REM1.3's phospho-status defines its plasma membrane nanodomain organization and activity in restricting PVX cell-to-cell movement

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

SEC14-like condensate phase transitions at plasma membranes regulate root growth in Arabidopsis

Protein function can be modulated by phase transitions in their material properties, which can range from liquid- to solid-like; yet, the mechanisms that drive these transitions and whether they are important for physiology are still unknown. In the model plant Arabidopsis, we show that developmental robustness is reinforced by phase transitions of the plasma membrane-bound lipid-binding protein SEC14-like. Using imaging, genetics, and in vitro reconstitution experiments, we show that SEC14-like undergoes liquid-like phase separation in the root stem cells. Outside the stem cell niche, SEC14-like associates with the caspase-like protease separase and conserved microtubule motors at unique polar plasma membrane interfaces. In these interfaces, SEC14-like undergoes processing by separase, which promotes its liquid-to-solid transition. This transition is important for root development, as lines expressing an uncleavable SEC14-like variant or mutants of separase and associated microtubule motors show similar developmental phenotypes. Furthermore, the processed and solidified but not the liquid form of SEC14-like interacts with and regulates the polarity of the auxin efflux carrier PINFORMED2. This work demonstrates that robust development can involve liquid-to-solid transitions mediated by proteolysis at unique plasma membrane interfaces.The mechanisms that drive protein phase transitions are unclear. This study in plants shows that an intracellular liquid condensate formed by the lipid transferase SFH8 associates with membranes; when a short fragment of SFH8 is removed by the caspase-like protease ESP, it transforms into a solid filament that can modulate root development

Comparing Arabidopsis receptor kinase and receptor protein-mediated immune signaling reveals BIK1-dependent differences

Pattern recognition receptors (PRRs) sense microbial patterns and activate innate immunity against attempted microbial invasions. The leucine‐rich repeat receptor kinases (LRR‐RK) FLS2 and EFR, and the LRR receptor protein (LRR‐RP) receptors RLP23 and RLP42, respectively, represent prototypical members of these two prominent and closely related PRR families. We conducted a survey of Arabidopsis thaliana immune signaling mediated by these receptors to address the question of commonalities and differences between LRR‐RK and LRR‐RP signaling. Quantitative differences in timing and amplitude were observed for several early immune responses, with RP‐mediated responses typically being slower and more prolonged than those mediated by RKs. Activation of RLP23, but not FLS2, induced the production of camalexin. Transcriptomic analysis revealed that RLP23‐regulated genes represent only a fraction of those genes differentially expressed upon FLS2 activation. Several positive and negative regulators of FLS2‐signaling play similar roles in RLP23 signaling. Intriguingly, the cytoplasmic receptor kinase BIK1, a positive regulator of RK signaling, acts as a negative regulator of RP‐type immune receptors in a manner dependent on BIK1 kinase activity. Our study unveiled unexpected differences in two closely related receptor systems and reports a new negative role of BIK1 in plant immunity

Tyrosine-610 in the receptor kinase BAK1 does not play a major role in brassinosteroid signaling or innate immunity

The plasma membrane-localized BRI1-ASSOCIATED KINASE1 (BAK1) functions as a co-receptor with several receptor kinases including the brassinosteroid (BR) receptor BRASSINOSTEROID-INSENSITIVE 1 (BRI1), which is involved in growth, and the receptors for bacterial flagellin and EF-Tu, FLAGELLIN-SENSING 2 (FLS2) and EF-TU RECEPTOR (EFR), respectively, which are involved in immunity. BAK1 is a dual specificity protein kinase that can autophosphorylate on serine, threonine and tyrosine residues. It was previously reported that phosphorylation of Tyr-610 in the carboxy-terminal domain of BAK1 is required for its function in BR signaling and immunity. However, the functional role of Tyr-610 in vivo has recently come under scrutiny. Therefore, we have generated new BAK1 (Y610F) transgenic plants for functional studies. We first produced transgenic Arabidopsis lines expressing BAK1 (Y610F)-Flag in the homozygous bak1-4 bkk1-1 double null background. In a complementary approach, we expressed untagged BAK1 and BAK1 (Y610F) in the bak1-4 null mutant. Neither BAK1 (Y610F) transgenic line had any obvious growth phenotype when compared to wild-type BAK1 expressed in the same background. In addition, the BAK1 (Y610F)-Flag plants responded similarly to plants expressing BAK1-Flag in terms of brassinolide (BL) inhibition of root elongation, and there were only minor changes in gene expression between the two transgenic lines as monitored by microarray analysis and quantitative real-time PCR. In terms of plant immunity, there were no significant differences between plants expressing BAK1 (Y610F)-Flag and BAK1-Flag in the growth of the non-pathogenic hrpA- mutant of Pseudomonas syringae pv. tomato DC3000. Furthermore, untagged BAK1 (Y610F) transgenic plants were as responsive as plants expressing BAK1 (in the bak1-4 background) and wild-type Col-0 plants toward treatment with the EF-Tu- and flagellin-derived peptide epitopes elf18- and flg22, respectively, as measured by reactive oxygen species production, mitogen-activated protein kinase activation, and seedling growth inhibition. These new results do not support any involvement of Tyr-610 phosphorylation in either BR or immune signaling.This work was supported in part by the National Science Foundation (IOS-1022177 and MCB-1021363) and the United States Department of Agriculture (USDA)-Agricultural Research Service (ARS) (SH), as well as by the Gatsby Charitable Foundation and the European Research Council (grant ‘PHOSPHinnATE’) (CZ)

