Biology subjects

Weijers, D.

Publications and source records attributed to Weijers, D..

At least 19 recordsLinked to original sources

bioRxiv2026

Phytosulfokine signaling modulates salt stress responses and cell wall remodeling in Arabidopsis

Salinity severely impairs plant growth and development. Increasing evidence suggests that the cell wall (CW) plays a central role in salt stress acclimation, not only as a structural barrier but also as a dynamic sensor that activates downstream stress signaling pathways. To identify extracellular factors involved in cell wall remodeling and integrity signaling during salt stress, we performed a comparative proteomic analysis of the Arabidopsis seedling apoplast. Among the proteins that accumulated under salt stress, we focused on three members of the Subtilisin-Like Protease family (SBT1.1, SBT1.6, and SBT5.3), which have previously been implicated in the processing of signaling peptides and CW-associated proteins. Functional analyses revealed that sbt1.1 and sbt5.3 mutants exhibit enhanced lignin deposition under salt stress, suggesting altered CW remodeling during stress application. Given the established role of SBT1.1 in processing PHYTOSULFOKINE (PSK) peptides, we investigated the involvement of PSK signaling in salt stress responses. Mutants lacking the two PSK RECEPTORS (pskr1 pskr2) displayed reduced growth and increased lignification under salt treatment. Exogenous application of PSK attenuated salt-induced responses, including MITOGEN ACTIVATED PROTEIN KINASE 6 (MPK6) phosphorylation, salt stress marker gene expression, and lignin accumulation, ultimately promoting root elongation under salt stress. Furthermore, PSK treatment mitigated salt-induced changes in CW composition. In cell wall integrity (CWI) mutants, PSK treatment failed to restore wild-type root growth under salt stress and induced a pronounced root bending phenotype in fer-4, indicating that CWI signaling influences PSK-mediated root growth responses. Together, these results support a role for PSK signaling in modulating CW-associated responses to salinity stress in Arabidopsis.

plant biology↗

bioRxiv2026

Systematic proteomics identifies a conserved mechanism for polar targeting of plant cortical proteins

Multicellular development is tightly coupled to the polarization of individual cells, which partitions polar proteins along the cell cortex and can control asymmetric cell division, anisotropic growth, local differentiation or physiology. Mechanisms driving cell polarization have been described in fungi and animals, but these lack counterparts in plants. While several polarized proteins have been identified in plants, the overall mechanisms guiding their polar localization are poorly characterized. Through iterative affinity proteomics on the recently identified SOSEKI polar proteins, we discovered a network of polar proteins that is conserved in the flowering plant Arabidopsis and the liverwort Marchantia. We next used this collection of novel polarized proteins for systematic proximity ligation proteomics in these two species to map their global polar proteome. We identified a subfamily of polarized Protein S-acyl transferase (PAT) enzymes that are required for membrane targeting of Arabidopsis SOSEKI proteins. Using human and fruit fly models, we showed that PAT19 is sufficient for membrane targeting of SOSEKI1, likely through direct palmitoylation. This work demonstrates a conserved mechanism for polar protein targeting in plants and offers a resource for studying polar protein localization.

plant biology↗

bioRxiv2026

Subnuclear cofactor partitioning underlies auxin-dependent transcriptional regulation

Cellular and organismal function relies on the precise activation and repression of gene expression by DNA-binding transcription factors (TFs). Many TFs occur in large gene families, and a key question in biology is how divergent functions emerge in TF families during evolution. Here we discover the biochemical mechanism for transcriptional activation by the Marchantia polymorpha AUXIN RESPONSE FACTOR1 (MpARF1) TF, which relies on direct recruitment of the Mediator complex into subnuclear MpARF1 clusters. We find that the Mediator recruitment region was the evolutionary innovation that converted ARF repressors into activators, switching binding specificity from co-repressor to co-activator. We demonstrate that this evolutionary innovation can be recreated, thereby revealing a deeply conserved mechanism based on competition between ARF clusters at the heart of the transcriptional auxin response.

plant biology↗

bioRxiv2026

High resolution, proteome-wide mapping of subcellular protein localization in plants

