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bioRxiv2018

Dynamic regulation of HYL1 provides new insights into its multifaceted role in Arabidopsis

MicroRNAs (miRNAs) are 21 to 24 nucleotide non-coding RNAs that regulate gene expression. Biogenesis of miRNAs is fine-tuned by specialized microprocessor complex, the regulation of which is being continuously understood. Recruitment of HYL1 to the microprocessor complex is crucial for accurate primary-miRNA (pri-miRNA) processing and accumulation of mature miRNA in Arabidopsis thaliana. HYL1 is a double-stranded RNA binding protein also termed as DRB1, has two double-stranded RNA binding domain at N-terminal and a highly disordered C-terminal region. Also, the biological activity of HYL1 is dynamically regulated through transition from hyperphosphorylation to hypophosphorylation state. HYL1 is known to be phosphorylated by a MAP kinase MPK3 and SnRK2. However, the precise role of its phosphorylation are still unknown. Recently, the stability of HYL1 protein has been shown to be regulated by an unknown protease X. However, the identity of the protease and its molecular mechanisms are poorly understood. Here, we describe, three functionally important facets of HYL1, which provide a better picture of its association with molecular processes. First, we identified a conserved MPK3 phosphorylation site on HYL1 and its possible role in the miRNA biogenesis. Secondly, the C-terminal region of HYL1 displays tendencies to bind dsDNA. Lastly, the role of C-terminal region of HYL1 in the regulation of its protein stability and the regulation of miRNA biogenesis is documented. We show the unexplored role of C-terminal and hypothesize the novel functions of HYL1 in addition to miRNA biogenesis. We anticipate that the data presented in this study, will open a new dimension of understanding the role of double stranded RNA binding proteins in diverse biological processes of plants and animal.

plant biology↗

bioRxiv2019

Generation of strong casein kinase 1 inhibitor of Arabidopsis thaliana

Casein kinase 1 (CK1) is an evolutionarily conserved protein kinase among eukaryotes. Studies on yeast, fungi, and animals have revealed that CK1 plays roles in divergent biological processes. By contrast, the collective knowledge regarding the biological roles of plant CK1 lags was behind those of animal CK1. One of reasons for this is that plants have more multiple genes encoding CK1 than animals. To accelerate the research for plant CK1, a strong CK1 inhibitor that efficiently inhibits multiple members of CK1 proteins in vivo (in planta) is required. Here, we report a novel strong CK1 inhibitor of Arabidopsis (AMI-331). Using a circadian period-lengthening activity as estimation of the CK1 inhibitor effect in vivo, we performed a structure-activity relationship (SAR) study of PHA767491 (1,5,6,7-tetrahydro-2-(4-pyridinyl)-4H-pyrrolo[3,2-c]pyridin-4-one hydrochloride), a potent CK1 inhibitor of Arabidopsis, and found that PHA767491 analogues bearing a propargyl group at the pyrrole nitrogen atom (AMI-212) or a bromine atom at the pyrrole C3 position (AMI-23) enhance the period-lengthening activity. The period lengthening activity of a hybrid molecule of AMI-212 and AMI-23 (AMI-331) is about 100-fold stronger than that of PHA767491. An in vitro assay indicated a strong inhibitory activity of CK1 kinase by AMI-331. Also, affinity proteomics using an AMI-331 probe showed that targets of AMI-331 are mostly CK1 proteins. As such, AMI-331 is a strong potent CK1 inhibitor that shows promise in the research of CK1 in plants.

plant biology↗

bioRxiv2026

Rice Jumonji706 confers the photoperiod sensitivity in rice by distinct regulation of short-day and long-day flowering time regulatory pathways.

