Biology subjects

Weijers, D.

Publications and source records attributed to Weijers, D..

At least 37 records · Page 2Linked to original sources

bioRxiv2022

An ultra-fast, proteome-wide response to the plant hormone auxin

The plant signaling molecule auxin controls growth and development through a simple nuclear pathway that regulates gene expresion. There are however several cellular and physiological responses to auxin that occur within seconds, far too rapid to be mediated by transcriptional changes, for which no molecular mechanism has yet been identified. Using a phosphoproteomic strategy in Arabidopsis thaliana roots, we identify an ultra-rapid auxin response system that targets over 1700 proteins, many within 30 seconds. Auxin response is chemically specific, requires known auxin-binding proteins, and targets various pathways. Through exploring its temporal dynamics, we infer auxin-triggered kinase-substrate networks and identify apoplastic pH changes as a target of signaling and as part of a relay mechanism. By generating a variety of phosphoproteomic datasets, integrated with structural information in a web-app, and by demonstrating analysis and inference strategies, we offer a resource to explore rapid and dynamic signaling in plants.

plant biology↗

bioRxiv2022

A RAF-like kinase mediates a deeply conserved, ultra-rapid auxin response

The plant signaling molecule auxin triggers both fast and slow cellular responses across the plant kingdom, including both land plants and algae. A nuclear response pathway mediates auxin-dependent gene expression, and controls a range of growth and developmental processes in land plants. It is unknown what mechanisms underlie both the physiological responses occurring within seconds, and the responses in algae, that lack the nuclear auxin response pathway. We discovered an ultra-fast proteome-wide phosphorylation response to auxin across 5 land plant and algal species, converging on a core group of shared target proteins. We find conserved rapid physiological responses to auxin in the same species and identified a RAF-like protein kinase as a central mediator of auxin-triggered phosphorylation across species. Genetic analysis allowed to connect this kinase to both auxin-triggered protein phosphorylation and a rapid cellular response, thus identifying an ancient mechanism for fast auxin responses in the green lineage.

plant biology↗

bioRxiv2022

Cooperative action of separate interaction domains promotes high-affinity DNA binding of Arabidopsis thaliana ARF transcription factors

The signaling molecule auxin is pivotal in coordinating many growth and development processes in plants mainly through the modulation of gene expression. The transcriptional response to auxin is mediated by the family of auxin response factors (ARF). Monomers of this family recognize a DNA motif (TGTC[TC]/[GG]) called the auxin-response element (AuxRE). ARFs can homodimerize through their DNA binding domains (DBD) thereby enabling cooperative binding for a bipartite inverted AuxRE (IR7). In addition to the DBD, most ARFs contain a C-terminal Phox and Bem1p (PB1) domain both capable of homotypic interactions, and mediating interactions with Aux/IAA repressors. Given the dual role of the PB1 domain, and the ability of both DBD and PB1 domain to mediate dimerization, a key question is how each of these domains contributes to conferring DNA-binding specificity and affinity. So far, ARF-ARF and ARF-DNA interactions have mostly been approached using qualitative methods that do not provide a quantitative and dynamic view on the binding equilibria. Here, we utilize a DNA binding assay based on single-molecule Forster resonance energy transfer (smFRET) to study the affinity and kinetics of the interaction of several Arabidopsis thaliana ARFs with an IR7 AuxRE. We show that both DBD and PB1 domains of AtARF2 contribute toward DNA binding, and we identify ARF dimer stability as a key parameter in defining affinity and kinetics seen for the DBDs of different AtARFs. Lastly, we derived an analytical solution for a four-state cyclic model that explains both the kinetics and the affinity of the interaction between AtARF2 and IR7. Our work demonstrates that the affinity of ARFs towards composite DNA response elements can be tuned by small changes of their dimerization equilibrium suggesting that this effect has major implications for ARF-mediated transcriptional activity.

