Biology subjects

Gronnier, J.

Publications and source records attributed to Gronnier, J..

18 recordsLinked to original sources

bioRxiv2026

Structural and functional insights into the role of Cysteine-Rich Receptor-Like Kinase 18 (CRK18) in Arabidopsis

Plants perceive and integrate diverse environmental signals through receptor kinase (RK) networks at the plasma membrane. Within this, Cysteine-Rich Receptor-Like Kinases (CRKs) constitute a large but not well-understood subfamily characterised by extracellular domains (ECDs) enriched in cysteine residues. CRKs have been implicated in plant responses to biotic and abiotic stress, as well as in developmental processes. Additionally, several CRKs have been proposed to act as redox sensors. Here, we investigate the homodimerization mechanism of Arabidopsis CRK18 and its regulation by redox conditions. By modulating pH and redox state, we assessed the stability and binding dynamics of the CRK18 ECD and its cysteine mutants. We also tested the plasma membrane localisation of all the cysteine mutants involved in the predicted disulfide bonds, and only CRK18C227A, C228A-ECD resembled the plasma membrane localization of wild-type CRK18. We combine co-immunoprecipitation, Forster resonance energy transfer-fluorescence lifetime imaging microscopy and microscale thermophoresis to quantify CRK18 self-association in planta and in vitro. Furthermore, we place CRK18 dimerization in a broader signalling context by identifying CRK18 interaction partners and CRK18-dependent signalling outputs. Using Arabidopsis CRK18 overexpression lines, we perform (phospho)proteomic and immunoprecipitation-mass spectrometry (IP-MS) analyses to map CRK18-centred signalling networks associated with stress-related responses. The constitutive activation of the CRK18 kinase domain and its interaction with many putative (cell wall) glycan-sensing RKs, including PERK15, suggest a regulatory role for CRK18. The absence of significant changes in the proteome and phosphoproteome of CRK18 overexpression lines in the absence of any trigger, and its restricted mobility after elicitation, suggest that CRK18 requires a stimulus for activation and possibly induces membrane microdomain reorganisation. This is in line with the infection assay with the nematode Heterodera schachtii, which causes modification and targeted damage to plant cell walls during infection, revealing that CRK18 acts as negative regulator of this process.

biochemistry↗

bioRxiv2026

A bacterial lipid triggers membrane mechanosensing immunity in Arabidopsis

The plant immune system engages cell-surface receptors that detect microbe-associated molecular patterns to initiate pattern-triggered immunity (PTI), and intracellular receptors that sense microbe-secreted effectors to activate effector-triggered immunity (ETI). Whether additional modes of microbial detection exist remains unclear. Here, we define membrane mechanosensing immunity (MSI), a third layer of immune signaling. A bacterial lipid, the main diffusible signal factor (DSF) from Xanthomonas campestris pv. campestris, acts as a membrane-active molecule that alters plasma membrane biophysical properties, activates Mechanosensitive Channel of Small Conductance (MscS)-like (MSL)-dependent immune signaling, and triggers a broad transcriptional reprogramming that overlaps with PTI and ETI. MSI modulates PTI signaling and requires both PTI and ETI components for effective disease resistance. These findings establish the sensing of metabolite-induced membrane perturbations as a mechanism of microbial detection.

plant biology↗

bioRxiv2026

Plasma membrane nanoscale dynamics of Arabidopsis leucine-rich repeat receptor kinase complexes