Photoperiod sensitivity (PS) is a key biological response in plants as they adapt to specific environments. Rice (Oryza sativa L.) exhibits a clear PS, as it implements critical phase transition decisions based on PS signals. In this study, we identified a novel PS gene, JMJ706, that is expected to deliver photoperiod-related signals to the flowering-time regulatory network in a day-length-dependent manner. The JMJ706 mutants exhibit early flowering under LD and later flowering under SD compared to WT plants. The gene encodes an H3K9me2 demethylase, and under long-day (LD) conditions, its demethylase activity facilitates the expression of Grain number, Plant height, and Heading-date7 (Ghd7). Since Ghd7 is a floral repressor in LD, it promotes the vegetative phase by delaying flowering. Under short-day conditions (SD), H3K9me2 demethylase activity facilitates Early heading-date 1 (Ehd1) expression, and it acts as a floral accelerator by inducing Heading date 3 (Hd3a) and RICE FLOWERING LOCUS T 1 (RFT1). Furthermore, we propose that the daylength-dependent promotion of target genes (Ghd7 and Ehd1) occurs through demethylation of specific promoter regions at a crucial time window. In addition, JMJ706 may play an important role in regulating plant architecture, including plant height. The natural variation in JMJ706 alleles shows high frequencies across major rice subpopulations, suggesting that JMJ706 could play an important role in the geographical distribution and adaptation of rice cultivars. Our results may add a new layer to the rice flowering-time regulatory pathway, supporting regional adaptation and potential for future breeding.

plant biology↗

bioRxiv2021

ACORBA: Automated workflow to measure Arabidopsis thaliana root tip angle dynamic

Plants respond to the surrounding environment in countless ways. One of these responses is their ability to sense and orient their root growth toward the gravity vector. Root gravitropism is studied in many laboratories as a hallmark of auxin-related phenotypes. However, manual analysis of images and microscopy data is known to be subjected to human bias. This is particularly the case for manual measurements of root bending as the selection lines to calculate the angle are set subjectively. Therefore, it is essential to develop and use automated or semi-automated image analysis to produce reproducible and unbiased data. Moreover, the increasing usage of vertical-stage microscopy in plant root biology yields gravitropic experiments with an unprecedented spatiotemporal resolution. To this day, there is no available solution to measure root bending angle over time for vertical-stage microscopy. To address these problems, we developed ACORBA (Automatic Calculation Of Root Bending Angles), a fully automated software to measure root bending angle over time from vertical-stage microscope and flatbed scanner images. Moreover, the software can be used semi-automated for camera, mobile phone or stereomicroscope images. ACORBA represents a flexible approach based on both traditional image processing and deep machine learning segmentation to measure root angle progression over time. By its automated nature, the workflow is limiting human interactions and has high reproducibility. ACORBA will support the plant biologist community by reducing time and labor and by producing quality results from various kinds of inputs. Significance statementACORBA is implementing an automated and semi-automated workflow to quantify root bending and waving angles from images acquired with a microscope, a scanner, a stereomicroscope or a camera. It will support the plant biology community by reducing time and labor and by producing trustworthy and reproducible quantitative data.

plant biology↗

bioRxiv2021

Mobius Assembly for Plant Systems highlights promoter-terminator interaction in gene regulation

Plant synthetic biology is a fast-evolving field that employs engineering principles to empower research and bioproduction in plant systems. Nevertheless, in the whole synthetic biology landscape, plant systems lag compared to microbial and mammalian systems. When it comes to multigene delivery to plants, the predictability of the outcome is decreased since it depends on three different chassis: E. coli, Agrobacterium, and the plant species. Here we aimed to develop standardised and streamlined tools for genetic engineering in plant synthetic biology. We have devised Mobius Assembly for Plant Systems (MAPS), a user-friendly Golden Gate Assembly system for fast and easy generation of complex DNA constructs. MAPS is based on a new group of small plant binary vectors (pMAPs) that contains an origin of replication from a cryptic plasmid of Paracoccus pantotrophus. The functionality of the pMAP vectors was confirmed by transforming the MM1 cell culture, demonstrating for the first time that plant transformation is dependent on the Agrobacterium strains and plasmids; plasmid stability was highly dependent on the plasmid and bacterial strain. We made a library of new short promoters and terminators and characterised them using a high-throughput protoplast expression assay. Our results underscored the strong influence of terminators in gene expression, and they altered the strength of promoters in some combinations and indicated the presence of synergistic interactions between promoters and terminators. Overall this work will further facilitate plant synthetic biology and contribute to improving its predictability, which is challenged by combinatorial interactions among the genetic parts, vectors, and chassis.

synthetic biology↗

bioRxiv2025

Remote sensing of endogenous pigmentation by inducible synthetic circuits in grasses