Protein function is intimately connected to subcellular localization, and experimental determination of protein localization is a key element of understanding biological roles. However, even in the best-studied model plants, such as Arabidopsis thaliana, a minority of proteins has an experimentally defined subcellular localization. We present an experimental strategy to globally map plant subcellular proteomes by mass spectrometry. We annotated subcellular localization of 7815 proteins in Arabidopsis roots, 4672 in Arabidopsis seedlings, and 2782 in the liverwort Marchantia polymorpha. By independent validation, we find that these annotations are highly predictive and can be integrated with other proteomics datasets. Cross-species comparisons reveal substantial global conservation of subcellular localization. Furthermore, we demonstrate that the same approach can be used to identify dynamically translocating proteins upon treatment or in a mutant. This work shows the power of global spatial proteome mapping in plants and offers an extensive resource for protein subcellular localization in plants. HighlightsO_LIOptimized approach for global mapping of protein subcellular localization by differential centrifugation in plants C_LIO_LIInteractive resource of subcellular localization of plant proteins at unprecedented depth and resolution C_LIO_LICross-species comparison reveals that the plant subcellular proteome is deeply conserved C_LIO_LIComparative subcellular proteomics of a Brefeldin A treatment and a gnom mutant robustly describes global shifts in protein localization C_LI

plant biology↗

bioRxiv2025

Nanoclustering of a plant transcription factor enables strong yet specific DNA binding

Transcription factors (TFs) are traditionally depicted as monomeric, or oligomeric, units that recognize and bind specific DNA sequences to regulate transcription. Emerging evidence suggests that many TFs can undergo liquid phase separation, resulting in condensates that bind DNA through a different mechanism. For the Auxin Response Factors (ARFs), canonical plant TFs, evidence for both scenarios exists. But which of these scenarios is operational in the plant nucleus is unclear. Here, we demonstrate using MpARF2 of Marchantia polymorpha that a third scenario is operational: MpARF2 forms nanoscopic clusters under physiological conditions. Nanoclusters combine DNA-binding features that cannot be accessed by the other two scenarios: high-affinity, switch-like, and sequence-specific. Our results suggest nanoclustering as a mechanism that equips TFs with the DNA-binding properties necessary for transcriptional regulation. TeaserTranscription factor nanoclusters balance the DNA-binding trade-off between weakly binding oligomers and nonspecific condensates.

plant biology↗

bioRxiv2025

Diversification of functional requirements for proteolysis of Auxin Response Factors

Auxin signaling through the Nuclear Auxin Pathway is essential for plant development and is mediated by competing A-class and B-class Auxin Response Factor (ARF) transcription factors. Recently, proteasomal ARF degradation through a degradation signal embedded within the DNA-Binding Domain, was identified as a key component of auxin response. Here, we investigate the structural requirements and biological relevance of ARF degradation in the bryophyte Marchantia polymorpha. We identify a critical residue for proteolysis of the repressive, B-class MpARF2, and find it to be functionally conserved in degradation of the activating, A-class MpARF1. Unlike MpARF2, however, impaired MpARF1 degradation had little effect on auxin response and development, suggesting differential integration in biological function. We find that MpARF2 degradation occurs across all developmental stages of the life cycle, and is required for MpARF2 function during development. Our findings reveal a degradation mechanism targeting A- and B-class ARFs which shares the same origin, but has evolved along unique evolutionary trajectories.

plant biology↗

bioRxiv2025

Transgenerational polarity axis inheritance during Ceratopteris embryogenesis

For sexually reproducing organisms to pass on their genetic information, progeny must successfully establish. Various life history strategies have evolved, using either dispersal of large numbers of progeny or intensive nurturing of a few. Most plants use the former strategy, but ferns generate a single embryo in the exact same location as the mother, and it is unknown how progeny success is promoted, or how embryogenesis is adapted. By studying Ceratopteris richardii embryogenesis, we find that maternal tissues guide orientation of the early embryo body axis, thus aligning its root pole towards the homologous maternal rhizoids. We find that axis polarity inheritance is mediated by maternal tissue mechanical patterns, and thus identify a robust mechanism for progeny establishment as a nurturing strategy in plants.

plant biology↗

bioRxiv2024

Auxin and tryptophan trigger common responses in the streptophyte alga Penium margaritaceum