Plasma membrane-localized receptors operate as dynamic signaling complexes and integrative networks1-3, yet the spatial and temporal regulation of these interactions remain largely unknown. Here, by analyzing the components of a minimal Arabidopsis leucine-rich repeat receptor kinase network, we describe the differential diffusion and organization of receptor complex components and unveil the nanoscale spatial and temporal logic underlying the formation of receptor kinase complexes. The ligand-binding receptors FLS2 and BRI1, and the accessory receptor BIR3, are organized in plasma membrane nanodomains, within which the co-receptor BAK1 diffuses and is spatially arrested upon ligand perception. BAK1s spatial arrest relies on extracellular domain (ECD)-ECD interactions but does not require receptor complex activation. Mathematical modelling, single molecule imaging and bio-assays infer that accessory receptors maintain a dynamic pool of co-receptors in the vicinity of ligand-binding receptors to promote ligand-induced complex formation and signaling. We propose that ligand-induced receptor kinase complex formation is a deterministic process defined by the relative nanoscale spatial positioning of individual signaling and regulatory components.

cell biology↗

bioRxiv2025

Membrane-binding domains define REMORIN phylogeny and provide a predicted structural basis for distinctive membrane nano-environments

REMORIN (REM) proteins are structural components of the plant plasma membrane that modulate membrane nano-organization and biophysics. They have been proposed to function as versatile scaffolds in the context of hormone signaling, immunity and symbiosis. REMs have been classified into six groups based the length and the amino acid composition of their intrinsically disordered N-terminal domain. Here we show that REM phylogeny is dominated by the evolution of their conserved C-terminal domain and defines four major REM clades. Structural bioinformatics analyses predict the conservation of a putative membrane binding interface formed by REM C-terminal domains and reveal a striking diversity in their curvatures and lengths. A subset of REMs is predicted to form C-terminal domain-mediated higher-order oligomers providing an additional level of diversity in REM membrane-binding interfaces. We discuss the implications of the predicted variations in REM C-terminal domain structure for their molecular function and membrane organization.

plant biology↗

bioRxiv2025

Manipulation of REMORIN gene dosage affects salt signaling and tolerance in Arabidopsis thaliana

Salinity is among the most detrimental environmental plant stressors hampering growth, development and immunity. Its impact is increasing due to intensive agricultural practices and climate change. The molecular components linked to the early cellular and molecular events triggered by salt are mostly unknown. Here we show that genes encoding for members of the REMORIN proteins family exhibit dynamic temporal regulation following salt perception. Analogously to pattern-triggered immune signaling, we observed that salt rapidly induces the reorganization of REMORIN1.2 (REM1.2) into static plasma membrane nanodomains which colocalize with the actin-nucleating protein FORMIN 6. Overexpression of REM1.2 inhibits early signaling and late cell morphological responses to salt, defects which correlated with an increase in salt sensitivity. Our findings connect early salt-triggered signaling events to changes in cell architecture and root resilience and describe REMORIN nanodomains as a convergent articulation point in biotic and abiotic signaling pathways.

plant biology↗

bioRxiv2025

An ancient alkalinization factor informs Arabidopsis root development

The power of hydrogen (pH) regulates virtually all cellular activities. In both plants and animals, cell-to-cell variations in pH correlate with key developmental transitions1-5, yet the underlying regulators and associated functions remain elusive. Here, we report that members of the REMORIN (REM) protein family function as inhibitors of the H+-ATPases thereby promoting extracellular pH (pHe) alkalinization. This, in turn, regulates various cell surface processes, including steroid hormone signaling, and coordinates developmental transitions in the Arabidopsis thaliana root. Inhibition of H+-ATPases by REMs represents an evolutionary innovation that predates the origin of the root system itself. This study thus uncovers an ancient alkalinization mechanism co-opted by the root developmental program and infers that pHe patterning may have shaped morphogenesis evolution.

plant biology↗

bioRxiv2025

Salicylic acid-induced alkalinization of the apoplast requires TRANSMEMBRANE KINASE 1 and results in growth attenuation