Plant synthetic biology holds great promise for engineering plants to meet future demands. Genetic circuits are being designed, built, and tested in plants to demonstrate proof of concept. However, developing these components in monocots, which the world relies on for grain, lags behind dicot models, such as Arabidopsis thaliana and Nicotiana benthamiana. Here, we show the successful adaptation of a ligand-inducible sensor to activate an endogenous anthocyanin pathway in the C4 monocot model Setaria viridis. We identify two transcription factors sufficient to induce endogenous anthocyanin production in S. viridis protoplasts and whole plants in a constitutive or ligand-inducible manner. We also test multiple ligands to overcome physical barriers to ligand uptake, identifying triamcinolone acetonide (TA) as a highly potent inducer of this system. Using hyperspectral imaging and a discriminative target characterization method in a near-remote configuration, we can non-destructively detect anthocyanin production in leaves in response to ligands. This work demonstrates the use of inducible expression systems in monocots to manipulate endogenous pathways, stimulating plants to overproduce secondary metabolites with value to human health. Applying inducible pigmentation coupled with sensitive detection algorithms could enable crop plants to report on the status of field contamination or detect undesirable chemicals impacting agriculture, ushering in an era of agriculture-based sensor systems. SummaryO_ST_ABSSynbio tools for C4 grass modelC_ST_ABS[bullet] Advantage of synthetic switches as tools for biopharming and functional genomics [bullet]Our workflow to optimize the gene circuits from a transient system to a stable transgenic [bullet]Testing and taming golden gate elements in monocot system, S. viridis

plant biology↗

bioRxiv2025

Evaluation of computational tools for predicting CRISPR gRNA on-target efficiency in plants.

CRISPR technologies has become an integral part of plant biotechnology, synthetic biology and basic plant research, routinely used by researchers for targeted genome modifications. CRISPR guide RNAs (gRNAs) undermines the highly programmable nature of CRISPR, enabling site-specific genome editing. However, different gRNA targets showed highly variable on-target effectiveness and poor gRNA design could amount to wasting valuable scientific resources. There has been broad development of computational and web-based tools for gRNA efficiency predictions but their performances in plant genome editing remains controversial or untested. Hence, in this study, we systematically evaluated over 20 accessible, web-based in silico gRNA on-target efficiency prediction tools using an experimental plant genome editing dataset. Excitingly, we identified multiple tools, mostly developed using machine learning, that were highly predictive of gRNA on-target genome editing efficiency in planta. The prediction scores assigned to gRNAs in the dataset by these tools were significantly correlated with the frequency of CRISPR-mediated InDels in plants. Furthermore, we evaluated efficiency prediction scores available on popular platforms such as CRISPOR and CRISPR-P which contain large numbers of non-model plant genomes. Our analysis showed that some prediction scores on CRISPOR performed quite well which allows efficient integration of on-target and off-target predictions. Overall, we believe that our study provided insights on improving gRNA design during conventional plant genome editing workflows and should also help unfamiliar researchers interested in CRISPR/SpCas9 genome editing.

plant biology↗

bioRxiv2026

Extracellular vesicles associate with infectious geminiviral particles in the apoplast of infected plants.

Plant viruses have evolved diverse strategies to facilitate their movement and survival within the host. Among them, geminiviruses co-opt host cellular machinery to replicate and disseminate. Traditionally, viral propagation has been associated with intercellular symplastic trafficking mediated by plasmodesmata and viral movement proteins. However, recent evidence demonstrated that the plant RNA virus turnip mosaic virus (TuMV) components are associated with extracellular vesicles (EVs). EVs are membrane-bound structures secreted to the extracellular space to potentially mediate several plant-pathogen interactions such as cross-kingdom RNA interference or the delivery of stress response proteins. In animals, EVs facilitate viral transmission both within the host and across species, but knowledge about their potential roles in plant viral infection is scarce. In this study, we demonstrate that EVs isolated from geminivirus-infected plants contain complete viral genomes and both capsid and viral movement proteins. Furthermore, these EV fractions were demonstrated to be infectious when mechanically inoculated onto naive plants. This discovery suggests that EVs may serve as alternative carriers for geminivirus components, enabling long-range transport or potentially modulating host immune responses, and highlights geminiviral capacity to transverse membrane boundaries, essential for circulative arbovirus propagation in their insect vectors. Significance StatementViruses are obligate intracellular parasites that shape ecological communities and crucially challenge animal and plant health worldwide. Conversely to animal viruses, plant-infecting viruses rely on plasmodesmata to disseminate through their host and establish systemic infection. Nonetheless, most plant viruses are insect-transmitted whose ecological cycle relies on their insect vector spread. Thus, strategies to cross continuous membrane barriers are essential for their dissemination and may be potentially conserved in plant hosts. Our discovery that infectious viral entities are associated with EVs reveals an alternative pathway for geminiviral movement within plant hosts that could facilitate vector transmission, challenging our long-standing understanding of plant virus biology and expanding the current conception of plant viral pathology.