Auxin is a signaling molecule that regulates multiple processes in the growth and development of land plants. Research gathered from model species, particularly Arabidopsis thaliana, has revealed that the nuclear auxin pathway controls many of these processes through transcriptional regulation. Recently, a non-transcriptional pathway based on rapid phosphorylation mediated by kinases has been described, complementing the understanding of the complexity of auxin-regulated processes. Phylogenetic inferences of both pathways indicate that only some of the components are conserved beyond land plants. This raises fundamental questions about the evolutionary origin of auxin responses and whether algal sisters share mechanistic features with land plants. Here we explore auxin responses in the unicellular streptophyte alga Penium margaritaceum. By assessing physiological, transcriptomic and cellular responses we found that auxin triggers cell proliferation, gene regulation and acceleration of cytoplasmic streaming. Notably, all these responses are also triggered by the structurally related tryptophan. These results identify shared auxin response features among land plants and algae, and suggest that less chemically specific responses preceded the emergence of auxin-specific regulatory networks in land plants.

plant biology↗

bioRxiv2024

Metapredict enables accurate disorder prediction across the Tree of Life

Intrinsically disordered proteins and protein regions (collectively IDRs) are critical in numerous cellular processes. To understand how IDRs facilitate function, we need tools to accurately and rapidly identify them from sequence. While many methods for disorder prediction exist, we are currently limited by throughput and accuracy for evolutionary scale analyses. To bridge this gap, we developed metapredict V3, an updated version of our disorder predictor that enables evolutionary-scale disorder prediction. Metapredict V3 enables proteome-scale prediction with state-of-the-art accuracy in seconds and was developed with a focus on usability. It is distributed as a web server, Python software package, command-line interface, and Google Colab notebook. Here, we leverage the accuracy and throughput of metapredict V3 to predict disorder for over 20,000 proteomes to evaluate the prevalence of disorder across the kingdoms of life.

bioinformatics↗

bioRxiv2024

An ultra-conserved ARF-DNA interface underlies auxin-triggered transcriptional response

Auxin Response Factor (ARF) plant transcription factors are the key effectors in auxin signalling. Their DNA-Binding Domain (DBD) contains a B3 domain that allows base-specific interactions with Auxin Response Elements (AuxREs) in DNA target sites. Land plants encode three phylogenetically distinct ARF classes: the closely related A- and B-classes have overlapping DNA binding properties, contrasting with the different DNA-binding properties of the divergent C-class ARFs. ARF DNA-binding divergence likely occurred early in the evolution of the gene family, but the molecular determinants underlying it remain unclear. Here, we show that the B3 DNA-binding residues are deeply conserved in ARFs, and variability within these is only present in tracheophytes, correlating with greatly expanded ARF families. Using the liverwort Marchantia polymorpha, we confirm the essential role of conserved DNA-contacting residues for ARF function. We further show that ARF B3-AuxRE interfaces are not mutation-tolerant, suggesting low evolvability that has led to the ultra-conservation of the B3-DNA interface between ARF classes. Our data support the almost complete interchangeability between A/B-class ARF B3 by performing interspecies domain swaps, even between lineages that diverged over half a billion years ago. Our analysis further suggests that DNA-binding specificity diverged early during ARF evolution in a common streptophyte ancestor, followed by strong selection as part of a competition-based auxin response system Significance StatementAuxin response evolved nearly half a billion years ago in the earliest land plants. Auxin response is mediated by a family of DNA-binding ARF transcription factors. It has been unclear if and how the ARF family has evolved. In this paper, the authors show that the central protein-DNA interface that defines the genes that are under auxin control has remained essentially unchanged throughout auxin response evolution, explaining how auxin has become a dominant signal controlling growth and development in all land plants.

plant biology↗

bioRxiv2024

The role of IAA and its transport in the complex streptophyte algae Chara braunii