The phytohormone salicylic acid (SA) has a key role in regulating plant growth and stress response. In the past, most of the growth-related SA functions have been explained by crosstalk with the master growth regulator auxin. By affecting polarity of auxin transporters, SA changes auxin distribution in the root and inhibits growth of the main root and activates lateral root formation. However, only recently there is evidence emerging that SA impacts growth processes independently of nuclear auxin signalling, possessing yet unknown mechanistic functions. Here we show that SA activity depends on TRANSMEMBRANE KINASE 1 (TMK1) resulting in apoplast alkalinization and growth restriction. SA treatment prevents phosphorylation and activation of plasma membrane (PM) H+-ATPases at the cell surface and does not depend on the auxin receptor Auxin Binding Protein 1 (ABP1). We suggest that alkalinization of the apoplast by SA serves as mechanism to balance stress response and growth.

plant biology↗

bioRxiv2025

Cell wall integrity and elicitor peptide signaling modulate antimicrobial defense in Arabidopsis via jasmonic acid

Plant cell walls constitute dynamic barriers that are essential for defense against pathogens. The receptor kinase THESEUS1 (THE1) monitors cell wall integrity (CWI) and contributes to pathogen resistance in Arabidopsis, but the underlying mechanisms remain unclear. Here we show that THE1-dependent CWI signaling induces accumulation of the antimicrobial metabolite camalexin upon cell wall damage (CWD) caused by cellulose biosynthesis inhibition or fungal infection. CWD alters THE1 plasma membrane nanodomain organization and involves calcium signaling components that modulate camalexin production. Induction of camalexin requires jasmonic acid (JA)-dependent expression of the transcription factors MYB47 and MYB95. In line with its antagonistic function on CWI signaling, the plant elicitor peptide Pep3 suppresses camalexin biosynthesis downstream of THE1 by inhibiting JA-dependent pathways. Our findings reveal a regulatory network where CWI and Pep3 signaling modulate antimicrobial defense via JA-mediated camalexin production. This network requires independent CWD-induced pathways, providing insights into how plants balance defense activation and suppression in response to cell wall stress.

plant biology↗

bioRxiv2025

Arabidopsis HYPERSENSITIVE INDUCED REACTION 2 affects plasma membrane receptor pathways and organization

The distribution of proteins across the plasma membrane is not uniform; however, the principles governing their organization remain not fully understood. Hypersensitive-induced reaction (HIR) proteins are plant-specific members of the stomatin/prohibitin/flotillin/HflK/C (SPFH) family that have been shown to influence membrane organization. Arabidopsis thaliana HIR2 interacts with multiple plasma membrane proteins, including receptor kinases such as BAK1-INTERACTING RECEPTORS 2 and 3 (BIR2 and 3), BRI1-ASSOCIATED KINASE 1 (BAK1), FLAGELLIN SENSING 2 (FLS2), and BRASSINOSTEROID INSENSITIVE 1 (BRI1). These interactions connect HIR2 to BAK1-mediated signaling pathways, as evidenced by impaired growth and immunity phenotypes in hir2 mutants. HIR2 is anchored to the inner leaflet of the plasma membrane through a hydrophobic interaction domain and S-acylation. Using single-particle tracking photoactivated localization microscopy (sptPALM), we showed that HIR2 affects receptor kinase dynamics and clustering, suggesting a role in spatially coordinating receptor complex activities. Structural modeling with AlphaFold 3 predicts a multimeric circular cup-like assembly for HIR2, consistent with high molecular weight complexes identified through blue native polyacrylamide gel electrophoresis. These findings indicate that HIR2 forms a discrete membrane compartment, providing a novel structural framework for spatial membrane organization and thereby modulating the function of membrane-resident receptors.

plant biology↗

bioRxiv2024

Photochromic reversion enables long-term tracking of single molecules in living plants.