plant biology↗

bioRxiv2024

Root Expansion Microscopy (ROOT-ExM): A streamlined super resolution method for plants

Expansion microscopy (ExM) has revolutionized biological imaging by physically enlarging samples, surpassing the light diffraction limit and enabling nanoscale visualization using standard microscopes. While extensively employed across a wide range of biological samples, its application to plant tissues is sparse. In this work, we present ROOT-ExM, an expansion method suited for stiff and intricate multicellular plant tissues, focusing on the primary root of Arabidopsis thaliana. ROOT-ExM achieves isotropic expansion with a fourfold increase in resolution, enabling super-resolution microscopy comparable to STimulated Emission Depletion (STED) microscopy. Labelling is achieved through immunolocalization, compartment-specific dyes, and native fluorescence preservation, while N-Hydroxysuccinimide (NHS) ester-dye conjugates reveal the ultrastructural context of cells alongside specific labelling. We successfully applied ROOT-ExM to image various cellular structures, including the Golgi apparatus, the endoplasmic reticulum, the cytoskeleton, and wall-embedded structures such as plasmodesmata. When combined with lattice light sheet microscopy (LLSM), ROOT-ExM achieves 3D quantitative analysis of nanoscale cellular process, revealing increased vesicular fusion in close proximity of the cell plate during cell division. Achieving super-resolution fluorescence imaging in plant biology remains a formidable challenge. Our findings underscore that ROOT-ExM provides a remarkable, cost-effective solution to this challenge, paving the way for unprecedented insights into plant cellular subcellular architecture. One sentence summaryROOT-ExM achieves super-resolution expansion microscopy in plants

plant biology↗

bioRxiv2023

Multiplicity of Agrobacterium infection of Nicotiana benthamiana for transient DNA delivery

Biological DNA transfer into plant cells mediated by Agrobacterium represents one of the most powerful tools for the engineering and study of plant systems. Transient expression of transfer DNA (T-DNA) in particular enables rapid testing of gene products and has recently been harnessed for facile combinatorial expression of multiple genes. In analogous mammalian cell-based gene expression systems, a clear sense of the multiplicity of infection (MOI) allows users to predict and control viral transfection frequencies for applications requiring single vs. multiple transfection events per cell. Despite the value of Agrobacterium-mediated transient transformation of plants, MOI has not been quantified. Establishing MOI for Agrobacterium T-DNA delivery at the level of single plant cells would allow users to design genomic library delivery conditions (at most 1 event/cell), or maximize co-delivery of T-DNA loads from separate Agrobacterium (>1 event/cell). Here, we analyze the Poisson probability distribution of T-DNA transfer in leaf pavement cells to determine the MOI for the widely used model system Agrobacterium GV3101/Nicotiana benthamiana. These data delineate the relationship between an individual Agrobacterium strain infiltration OD600, plant cell perimeter and leaf age, as well as plant cell co-infection rates. Our analysis also establishes experimental regimes where the probability of near-simultaneous delivery of >20 unique T-DNAs to a given plant cell remains high throughout the leaf. We anticipate that these data will enable users to develop new approaches to in-leaf library development using Agrobacterium transient expression and the reliable combinatorial assaying of multiple heterologous proteins in a single plant cell.

plant biology↗

bioRxiv2023

Unraveling the Interplay of Different Traits and Parameters Related to Nitrogen Use Efficiency in Wheat: Insights for Grain Yield Influence