biochemistry↗

bioRxiv2022

Selective degradation of ARF monomers controls auxin response in Marchantia

The plant signaling molecule auxin controls a variety of growth and developmental processes in land plants. Auxin regulates gene expression through a nuclear auxin signaling pathway (NAP) consisting of a ubiquitin ligase auxin receptor TIR1/AFB, its Aux/IAA degradation substrate, and the DNA-binding ARF transcription factors. While extensive qualitative understanding of the pathway and its interactions has been obtained by studying the flowering plant Arabidopsis thaliana, it is so far unknown how these translate to quantitative system behaviour in vivo, a problem that is confounded by large NAP gene families in this species. Here we used the minimal NAP of the liverwort Marchantia polymorpha to quantitatively map NAP protein accumulation and dynamics in vivo through the use of knock-in fluorescent fusion proteins. Beyond revealing the native accumulation profile of the entire NAP protein network, we discovered that the two central ARFs MpARF1 and MpARF2 are proteasomally degraded. This degradation serves two functions: it tunes the stoichiometry of auxin-responsive, positively acting MpARF1 and auxin-independent, negatively acting MpARF2, thereby permitting auxin response. Secondly, through mapping a minimal degradation motif, we found that degradation is likely selective for MpARF2 monomers and favours accumulation of dimers. Interfering with MpARF1:MpARF2 stoichiometry or preventing degradation of MpARF2 monomers caused strong growth defects associated with auxin response defects. Thus, quantitative analysis of the entire Marchantia NAP, allowed to identify a novel regulatory mechanism in auxin response, built on regulated ARF degradation.

plant biology↗

bioRxiv2022

The Proxitome of Arabidopsis Processing Bodies Reveals a Condensate-Membrane Interface Instructing Actin-Driven Directional Growth

Cellular condensates can comprise membrane-less ribonucleoprotein assemblies with liquid-like properties. These cellular condensates influence various biological outcomes, but their liquidity hampers their isolation and characterization. Here, we investigated the composition of the condensates known as processing bodies (PBs) in the model plant Arabidopsis through a proximity-biotinylation pipeline. Using in situ protein-protein interaction approaches, genetics and high-resolution imaging, we show that PBs comprise networks that interface with membranes. Surprisingly, the conserved component of PBs, DECAPPING PROTEIN 1 (DCP1), can interface with unique plasma membrane subdomains that include cell edges and vertices. We characterized these plasma membrane interfaces and discovered a developmental module that can control cell shape. This module is modulated by the liquid-like properties of DCP1 and the actin-nucleating SCAR-WAVE complex, whereby the DCP1-SCAR-WAVE interaction confines actin nucleation. This study reveals an unexpected repertoire for a conserved condensate at unique membrane-condensate interfaces.

cell biology↗

bioRxiv2022

Local light signalling at the leaf tip drives remote differential petiole growth through auxin-gibberellin dynamics

Although plants are immobile, many of their organs are flexible to move in response to environmental cues. In dense vegetation plants detect neighbours through far-red light perception with their leaf tip. They respond remotely, with asymmetrical growth between the abaxial and adaxial sides of the leafstalk, the petiole. This results in upward movement that brings the leaf blades into better lit zones of the canopy. The plant hormone auxin is required for this response, but it is not understood how non-differential leaf tip-derived auxin can remotely regulate movement. Here we show that remote light signalling promotes auxin accumulation in the abaxial petiole by reinforcing an intrinsic auxin transport directionality. In the petiole, auxin elicits a response of both auxin as well as a second growth promoter; gibberellin. We show that this dual regulation is necessary for hyponastic leaf movement in response to light. Our results reveal how plants can spatially relay information about neighbour proximity from their sensory leaf tips to the petiole base, thus driving adaptive growth.

plant biology↗

bioRxiv2021

Probing DNA - transcription factor interactions using single-molecule fluorescence detection in nanofluidic devices

Single-molecule fluorescence detection offers powerful ways to study biomolecules and their complex interactions. Here, we combine nanofluidic devices and camera-based, single-molecule Forster resonance energy transfer (smFRET) detection to study the interactions between plant transcription factors of the auxin response family (ARF) and DNA oligonucleotides that contain target DNA response elements. In particular, we show that the binding of the unlabelled ARF DNA binding domain (ARF-DBD) to donor and acceptor labelled DNA oligonucleotides can be detected by changes in the FRET efficiency and changes in the diffusion coefficient of the DNA. In addition, our data on fluorescently labelled ARF-DBDs suggest that, at nanomolar concentrations, ARF-DBDs are exclusively present as monomers. In general, the fluidic framework of freely diffusing molecules minimizes potential surface-induced artifacts, enables high-throughput measurements and proved to be instrumental in shedding more light on the interactions between ARF-DBDs monomers and between ARF-DBDs and their DNA response element.

biophysics↗

bioRxiv2021

Rapid auxin-mediated phosphorylation of Myosin regulates trafficking and polarity in Arabidopsis