Single-molecule imaging enables the observation of individual molecules in living cells (DEste et al., 2024; Kusumi et al., 2014; Lelek et al., 2021; Nguyen et al., 2023). In plants, however, the tracking of single molecules is typically limited to a few hundred milliseconds (Bayle et al., 2021; Gronnier et al., 2017; Hosy et al., 2015), precluding the observation of dynamic cellular processes at molecular resolution. Here, we describe photochromic reversion, an imaging modality that enables long-term single-molecule tracking of genetically encoded translational fusions. Using this approach, we achieve minute-long tracking of individual cell-surface receptors and reveal previously inaccessible dynamic spatial arrest events of single plasma membrane proteins. We further developed and benchmarked computational analysis of spatial arrests (CASTA), a machine learning-based tool that automatically detects and analyses spatial, temporal, and diffusional properties of these events, thereby enabling precise nanoscale kinetic measurements. Together, these advances provide a powerful framework for deciphering the principles governing membrane dynamics and function.

cell biology↗

bioRxiv2024

A RALF-Brassinosteroid morpho-signaling circuit regulates Arabidopsis hypocotyl cell shape

Plant cells survey and modulate their cell wall to control their shape and anisotropic growth. Signaling mediated by the plant steroid hormones brassinosteroids (BR) plays a central role in coordinating cell wall status and cell growth, and alterations in the cell wall - BR feedback loop leads to life-threatening defects in tissue and cellular integrity. How the status of the cell wall is relayed to BR signaling remains largely unclear. Increasing evidence shows that RAPID ALKALANIZATION FACTORs (RALFs), a class of secreted peptides, play structural and signaling roles at the cell surface. Here we show that perception of RALF23 promotes the formation and signaling of the main BR receptor complex formed by BRASSINOSTEROID INSENSITIVE 1 (BRI1) and BRI1-BRASSINOSTEROID INSENSITIVE1-BRASSINOSTEROID-ASSOCIATED KINASE 1 (BAK1). The loss of the plasma membrane-localized RALF receptor complex FERONIA (FER)-LORELEI LIKE GPI-anchor protein 1 (LLG1) leads to defects in cell expansion and anisotropy, as well as uncontrolled BRI1-BAK1 complex formation and signaling. RALF23 bioactivity relies on pectin status and its perception induces changes in pectin composition and the activity of pectin-modifying enzymes. Our observations suggest a model in which RALF23 functions as a cell wall-informed signaling cue initiating a feedback loop that solicits BR signaling, modifies the cell wall, and coordinates cell morphogenesis. Highlights-The RALF receptor complex FER-LLG1 regulates cell anisotropic growth -RALF23 promotes BRI1-BAK1 complex formation and signaling -RALF23 functions as a cell wall-informed and wall-modifying signaling cue

plant biology↗

bioRxiv2023

Interdependence of a kinase and its cognate substrate plasma membrane nanoscale dynamics underlies Arabidopsis response to viral infection

Plant viruses represent a risk to agricultural production and as only a few treatments exist, it is urgent to identify resistance mechanisms and factors. In plant immunity, plasma membrane (PM)-localized proteins play an essential role in sensing the extracellular threat presented by bacteria, fungi or herbivores. Viruses are intracellular pathogens and as such the role of the plant PM in detection and resistance against viruses is often overlooked. We investigated the role of the partially PM-bound Calcium-dependent protein kinase 3 (CPK3) in viral infection and we discovered that it displayed a specific ability to hamper viral propagation over CPK isoforms that are involved in immune response to extracellular pathogens. More and more evidence support that the lateral organization of PM proteins and lipids underlies signal transduction in plants. We showed here that CPK3 diffusion in the PM is reduced upon activation as well as upon viral infection and that such immobilization depended on its substrate, Remorin (REM1.2), a scaffold protein. Furthermore, we discovered that the viral infection induced a CPK3-dependent increase of REM1.2 PM diffusion. Such interdependence was also observable regarding viral propagation. This study unveils a complex relationship between a kinase and its substrate that contrasts with the commonly described co-stabilisation upon activation while it proposes a PM-based mechanism involved in decreased sensitivity to viral infection in plants.

plant biology↗

bioRxiv2023

The actin depolymerizing factor StADF2 alters StREM1.3 plasma membrane nanodomains to inhibit the Potato Virus X