Enhancing Nitrogen Use Efficiency (NUE) in wheat to optimize grain yield is a significant challenge. To address this challenge, a comprehensive study was conducted to investigate various morphological, biochemical, molecular parameters, and agronomic traits related to NUE. By examining various traits under both optimum-N (ON) and stressed-N (SN) conditions, the study explores the interrelationships among these traits, providing novel insights not previously reported. A set of 278 diverse wheat genotypes were assessed, encompassing eight NUE-related traits: Grain Yield, Biomass, Grain nitrogen, N at head, N at harvest, N-uptake, Nitrogen Uptake Efficiency, Nitrogen Utilization Efficiency, and NUE under both ON and SN conditions in the field. The findings demonstrated a significant positive correlation between grain yield and all NUE-related traits, highlighting their significance in comprehending the biological NUE of wheat plants. Notably, the study identified N-uptake and N-uptake related traits as key factors influencing the impact of soil nitrogen status on yield and associated parameters. These traits hold particular importance for selecting wheat genotypes with optimal yield and NUE in wheat cultivation. To complement the field data, representative genotypes were further subjected to a hydroponics experiment under absolute nitrogen control. This experiment provided insights into the effects of nitrogen stress on morphological parameters and the performance of eight essential nitrogen and carbon metabolizing enzymes. Correlation analysis highlighted the substantial influence of four key N-metabolizing enzymes, namely Nitrate Reductase, Glutamine Synthetase, Glutamate Oxo-Glutarate Amino Transferase, and Glutamate Dehydrogenase, on grain yield. Additionally, this study underscored the direct and indirect associations between seedling parameters and field traits, emphasizing the significance of shoot and root length parameters in nitrogen acquisition under nitrogen stress. In conclusion, these findings offer valuable insights into the intricate network of traits and parameters that influence wheat grain yield under varying nitrogen regimes. This knowledge can aid in the selection of wheat genotypes with enhanced NUE and grain yield, particularly in scenarios of reduced nitrogen application. Key PointsO_LIA comprehensive study in field and hydroponics conditions revealed differential responses of various morphological, biochemical, molecular parameters, and agronomic traits to different nitrogen levels. C_LIO_LIN-uptake related traits in field condition and chlorophyll content and morphological parameters in hydroponics condition were found as essential factors contributing to variations under both optimum and stressed N conditions. C_LIO_LIAmong the parameters observed in the seedling stage, SL and RL, along with the enzymes NR, GS, GOGAT, and GDH, demonstrated their influence on GY. C_LI

plant biology↗

bioRxiv2023

Proteomic and Physiological Signatures of Altitude Adaptation in a Myrsine coriacea Population under Common Garden Conditions

Plants exhibit phenotypic plasticity in response to environmental variations, which can lead to stable genetic and physiological adaptations if exposure to specific conditions is prolonged. Myrsine coriacea demonstrates this through its ability to thrive in diverse environments. The objective of the article is to investigate the adaptive responses of M. coriacea by cultivating plants from seeds collected at four different altitudes in a common garden experiment. Through integrated physiological and proteomic analyses, we identified 170 differentially accumulated proteins and observed significant physiological differences among the populations. The high-altitude population (POP1) exhibited a unique proteomic profile with significant down-regulation of proteins involved in carbon fixation and energy metabolism, suggesting a potential reduction in photosynthetic efficiency. Physiological analyses showed lower leaf nitrogen content, net CO2 assimilation rate, specific leaf area, and relative growth rate in stem height for POP1, alongside higher leaf carbon isotopic composition ({delta}13C) and leaf carbon (C) content. These findings provide insight into the complex interplay between proteomic and physiological adaptations in M. coriacea and underscore the importance of local adaptations. This study enhances our understanding of how altitude-specific selection pressures can shape plant molecular biology and physiology, offering valuable perspectives for predicting plant responses to global environmental changes. HighlightThis study unveils proteomic and physiological adaptations in a high-altitude M. coriacea population with reduced carbon fixation and energy metabolism.