The signaling molecule auxin controls plant development through a well-known transcriptional mechanism that regulates many genes. However, auxin also triggers cellular responses within seconds or minutes, and mechanisms mediating such fast responses have remained elusive. Here, we identified an ultrafast auxin-mediated protein phosphorylation response in Arabidopsis roots that is largely independent of the canonical TIR1/AFB receptors. Among targets of this novel response are Myosin XI and its adaptor protein MadB2. We show that their auxin-mediated phosphorylation regulates trafficking and polar, subcellular distribution of PIN auxin transporters. This phosphorylation-based auxin signaling module is indispensable during developmental processes that rely on auxin-mediated PIN repolarization, such as termination of shoot gravitropic bending or vasculature formation and regeneration. Hence, we identified a fast, non-canonical auxin response targeting multiple cellular processes and revealed auxin-triggered phosphorylation of a myosin complex as the mechanism for feedback regulation of directional auxin transport, a central component of auxin canalization, which underlies self-organizing plant development.

plant biology↗

bioRxiv2021

Conserved, divergent and heterochronic gene expression during Brachypodium and Arabidopsis embryo development

Embryogenesis, transforming the zygote into the mature embryo, represents a fundamental process for all flowering plants. Current knowledge of cell specification and differentiation during plant embryogenesis is largely based on studies of the dicot model plant Arabidopsis thaliana. However, the major crops are monocots and the transcriptional programs associated with the differentiation processes during embryogenesis in this clade were largely unknown. Here, we combined analysis of cell division patterns with development of a temporal transcriptomic resource during embryogenesis of the monocot model plant Brachypodium distachyon. We found that early divisions of the Brachypodium embryo were highly regular, while later stages were marked by less stereotypic patterns. Comparative transcriptomic analysis between Brachypodium and Arabidopsis revealed that the early and late embryogenesis shared a common transcriptional program, whereas mid-embryogenesis was divergent between species. Analysis of orthology groups revealed widespread heterochronic expression of potential developmental regulators between the species. Interestingly, Brachypodium genes tend to be expressed at earlier stages than Arabidopsis counterparts, which suggests that embryo patterning may occur early during Brachypodium embryogenesis. Detailed investigation of auxin-related genes shows that the capacity to synthesize, transport, and respond to auxin is established early in the embryo. However, while early PIN1 polarity could be confirmed, it is unclear if an active response is mounted. This study presents a resource for studying Brachypodium and grass embryogenesis, and shows that divergent angiosperms share a conserved genetic program that is marked by heterochronic gene expression. Key messageDevelopmental and transcriptomic analysis of Brachypodium embryogenesis, and comparison with Arabidopsis, identifies conserved and divergent phases of embryogenesis, and reveals widespread heterochrony of developmental gene expression.

plant biology↗

bioRxiv2021

A molecular framework for control of oriented cell division in the Arabidopsis embryo

Premitotic control of cell division orientation is critical for plant development, as cell walls prevent extensive cell remodelling or migration. Whilst many divisions are proliferative and add cells to existing tissues, some divisions are formative, and generate new tissue layers or growth axes. Such formative divisions are often asymmetric in nature, producing daughters with different fates. We have previously shown that in the Arabidopsis thaliana embryo, developmental asymmetry is correlated with geometric asymmetry, creating daughter cells of unequal volume. Such divisions are generated by division planes that deviate from a default "minimal surface area" rule. Inhibition of auxin response leads to reversal to this default, yet the mechanisms underlying division plane choice in the embryo have been unclear. Here we show that auxin-dependent division plane control involves alterations in cell geometry, but not in cell polarity or nuclear position. Through transcriptome profiling, we find that auxin regulates genes controlling cell wall and cytoskeleton properties. We confirm the involvement of microtubule (MT)-binding proteins in embryo division control. Topology of both MT and Actin cytoskeleton depend on auxin response, and genetically controlled MT or Actin depolymerization in embryos leads to disruption of asymmetric divisions, including reversion to the default. Our work shows how auxin-dependent control of MT- and Actin cytoskeleton properties interacts with cell geometry to generate asymmetric divisions during the earliest steps in plant development.