The dynamic regulation of the plasma membrane (PM) organization at the nanoscale emerged as a key element shaping the outcome of host-microbe interactions. Protein organization into nanodomains (ND) is often assumed to be linked to the activation of cellular processes. In contrast, we have previously shown that the phosphorylation of the Solanum tuberosum REM1.3 (StREM1.3) N-terminal domain disperses its native ND organization and promotes its inhibitory effect on Potato Virus X (PVX) cell-to-cell movement. Here, we show that the phosphorylation of StREM1.3 modify the chemical environment of numerous residues in its intrinsically-disordered N-terminal domain. We leveraged exploratory screens to identify potential phosphorylation-dependent interactors of StREM1.3. Herewith, we uncovered uncharacterized regulators of PVX cell-to-cell movement, linking StREM1.3 to autophagy, water channels and the actin cytoskeleton. We show that the Solanum tuberosum actin depolymerizing factors 2 (StADF2) alters StREM1.3 NDs and limits PVX cell-to-cell movement in a REMORIN-dependent manner. Mutating a conserved single residue reported to affect ADFs affinity to actin inhibits StADF2 effect on StREM1.3 ND organization and PVX cell-to-cell movement. These observations provide functional links between the organization of plant PM and the actin cytoskeleton and suggests that the alteration of StREM1.3 ND organization promotes plant anti-viral responses. We envision that analogous PM re-organization applies for additional signaling pathways in plants and in other organisms.

cell biology↗

bioRxiv2021

S-acylation is a positive regulator of FLS2-mediated plant immunity

Plant receptor kinases are key transducers of extracellular stimuli, such as the presence of beneficial or pathogenic microbes or secreted signalling molecules. Receptor kinases are regulated by numerous post-translational modifications. Here, using the immune receptor kinases FLS2 and EFR, we show that S-acylation at a cysteine conserved in all plant receptor kinases is crucial for function. S-acylation involves the addition of long-chain fatty acids to cysteine residues within proteins, altering their biophysical properties and behaviour within the membrane environment. We observe S-acylation of FLS2 at C-terminal kinase domain cysteine residues within minutes following perception of its ligand flg22, in a BAK1 co-receptor dependent manner. We demonstrate that S-acylation is essential for FLS2-mediated immune signalling and resistance to bacterial infection. Similarly, mutating the corresponding conserved cysteine residue in EFR supressed elf18 triggered signalling. Analysis of unstimulated and activated FLS2-containing complexes using microscopy, detergents and native membrane DIBMA nanodiscs indicates that S-acylation stabilises and promotes retention of activated receptor kinase complexes at the plasma membrane to increase signalling efficiency.

plant biology↗

bioRxiv2021

A hetero-oligomeric remorin-receptor complex regulates plant development

Plant growth and development are modulated by both biotic and abiotic stress. Increasing evidence suggests that cellular integration of the corresponding signals occurs within preformed hubs at the plasma membrane called nanodomains. These membrane sub-compartments are organized by multivalent molecular scaffold proteins, such as remorins. Here, we demonstrate that Group 1 remorins form a hetero-oligomeric complex at the plasma membrane. While these remorins are functionally redundant for some pathways their multivalency also allows the recruitment of specific interaction partners. One of them, the receptor-like kinase REMORIN-INTERACTING RECEPTOR 1 (RIR1), that acts redundantly with the closely related receptor NILR2, is specifically recruited by REM1.2 in a phosphorylation-dependent manner. Overlapping developmental phenotypes suggest that the REM/RIR complex regulates key developmental pathways.

plant biology↗

bioRxiv2020

FERONIA regulates FLS2 plasma membrane nanoscale dynamics to modulate plant immune signaling