plant biology↗

bioRxiv2019

EXO70A2 is critical for the exocyst complex function in Arabidopsis pollen

Pollen development, pollen grain germination and pollen tube elongation are crucial biological processes in angiosperm plants that need precise regulation to deliver sperm cells to fertilize ovules. Pollen grains undergo two major developmental switches: dehydration characterized by metabolic quiescent state, and rehydration upon pollination that leads to extraordinary metabolic and membrane trafficking activity, resulting in germination and rapid tip growth of pollen tubes. To sustain these processes, many plant housekeeping genes evolved their pollen-specific paralogs. Highly polarized secretion at a growing pollen tube tip requires the exocyst tethering complex responsible for specific targeting of secretory vesicles to the plasma membrane. Here, we describe that EXO70A2 (At5g52340) is the main exocyst EXO70 isoform in Arabidopsis pollen, which governs the conventional secretory function of the exocyst, analogically to EXO70A1 (At5g03540) in the sporophyte. Our analysis of a CRISPR-generated exo70a2 mutant revealed that EXO70A2 is essential for efficient pollen maturation, pollen grain germination and pollen tube growth. GFP:EXO70A2 was localized similarly to other exocyst subunits to the apical domain in growing pollen tube tips characterized by intensive exocytosis. Moreover, EXO70A2 could substitute for the EXO70A1 function in the sporophyte, indicating functional redundancy of these two closely related isoforms. Phylogenetic analysis revealed that the ancient duplication of EXO70A to two (or more) paralogs, one of which is highly expressed in pollen, occurred independently in monocots and dicots. In summary, EXO70A2 is a crucial component of the exocyst complex in the Arabidopsis pollen required for efficient plant sexual reproduction.

plant biology↗

bioRxiv2026

Intronised fluorophores improve reporter output and biosensor expression in plants

Genetically encoded biosensors are central to plant cell biology, but low reporter signal can limit their use. Here, we developed modular intronised fluorophores to improve reporter output in transient and stable plant expression systems. Matched nuclear-localised triple mCitrine and mScarlet3 reporters were compared with and without introduced introns in Nicotiana benthamiana and Arabidopsis thaliana. Intronisation increased fluorescence, with the magnitude of enhancement depending on the fluorophore and expression context. Immunoblot analyses supported increased reporter accumulation. In Arabidopsis, intronised reporters driven by the tissue-specific SCARECROW promoter, produced stronger fluorescence signals that remained detectable farther from the root tip than non-intronised versions. Intronised versions of the abscisic acid (ABA) biosensor ABACUS2 retained ABA responsiveness and showed a lower frequency of reduced fluorescence between generations. These modular components provide a practical toolbox for improving fluorescent reporter detection and maintaining usable biosensor expression, supporting their application in plant cell biology and quantitative imaging.

plant biology↗

bioRxiv2021

HSFA1 proteins mediate heat-induced accumulation of CPT7-derived polyprenols affecting thylakoid organization

Polyprenols are ubiquitous isoprenoid compounds that accumulate in large quantities in plant photosynthetic tissues. While our knowledge of polyprenol biochemistry is constantly expanding, the regulation of their biosynthesis as well as the molecular basis of their cellular action are still poorly understood. In Arabidopsis, the polyprenols Pren-9, -10 and -11, synthesized by cis-prenyltransferase 7 (CPT7), are localized in plastidial membranes and affect the photosynthetic performance of chloroplasts. In this report we present evidence that plastidial polyprenols are among the major constituents of thylakoid membranes. Disturbances in polyprenol level, caused by alterations in CPT7 expression, change chloroplast ultrastructure, affect aggregation of LHCII complexes and modulate non-photochemical quenching (NPQ). Moreover, we show that Arabidopsis responds to high temperature by upregulating expression of CPT7 and increasing the accumulation of CPT7-derived polyprenols. These heat-induced changes in polyprenol biosynthesis are mediated by Heat Shock Transcription Factors of the HSFA1 family, the master regulators of heat stress response. Collectively, results presented in this report bring us closer to understanding the mechanisms by which polyprenols affect plant physiology and provide an additional link between chloroplast biology and plant responses to changing environmental conditions. One sentence summaryHeat Shock Transcription Factors induce biosynthesis of polyprenols - isoprenoid compounds that affect the organization and function of chloroplasts.