The role of auxin indole-3-acetic acid (IAA), a phytohormone with numerous morphogenic functions, is now well-established in land plants. Although the role of IAA and its transport in algae remains unclear, PIN-driven auxin export is probably an ancient and conserved trait within Streptophytes. Among streptophyte algae, Chara represents a genus with a considerable degree of complexity in body arrangement. This study investigates the auxin response and characterizes homologs of land plant PIN auxin efflux carriers in Chara braunii. Through regeneration experiments, we observed that IAA significantly promotes elongation of thallus and side branch development upon thallus tip decapitation, indicating an effect on morphogenesis. We show that IAA is actively uptaken and metabolized by thallus cells and that this process is influenced by N-1-naphthylphthalamic acid (NPA). To elucidate the underlying mechanisms, we cloned and sequenced the most expressed Chara braunii PINs, CbPINa and CbPINc. Using epitope-specific antibodies, we showed their presence in the plasma membrane (PM) of vegetative internodal cells and generative antheridial cells. Functional tests in tobacco BY-2 cells, supported by in silico docking of IAA and NPA to CbPINa and CbPINc, showed that PM-localized CbPINa interferes with auxin transport in contrast to ER-localized CbPINc. Finally, our phosphoproteome analysis indicated that IAA rapidly induces specific phosphorylation events, including RAF-like kinase phosphorylation, highlighting a potential role for IAA in fast signaling processes in Chara braunii. Altogether, we provide new insights into IAA role in Chara braunii morphogenesis and suggest that while the canonical auxin transport mechanism may not be conserved, auxin still may likely play a role in rapid signaling pathways in this close relative of land plants.

plant biology↗

bioRxiv2024

ARF degradation defines a deeply conserved step in auxin response

Auxin response critically depends on the concentrations and stoichiometry of competing A- and B-class AUXIN RESPONSE FACTOR (ARF) proteins. In Marchantia polymorpha, both A- and B-ARFs are unstable, and here we identify a minimal necessary and sufficient region for ARF degradation that is critical for development, and auxin response. Through comparative analysis, we find that ARF instability likely preceded the emergence of the auxin response system.

plant biology↗

bioRxiv2024

ETTIN-mediated auxin signalling is an angiosperm-specific neofunctionalization for carpel development

The phytohormone auxin affects processes throughout plant growth and development. While auxin signalling has been mainly attributed to a repressor degradation-based pathway, numerous alternative mechanisms for how auxin mediates its effect have been revealed in recent years. One such mechanism involves a direct auxin-induced switch in the transcriptional regulatory activity of the Auxin Response Factor (ARF) ETTIN (ETT). ETT lacks a conserved C-terminus domain involved in canonical pathway interactions but contains a middle region domain mediating auxin binding. As the ETT clade only exists in the angiosperms, it remains unknown when the pathway evolved. Here we provide evidence for a two-step origin of the ETT clade and its neofunctionalisation through the gain of auxin perception in gynoecium patterning. Phylogenetic analyses reveal the structural divergence of ETT and its paralogue ARF4 after their duplication from an ancestral euphyllophyte ARF3/4 clade. Auxin sensitivity was identified as an ETT-specific innovation that likely originated in the last common angiosperm ancestor. Furthermore, in planta complementation experiments demonstrated the full genetic redundancy of ETT and ARF4 in leaf and ovary development, but a specialised role for the ETT-mediated auxin signalling pathway in style development. Our work thus provides evidence that ETT was recruited from an ancestral role in leaf development and subsequently underwent neofunctionalisation through the acquisition of direct auxin sensing for a novel role in gynoecium patterning.

evolutionary biology↗

bioRxiv2024

Evolutionary Origins and Functional Diversification of Auxin Response Factors

The Auxin Response Factors (ARFs) family of transcription factors are the central mediators of auxin-triggered transcriptional regulation. Functionally different classes of extant ARFs operate as antagonistic auxin-dependent and -independent regulators. While part of the evolutionary trajectory to the present auxin response functions has been reconstructed, it is unclear how ARFs emerged, and how early diversification led to functionally different proteins. Here, we use in silico and in vivo analyses to revisit the molecular events that led to the origin and subsequent evolution of the ARFs. We reveal the shared origin of ARFs from preexisting domains, uncovering a protein fold homologous to the ARF DNA-binding fold in a conserved eukaryotic chromatin regulator. Building on this, we reconstruct the complete evolutionary history of ARFs, including the divergence events leading to the appearance of the ARF classes and defining the main molecular targets for their functional diversification. We derive a complete evolutionary trajectory that led to the emergence of the nuclear auxin signalling pathway.

plant biology↗

bioRxiv2024

Mechanism of auxin-dependent gene regulation through composite auxin response elements

