to investigate genomic mechanisms of invasiveness. We generated independent chromosome-scale genome assemblies for invasive P. australis ssp. australis and co-occurring native ssp. americanus and used comparative genomic and transcriptomic analyses to identify lineage-specific innovations associated with invasive success. The invasive subspecies exhibits genomic novelties through functionally-biased single-copy orthologs, intronless genes, and subgenome expression asymmetry, along with a stress-ready basal transcriptome relative to the native subspecies. Following the removal of aboveground shoots ("cutback"), which measures the ability to recover from damage, the invasive subspecies undergoes stronger transcriptional reprogramming, increased shoot production, and higher biomass accumulation compared to the native. It also displays expansion of gene families and coordinately expressed gene modules that support resource mobilization, growth responses to light, and stress tolerance. Beyond Phragmites, comparative analyses across multiple grass genomes, including eight invasive species with related non-invasive species, revealed repeated expansion of gene families associated with abiotic stress tolerance and developmental regulation, suggesting convergent adaptive strategies in the grass family for invasive success. Together, these results demonstrate genomic architecture linked to invasion success and highlight potential targets for managing invasive grasses.

Plant Biology↗

bioRxiv2025

Production Status, Breeding Priorities and Genetic Resources of Cowpea in Post-Civil War Somalia

Over the past three decades, Somalia has faced recurrent civil conflicts and prolonged droughts, leading to widespread displacement of farming communities. These disruptions have weakened traditional seed systems and potentially damaged local crop diversity. In this context, to strengthen the countrys food security and climate change adaptation of smallholder farmers, there is an urgent need to revive agricultural research and breeding programs for the main food crops grown in the country. Cowpea (Vigna unguiculata (L.) Walp.) is the most important food crop after maize and sorghum in Somalia, providing inexpensive protein for humans and biologically fixed nitrogen for low-input crop production systems in the country. This study assessed the current status of cowpea production, identified breeding priorities across three agroecological zones in southern Somalia, and established a core germplasm collection for future breeding programs. Farmer surveys were conducted in Baidoa, Afgoie, and Jowhar, involving interviews with 150 farmers using a semi-structured questionnaire. The results showed that over 60% of farmers cultivate cowpea for both household consumption and market purposes. While the crop is predominantly grown as an intercrop, nearly 45% of farmers in Afgoie grow it as a sole crop. Marked regional differences were observed in varietal preferences, with farmers in Afgoie favouring erect, uniformly maturing cultivars with a dark-red seed colour, whereas other regions demonstrated more diverse preferences. Production constraints also varied by location, with drought identified as the primary limitation in Baidoa, and pests and diseases as the major challenges in Afgoie and Jowhar. Accordingly, farmers ranked early maturity as the top breeding priority in Baidoa, whereas disease and pest resistance were the primary targets in Afgoie. Furthermore, we established a germplasm collection representing Somalias cowpea diversity from both the pre- and post-war periods. The collection also includes a subset of global cowpea diversity and accessions with known desirable agronomic traits. These genetic resources, together with farmer survey data, provide a foundation for targeted breeding efforts aimed at improving cowpea productivity and enhancing food security in Somalia in the future.

plant biology↗

bioRxiv2024

A Genetic Toggle Switch in Plants

In synthetic biology, genetic components are assembled to make transcriptional units, and transcriptional units are assembled into circuits to perform specific and predictable functions of a genetic device. Genetic devices have been described in bacteria, mammalian cell cultures and small organoids, yet development of programmable genetic circuits for devices in plants has lagged. Programmable genetic devices require defining the components quantitative functions. Because plants have long life spans, studies often use transient analysis to define quantitative functions while verification in stably engineered plants is often neglected and largely unknown. This raises a question if unique attributes of plants such as environmental sensitivity, developmental plasticity, or alternation of generations, adversely impacts predictability of plant genetic circuits and devices. Alternatively, it is also possible that genetic elements to produce predictable genetic devices for plants require rigorous characterization with detailed mathematical modeling. Here we use plant genetic elements with quantitatively characterized transfer functions and developed in silico models to guide their assembly into a genetic device: a toggle switch or a mutually inhibitory gene-regulatory device. Our approach allows computational selection of plant genetic components and iterative refinement of the circuit if the desired genetic functions are not initially achieved. We show that our computationally selected genetic circuit functions as predicted in stably engineered plants including through tissue and organ differentiation. Developing abilities to produce predictable and programmable plant genetic devices opens the prospect of predictably engineering plants unique abilities in sustainable human and environmental systems.

synthetic biology↗

bioRxiv2026

A Comprehensive Epidermal Map from a Poplar Single-Cell Shoot Atlas Reveals New Trichome-Specific Genes

Poplar (Populus spp.) is a model system for tree biology. Specifically, P. tremula x P. alba INRA 717-1B4 (hereafter "poplar 717") has become an important platform for functional genomics and synthetic biology due to its rapid growth and ease of transgenesis. Here, we present a single-cell RNA-seq atlas of the poplar 717 shoot, including apical meristem, primary and secondary stems, and three stages of leaf development. Analysis of ca. 159,000 cells resolved 40 transcriptionally distinct clusters representing 7 major cell types, providing a high-resolution view of shoot development and tissue organization. We focused on the epidermis which constituted >15% of cells in the shoot atlas for in-depth characterization of epidermal heterogeneity. By integrating known marker genes with transcriptomic signatures consistent with established poplar leaf phytochemistry, we annotated epidermal cell subclusters corresponding to developmental stages, spatial location, and specialized cell types, including a distinct population of non-glandular trichomes. Coupling the single-cell RNA-seq atlas with bulk transcriptome data from glabrous mutants enabled the identification of novel trichome markers. Experimental validation of a representative trichome-specific promoter established a tool with potential to support cell type-targeted-metabolic engineering. We provide the poplar 717 atlas to the community through the BioPoplar Atlas Viewer (http://bio-poplar-atlas.com), providing a platform to explore the poplar transcriptome at single-cell resolution and a foundation for data-driven cell type-aware genetic engineering strategies in poplar.

plant biology↗

bioRxiv2025

Spatio-temporal plasticity of root exudation in three temperate tree species: effects of season, site and soil characteristics

Root exudation provides a constant carbon input to the rhizosphere and is therefore a very important factor in shaping this hotspot of biological activity. Nonetheless, root exudation data and its spatio-temporal plasticity is scarce. This study provides insights into compound-specific root exudation in three temperate tree species in two seasons (late spring and late summer) and two soil compartments (forest floor and the top mineral soil), including the effect of soil chemistry. At four sites with differing mean annual temperature and soil phosphorus level, root exudates were sampled using an in-situ cuvette-based system and analysed by gas chromatography-mass spectrometry. We found seasonally and spatially varying site- and species-specific exudation patterns. While the seasonal pattern was similar among species and sites, with higher exudation rates in late spring, soil compartment-specific exudation depended on species and site. Acer pseudoplatanus tended to exude more into the mineral soil at warmer sites, while Picea abies exuded more in the mineral soil at all sites. Exudation by Fagus sylvatica was independent from the soil compartment. Significant correlation between exchangeable soil cations and specific compounds exuded by F. sylvatica and P. abies were found. Exudation of specific compounds in F. sylvatica increased with the concentration of exchangeable Mg, Al and Fe, whereas exudation rates in P. abies decreased with most base cations concentration, while sugar exudation increased with the exchangeable non-base cations Al and Fe. These results demonstrate that root exudation is dynamically adjusted to the species-specific nutritional needs governed by site, season and soil characteristics.

plant biology↗