plant biology↗

bioRxiv2025

A molecular glue: LecRKI.9 creates stiff plasma membrane cell wall connections

Water stress challenges plasma membrane - wall attachment. When plants are exposed to strong hyperosmotic stress, water exits the cell leading to plasmolysis, where the plasma membrane detaches partially from the cell wall and reveals discrete and persistent attachment sites in the form of Hechtian strands. Although these structures have been observed in several species for more than a century, the molecular bases behind plasma membrane - wall attachment remains elusive. Through a screen of candidate proteins, we reveal that the overexpression of two lectin receptor-like proteins (LecRK-I.9 and LecTM) increases the density of plasma membrane - wall connections in N. Benthamiana. The extracellular lectin domain of LecRK-I.9 was able to remain in the wall during plasmolysis. Conversely, deletion of the lectin domain in the LecRK-I.9 overexpressor restored the density of Hechtian strands to WT levels. In Arabidopsis, upon hyperosmotic stress, LecRK-I.9 formed largely immobile clusters, whereas lectin-deleted versions of LecRK-I.9 clusters were mobile. Cluster density correlated with predicted tensile stress levels and mechanical reinforcement in the wall before plasmolysis, consistent with a scenario in which attachment of the lectin domain also reflects the wall properties. Last, while overexpressing LecRK-I.9 conferred resistance to low water potential conditions, deletion of the lectin domain in the LecRK-I.9 overexpressor restored a WT response to water stress. Altogether, this demonstrates that the lectin domain of LecRK-I.9 creates persistent plasma membrane - wall attachment sites, with physiological relevance for plant resistance to water stress. SIGNIFICANCE STATEMENTAs observed since the mid-19th century, when placed in hyperosmotic conditions, plant cells undergo plasmolysis and form thin membraneous threads called Hechtian strands. These structures reflect the presence of persistent plasma membrane - cell wall attachment sites. Yet, the molecular components behind these sites are unknown. Through a candidate screen approach, we identify and demonstrate that the lectin domain of a receptor-like kinase provides such molecular glue. We also show that the clustering of the receptor is patterned and correlate with mechanically reinforced cell walls. Conversely, we find that the promotion of attachment increases resistance to water stress. This work revisits an old plant cell biology classic, and opens new avenues of research for plant adaptation to their environment.

plant biology↗

bioRxiv2020

Computational modelling of cambium activity provides a regulatory framework for simulating radial plant growth

Precise organization of growing structures is a fundamental process in developmental biology. In plants, radial growth is mediated by the cambium, a stem cell niche continuously producing wood (xylem) and bast (phloem) in a strictly bidirectional manner. While this process contributes large parts to terrestrial biomass, cambium dynamics eludes direct experimental access due to obstacles in live cell imaging. Here, we present a cell-based computational model visualizing cambium activity and integrating the function of central cambium regulators. Performing iterative comparisons of plant and model anatomies, we conclude that the receptor- like kinase PXY and its ligand CLE41 are part of a minimal framework sufficient for instructing tissue organization. By integrating tissue-specific cell wall stability values, we moreover probe the influence of physical constraints on tissue geometry. Our model highlights the role of intercellular communication within the cambium and shows that a limited number of factors is sufficient to create radial growth by bidirectional tissue production. Impact statementRadial plant growth produces large parts of terrestrial biomass and can be computationally simulated with the help of an instructive framework of intercellular communication loops.

plant biology↗

bioRxiv2020

Foliar water uptake as a source of hydrogen and oxygen in plant biomass

Introductory biology lessons around the world typically teach that plants absorb water through their roots, but, unfortunately, absorption of water through leaves and subsequent transport and use of this water for biomass formation remains a field limited mostly to specialists. Recent studies have identified foliar water uptake as a significant but still unquantified net water source for terrestrial plants. The growing interest in the development of a new model that includes foliar uptake of liquid water to explain hydrogen and oxygen isotope ratios in leaf water and tree rings requires a method for distinguishing between these two water sources. I therefore devised a method utilizing two different heavy waters (HDO and H218O) to simultaneously label both foliar-uptake water and root-uptake water and quantify their relative contributions to plant biomass. Using this new method, I here present evidence that, in the case of well-watered Cryptomeria japonica, hydrogen and oxygen incorporated into new leaf cellulose in the rainy season derives mostly from foliar-uptake water, while that of new root cellulose derives mostly from root-uptake water, and new branch xylem is somewhere in between. Abandoning the assumption that these elements are supplied from soil water alone may have vast implications in fields ranging from isotope dendroclimatology, silviculture, to biogeochemistry.

plant biology↗