The plant signaling molecule auxin controls growth and development, largely through activating and repressing the expression of thousands of genes. Auxin-dependent transcriptional changes are mediated by DNA-binding Auxin Response Factors (ARF), whose AuxRE DNA binding sites are well-known. The identification of the first AuxRE showed this to be part of a composite element with a second motif. Indeed, systematic analysis showed other DNA motifs to be enriched in auxin-regulated promoters. Neither the basis for this enrichment nor the mechanisms for the activity of composite AuxREs is known. Here, we systematically mined Arabidopsis promoters for composite AuxRE elements enriched in auxin-responsive genes. We identified many and show that their presence is a reliable predictor of auxin response. Through mutating these elements and their higher-order modules, we demonstrate function in promoter activity. Lastly, we identified transcription factors (TFs) that bind AuxRE-associated motifs, showed their involvement in auxin response, and discovered that several of these TFs directly bind ARF proteins. We propose that ARF-TF complexes specifically bind compound motifs in promoters, and act as a source of diversification in auxin-dependent gene regulation.

plant biology↗

bioRxiv2024

CarboTag: a modular approach for live and functional imaging of plant cell walls

Plant cells are contained inside a rigid network of cell walls. Cell walls are highly dynamic structures that act both as a structural material and as a hub for a wide range of signaling processes. Despite its crucial role in all aspects of the plant life cycle, live dynamical imaging of the cell wall and its functional properties has remained challenging. Here, we introduce CarboTag, a modular toolbox for live functional imaging of plant walls. CarboTag relies on a small molecular motif, a pyridine boronic acid, that targets its cargo to the cell wall, is non-toxic and ensures rapid tissue permeation. We designed a suite of cell wall imaging probes based on CarboTag in any desired color for multiplexing. Moreover, we created new functional reporters for live quantitative imaging of key cell wall features: network porosity, cell wall pH and the presence of reactive oxygen species. CarboTag opens the way to dynamical and quantitative mapping of cell wall responses at subcellular resolution.

plant biology↗

bioRxiv2024

Analysis of auxin responses in the fern Ceratopteris richardii identifies tissue ontogeny as a major determinant for response properties

The auxin signalling molecule regulates a range of plant growth and developmental processes. The core transcriptional machinery responsible for auxin-mediated responses is conserved across all land plants. Genetic, physiological and molecular exploration in bryophyte and angiosperm model species have shown both qualitative and quantitative differences in auxin responses. Given the highly divergent ontogeny of the dominant gametophyte (bryophytes) and sporophyte (angiosperms) generations, however, it is unclear whether such differences derive from distinct phylogeny or ontogeny. Here, we address this question by comparing a range of physiological, developmental and molecular responses to auxin in both generations of the model fern Ceratopteris richardii. We find that auxin response in Ceratopteris gametophytes closely resembles that of a thalloid bryophyte, whereas the sporophyte mimics auxin response in flowering plants. This resemblance manifests both at phenotypic and transcriptional level. Furthermore, we show that disrupting auxin transport can lead to ectopic sporophyte induction on the gametophyte, suggesting a role for auxin in the alternation of generations. Our study thus identifies ontogeny, rather than phylogeny, as a major determinant of auxin response properties in land plants. Summary statementStudies in angiosperms and bryophytes have left unresolved the roles of tissue ontogeny and species phylogeny in auxin response. We address that problem by characterizing auxin response in a fern.

plant biology↗

bioRxiv2023

An elastic proteinaceous envelope encapsulates the early Arabidopsis embryo

Plant external surfaces are often covered by barriers that control the exchange of molecules, protect from pathogens, and offer mechanical integrity. A key question is when and how such surface barriers are generated. Post-embryonic surfaces have well-studied barriers, including the cuticle, and late Arabidopsis embryo was shown to be protected by an endosperm-derived sheath deposited onto a primordial cuticle. Here we show that both cuticle and sheath are preceded by another structure during the earliest stages of embryogenesis. This structure, which we named the embryonic envelope, is tightly wrapped around the embryonic surface but can be physically detached by cell wall digestion. We show that this structure is composed primarily of Extensin and Arabinogalactan O-glycoproteins and lipids, which appear to form a dense and elastic crosslinked embryonic envelope. The envelope forms in cuticle-deficient mutants and in a mutant that lacks endosperm. This embryo-derived envelope is therefore distinct from previously described cuticle and sheath structures. We propose that it acts as an expandable diffusion barrier, as well as a means to mechanically confine the embryo to maintain its tensegrity during early embryogenesis. Summary statementThe early Arabidopsis embryo is surrounded by a proteinaceous envelope that is distinct from the cuticle or embryo sheath

plant biology↗