plant biology↗

bioRxiv2020

Suspensor-derived somatic embryogenesis in Arabidopsis

In many flowering plants, including Arabidopsis thaliana, asymmetric division of the zygote generates apical and basal cells with different fates. The apical cell continues to produce the embryo while the basal cell undergoes a restricted number of anticlinal divisions leading to a suspensor of 6-9 quiescent cells that remain extra-embryonic and eventually senesce. In some genetic backgrounds, or upon ablation of the embryo, suspensor cells can however undergo periclinal cell divisions and eventually form a second, twin seedling. Likewise, embryogenesis can be induced from somatic cells by various genes, but the relation to suspensor-derived embryos is unclear. Here, we addressed the nature of the suspensor to embryo fate transformation, and its genetic triggers. We expressed most known embryogenesis-inducing transcriptional regulators and receptor-like kinases specifically in suspensor cells. Among these, only RKD1 and WUS could induce a heritable twin seedling phenotype. We next analyzed morphology and fate marker expression in embryos in which suspensor division were activated by different triggers to address the developmental paths towards reprogramming. Our results show that reprogramming of Arabidopsis suspensor cells towards embryonic identity is a specific cellular response that is triggered by defined regulators, follows a conserved developmental trajectory and shares similarity to the process of somatic embryogenesis from post-embryonic tissues.

plant biology↗

bioRxiv2020

Deep Evolutionary History of the Phox and Bem1 (PB1) Domain Across Eukaryotes

Protein oligomerization is a fundamental process to build complex functional modules. Domains that facilitate the oligomerization process are diverse and widespread in nature across all kingdoms of life. One such domain is the Phox and Bem1 (PB1) domain, which is functionally (relatively) well understood in the animal kingdom. However, beyond animals, neither the origin nor the evolutionary patterns of PB1-containing proteins are understood. While PB1 domain proteins have been found in other kingdoms, including plants, it is unclear how these relate to animal PB1 proteins. To address this question, we utilized large transcriptome datasets along with the proteomes of a broad range of species. We discovered eight PB1 domain-containing protein families in plants, along with three each in Protozoa and Chromista and four families in Fungi. Studying the deep evolutionary history of PB1 domains throughout eukaryotes revealed the presence of at least two, but likely three, ancestral PB1 copies in the Last Eukaryotic Common Ancestor (LECA). These three ancestral copies gave rise to multiple orthologues later in evolution. Tertiary structural models of these plant PB1 families, combined with Random Forest based classification, indicated family-specific differences attributed to the length of PB1 domain and the proportion of {beta}-sheets. This study identifies novel PB1 families and reveals considerable complexity in the protein oligomerization potential at the origin of eukaryotes. The newly identified relationships provide an evolutionary basis to understand the diverse functional interactions of key regulatory proteins carrying PB1 domains across eukaryotic life.

evolutionary biology↗

bioRxiv2019

Initiation and regulation of vascular tissue identity in the Arabidopsis embryo

Development of plant vascular tissues involves tissue specification, growth, pattern formation and cell type differentiation. While later steps are understood in some detail, it is still largely unknown how the tissue is initially specified. We have used the early Arabidopsis embryo as a simple model to study this process. Using a large collection of marker genes, we find that vascular identity is established in the 16-cell embryo. After a transient precursor state however, there is no persistent uniform tissue identity. Auxin is intimately connected to vascular tissue development. We find that while AUXIN RESPONSE FACTOR5/MONOPTEROS/ (ARF5/MP)-dependent auxin response is required, it is not sufficient for tissue establishment. We therefore used a large-scale enhanced Yeast One Hybrid assay to identify potential regulators of vascular identity. Network and functional analysis of suggest that vascular identity is under robust, complex control. We found that one candidate regulator, the G-class bZIP transcription factor GBF2, modulates vascular gene expression, along with its homolog GBF1. Furthermore, GBFs bind to MP and modulate its activity. Our work uncovers components of a gene regulatory network that controls the initiation of vascular tissue identity, one of which involves the interaction of MP and GBF2 proteins.

plant biology↗

bioRxiv2019

Design principles of a minimal auxin response system

Auxin controls numerous growth processes in land plants through a gene expression system that modulates ARF transcription factor activity1-3. Gene duplications in families encoding auxin response components have generated tremendous complexity in most land plants, and neofunctionalization enabled various unique response outputs during development2-4. However, it is unclear what fundamental biochemical principles underlie this complex response system. By studying the minimal system in Marchantia polymorpha, we derive an intuitive and simple model where a single auxin-dependent A-ARF activates gene expression. It is antagonized by an auxin-independent B-ARF that represses common target genes. Expression patterns of both ARF proteins define developmental zones where auxin response is permitted, quantitatively tuned, or prevented. This fundamental design likely represents the ancestral system, and formed the basis for inflated, complex systems.