SEC14-like condensate phase transitions at plasma membranes regulate root growth in Arabidopsis.

Protein function can be modulated by phase transitions in their material properties, which can range from liquid- to solid-like; yet, the mechanisms that drive these transitions and whether they are important for physiology are still unknown. In the model plant Arabidopsis, we show that developmental robustness is reinforced by phase transitions of the plasma membrane-bound lipid-binding protein SEC14-like. Using imaging, genetics, and in vitro reconstitution experiments, we show that SEC14-like undergoes liquid-like phase separation in the root stem cells. Outside the stem cell niche, SEC14-like associates with the caspase-like protease separase and conserved microtubule motors at unique polar plasma membrane interfaces. In these interfaces, SEC14-like undergoes processing by separase, which promotes its liquid-to-solid transition. This transition is important for root development, as lines expressing an uncleavable SEC14-like variant or mutants of separase and associated microtubule motors show similar developmental phenotypes. Furthermore, the processed and solidified but not the liquid form of SEC14-like interacts with and regulates the polarity of the auxin efflux carrier PINFORMED2. This work demonstrates that robust development can involve liquid-to-solid transitions mediated by proteolysis at unique plasma membrane interfaces

Remorin, a plant protein involved in virus movement; implication of the post-translational modifications

Les Rémorines (REM) du groupe 1 sont des protéines spécifiques du monde végétale. Malgré leur caractère hydrophile elles sont localisées à la membrane plasmique. La phosphorylation des REM serait potentiellement impliquée dans la signalisation précoce et la défense des végétaux contre les pathogènes. Benschop et al. (2007) détecte AtREM1.3 (Arabidopsis thaliana, groupe 1b) phosphorylée en réponse au traitement par l'éliciteur générale flg22, tandis que Widjaja et al. (2008) a suggéré que la phosphorylation de AtREM1.2 est potentiellement impliquée dans la signalisation précoce à l'infection par Pseudomonas syringae. La fonction précise de la phosphorylation des protéines REM du groupe 1 reste inconnue. Des travaux antérieurs dans le laboratoire ont montré que le mouvement du virus X de la pomme de terre (PVX) est inversement corrélée à l'accumulation de StREM1.3 (Solanum tuberosum) et que StREM1.3 peut interagir physiquement avec la protéine de mouvement TRIPLE GENE BLOC Protein 1 (TGBp1) du PVX (Raffaele et al., 2009). Dans ce travail, nous avons étudié les mécanismes qui sous-tendent les interactions REM-TGBp1 et nous avons essayé de caractériser biochimiquement la kinase qui phosphoryle REM. Les conséquences physiologiques de l'interaction TGBp1 / StREM1.3 et de la phosphorylation de REM en terme de propagation des virus, d’inactivation génique post-transcriptionnelle, de régulation de l’ouverture des plasmodesmes, et d’activation de kinase ont également été étudiés.The group 1 Remorin (REM) proteins are plant-specific oligomeric proteins that have been reported to localize to the plasma membrane despite their overall hydrophilic nature. There is evidence that the REM protein phosphorylation is potentially implicated in the early signaling and defense. Benschop et al. (2007) detected the AtREM1.3 (Arabidopsis thaliana group 1b of REM protein family) to be phosphorylated in response to treatment with the general elicitor flg22, while the Widjaja et al. (2008) suggested that the phosphorylation of AtREM1.2 is potentially implicated in early signaling upon infection with Pseudomonas syringae. The precise exact function of the group 1 REM protein phosphorylation remains unknown. Previous work in the laboratory showed that Potato virus X (PVX) movement is inversely correlated to potato StREM1.3 accumulation and that StREM1.3 can physically interacts with the movement protein TRIPLE GENE BLOCK PROTEIN 1 (TGBp1) from PVX (Raffaele et al., 2009). In this work, we studied the mechanism underlying the REM-TGBp1 interactions and we tried to characterise biochemically the kinase that phosphorylate REM. The physiological consequences of TGBp1/ StREM1.3 interaction and REM phosphorylation in terms of virus spreading, post-transcriptional gene silencing, plasmodesmata gating, kinase activation were also investigated