Spatial partitioning is a propensity of biological systems orchestrating cell activities in space and time. The dynamic regulation of plasma membrane nano-environments has recently emerged as a key fundamental aspect of plant signaling, but the molecular components governing it are still mostly unclear. The receptor kinase FERONIA (FER) controls complex formation of the immune receptor kinase FLAGELLIN SENSING 2 (FLS2) with its co-receptor BRASSINOSTEROID INSENSITIVE 1-ASSOCIATED KINASE 1 (BAK1), and this function is inhibited by the FER ligand RAPID ALKALANIZATION FACTOR 23 (RALF23). Here, we show that FER regulates the plasma membrane nanoscale organization of FLS2 and BAK1. Our study demonstrates that akin to FER, leucine-rich repeat (LRR) extensin (LRXs) proteins contribute to RALF23 responsiveness, regulate BAK1 nanoscale organization and immune signaling. Furthermore, RALF23 perception leads to rapid modulation of FLS2 and BAK1 nanoscale organization and its inhibitory activity on immune signaling relies on FER kinase activity. Our results suggest that perception of RALF peptides by FER and LRXs actively modulates the plasma membrane nanoscale landscape to regulate cell surface signaling by other receptor kinases.

plant biology↗

bioRxiv2020

Plastidic Δ6 Fatty-Acid Desaturases With Distinctive Substrate Specificity Regulate The Pool Of C18-PUFAs In The Ancestral Picoalga Ostreococcus tauri

Eukaryotic {Delta}6-desaturases are microsomal enzymes which balance the synthesis of {omega}-3 and {omega}-6 C18-polyunsaturated-fatty-acids (PUFA) accordingly to their specificity. In several microalgae, including O. tauri, plastidic C18-PUFA are specifically regulated by environmental cues suggesting an autonomous control of {Delta}6-desaturation of plastidic PUFA. Sequence retrieval from O. tauri desaturases, highlighted two putative {Delta}6/{Delta}8-desaturases sequences clustering, with other microalgal homologs, apart from other characterized {Delta}-6 desaturases. Their overexpression in heterologous hosts, including N. benthamiana and Synechocystis, unveiled their {Delta}6-desaturation activity and plastid localization. O. tauri lines overexpressing these {Delta}6-desaturases no longer adjusted their plastidic C18-PUFA amount under phosphate starvation but didnt show any obvious physiological alterations. Detailed lipid analyses from the various overexpressing hosts, unravelled that the substrate features involved in the {Delta}6-desaturase specificity importantly involved the lipid head-group and likely the non-substrate acyl-chain, in addition to the overall preference for the {omega}-class of the substrate acyl-chain. The most active desaturase displayed a broad range substrate specificity for plastidic lipids and a preference for {omega}-3 substrates, while the other was selective for {omega}-6 substrates, phosphatidylglycerol and 16:4-galactolipid species specific to the native host. The distribution of plastidial {Delta}6-desaturase products in eukaryotic hosts suggested the occurrence of C18-PUFA export from the plastid. One sentence summaryOsteococcus tauri plastidic lipid C18-PUFA remodelling involves two plastid-located cytochrome-b5 fused {Delta}6-desaturases with distinct preferences for both head-group and acyl-chain.

biochemistry↗

bioRxiv2017

The plant calcium-dependent protein kinase CPK3 phosphorylates REM1.3 to restrict viral infection

Plants respond to pathogens through dynamic regulation of plasma membrane-bound signaling pathways. To date, how the plant plasma membrane is involved in responses to viruses is mostly unknown. Here, we show that plant cells sense the Potato virus X (PVX) COAT PROTEIN and TRIPLE GENE BLOCK 1 proteins and subsequently trigger the activation of a membrane-bound calcium-dependent kinase. We show that the Arabidopsis thaliana CALCIUM-DEPENDENT PROTEIN KINASE 3-interacts with group 1 REMORINs in vivo, phosphorylates the intrinsically disordered N-terminal domain of the Group 1 REMORIN REM1.3, and restricts PVX cell-to-cell movement. REM1.3-s phospho-status defines its plasma membrane nanodomain organization and is crucial for REM1.3-dependent restriction of PVX cell-to-cell movement by regulation of callose deposition at plasmodesmata. This study unveils plasma membrane nanodomain-associated molecular events underlying the plant immune response to viruses.

plant biology↗