plant biology↗

bioRxiv2019

Evolution of vascular plants through redeployment of ancient developmental regulators

Vascular plants provide most of the biomass, food and feed on earth; yet the molecular innovations that led to the evolution of their conductive tissues are unknown. Here, we reveal the evolutionary trajectory for the heterodimeric TMO5/LHW transcription factor complex, which is rate-limiting for vascular cell proliferation in Arabidopsis thaliana. Both regulators have origins predating vascular tissue emergence, and even terrestrialization. We further show that TMO5 evolved its modern function, including dimerization with LHW, at the origin of land plants. A second innovation in LHW, coinciding with vascular plant emergence, conditioned obligate heterodimerization and generated the critical function in vascular development. In summary, our results suggest that division potential of vascular cells may have been a major driver in the evolution of vascular plants.

plant biology↗

bioRxiv2018

A SOSEKI-based coordinate system interprets global polarity cues in Arabidopsis

Multicellular development requires coordinated cell polarization relative to body axes, and translation to oriented cell division. In plants, it is unknown how cell polarities are connected to organismal axes and translated to division. Here, we identify Arabidopsis SOSEKI (SOK) proteins that integrate apical-basal and radial organismal axes to localize to polar cell edges. Localization does not depend on tissue context, requires cell wall integrity and is defined by a transferrable, protein-specific motif. SOK proteins structurally resemble the DIX oligomerization domain in the animal Dishevelled polarity regulator. The DIX-like domain self-interacts and is required for edge localization and for influencing division orientation. Our work identifies a plant compass, interpreted by SOK proteins. Furthermore, despite fundamental differences, polarity in plants and animals converge upon the same protein domain.

plant biology↗

bioRxiv2018

Maternal inflammation significantly impacts cortical interneuron development in a subtype-specific manner

Severe infections during pregnancy are one of the major risk factors for cognitive brain impairment in offspring. It has been suggested that maternal inflammation leads to dysfunction of cortical GABAergic interneurons that in turn underlies cognitive impairment of the affected offspring. However, the evidence comes largely from studies of adult or mature brain and how impairment of inhibitory circuits arises upon maternal inflammation is unknown. Here we show that maternal inflammation affects multiple steps of cortical GABAergic interneuron development, i.e. proliferation of precursor cells, migration and positioning of neuroblasts as well as neuronal maturation. Importantly, the development of distinct subtypes of cortical GABAergic interneurons was discretely impaired as a result of maternal inflammation. This translated into a reduction in cell numbers, redistribution across cortical regions and layers, changes in morphology and cellular properties. Furthermore, selective vulnerability of GABAergic interneuron subtypes was associated with the stage of brain development. Thus, we propose that maternally-derived insults have developmental stage-dependent effects which contribute to the complex etiology of cognitive impairment in the affected offspring.

neuroscience↗

bioRxiv2018

OsIQD14 regulates rice grain shape through modulating the microtubule cytoskeleton

Cortical microtubule (MT) arrays play a critical role in plant cell shape determination by defining the direction of cell expansion1-3. The control of plant organ shape and architecture is a major target of cereal crop improvement. Given the pleiotropic effects of MT modification, however, it is challenging to exploiting MT array organization for crop improvement. Moreover, as plants continuously adapt cell growth and expansion to ever-changing environmental conditions, multiple environmental (e.g. light4) and developmental (e.g. hormones5,6) inputs need to be translated into changes of the MT cytoskeleton. Here, we identify and functionally characterize an auxin-inducible and MT-localized protein OsIQ67-DOMAIN14 (OsIQD14), which is highly expressed in rice seed hull cells. While deficiency of OsIQD14 results in short and wide seeds and increases overall yield, overexpression leads to narrow and long seeds, caused by changes in the direction of MT arrangement. We further show that OsIQD14-mediated MT reordering is regulated through interacting with SPIRAL2, a MT-binding protein involved in KATANIN1-mediated MT rearrangement7,8, and with calmodulin proteins. As such, OsIQD14 acts as an integrator of auxin and calcium inputs into MT rearrangements, and allows effective local cell shape manipulation to improve a key rice yield trait.

plant biology↗