Remorin, a plant protein involved in virus movement; implication of the post-translational modifications

Les Rémorines (REM) du groupe 1 sont des protéines spécifiques du monde végétale. Malgré leur caractère hydrophile elles sont localisées à la membrane plasmique. La phosphorylation des REM serait potentiellement impliquée dans la signalisation précoce et la défense des végétaux contre les pathogènes. Benschop et al. (2007) détecte AtREM1.3 (Arabidopsis thaliana, groupe 1b) phosphorylée en réponse au traitement par l'éliciteur générale flg22, tandis que Widjaja et al. (2008) a suggéré que la phosphorylation de AtREM1.2 est potentiellement impliquée dans la signalisation précoce à l'infection par Pseudomonas syringae. La fonction précise de la phosphorylation des protéines REM du groupe 1 reste inconnue. Des travaux antérieurs dans le laboratoire ont montré que le mouvement du virus X de la pomme de terre (PVX) est inversement corrélée à l'accumulation de StREM1.3 (Solanum tuberosum) et que StREM1.3 peut interagir physiquement avec la protéine de mouvement TRIPLE GENE BLOC Protein 1 (TGBp1) du PVX (Raffaele et al., 2009). Dans ce travail, nous avons étudié les mécanismes qui sous-tendent les interactions REM-TGBp1 et nous avons essayé de caractériser biochimiquement la kinase qui phosphoryle REM. Les conséquences physiologiques de l'interaction TGBp1 / StREM1.3 et de la phosphorylation de REM en terme de propagation des virus, d’inactivation génique post-transcriptionnelle, de régulation de l’ouverture des plasmodesmes, et d’activation de kinase ont également été étudiés.The group 1 Remorin (REM) proteins are plant-specific oligomeric proteins that have been reported to localize to the plasma membrane despite their overall hydrophilic nature. There is evidence that the REM protein phosphorylation is potentially implicated in the early signaling and defense. Benschop et al. (2007) detected the AtREM1.3 (Arabidopsis thaliana group 1b of REM protein family) to be phosphorylated in response to treatment with the general elicitor flg22, while the Widjaja et al. (2008) suggested that the phosphorylation of AtREM1.2 is potentially implicated in early signaling upon infection with Pseudomonas syringae. The precise exact function of the group 1 REM protein phosphorylation remains unknown. Previous work in the laboratory showed that Potato virus X (PVX) movement is inversely correlated to potato StREM1.3 accumulation and that StREM1.3 can physically interacts with the movement protein TRIPLE GENE BLOCK PROTEIN 1 (TGBp1) from PVX (Raffaele et al., 2009). In this work, we studied the mechanism underlying the REM-TGBp1 interactions and we tried to characterise biochemically the kinase that phosphorylate REM. The physiological consequences of TGBp1/ StREM1.3 interaction and REM phosphorylation in terms of virus spreading, post-transcriptional gene silencing, plasmodesmata gating, kinase activation were also investigated

An update on plant membrane rafts

International audienceThe dynamic segregation of membrane components within microdomains, such as the sterol-enriched and sphingolipid-enriched membrane rafts, emerges as a central regulatory mechanism governing physiological responses in various organisms. Over the past five years, plasma membrane located raft-like domains have been described in several plant species. The protein and lipid compositions of detergent-insoluble membranes, supposed to contain these domains, have been extensively characterised. Imaging methods have shown that lateral segregation of lipids and proteins exists at the nanoscale level at the plant plasma membrane, correlating detergent insolubility and membrane-domain localisation of presumptive raft proteins. Finally, the dynamic association of specific proteins with detergent-insoluble membranes upon environmental stress has been reported, confirming a possible role for plant rafts as signal transduction platforms, particularly during biotic interactions