ABSTRACT
Plants spend most of their life oscillating around 1-3 Hz due to the effect of the wind. Therefore, stems and foliage experience repetitive mechanical stresses through these passive movements. However, the mechanism of the cellular perception and transduction of such recurring mechanical signals remains an open question. Multimeric protein complexes forming mechanosensitive (MS) channels embedded in the membrane provide an efficient system to rapidly convert mechanical tension into an electrical signal. So far, studies have mostly focused on nonoscillatory stretching of these channels. Here, we show that the plasma-membrane MS channel MscS-LIKE 10 (MSL10) from the model plant Arabidopsis thaliana responds to pulsed membrane stretching with rapid activation and relaxation kinetics in the range of 1 s. Under sinusoidal membrane stretching MSL10 presents a greater activity than under static stimulation. We observed this amplification mostly in the range of 0.3-3 Hz. Above these frequencies the channel activity is very close to that under static conditions. With a localization in aerial organs naturally submitted to wind-driven oscillations, our results suggest that the MS channel MSL10, and by extension MS channels sharing similar properties, represents a molecular component allowing the perception of oscillatory mechanical stimulations by plants.
Subject(s)
Arabidopsis Proteins/metabolism , Cell Membrane/metabolism , Mechanotransduction, Cellular/physiology , Membrane Proteins/metabolism , Arabidopsis/metabolism , Arabidopsis Proteins/physiology , Cell Membrane/physiology , Ion Channels/metabolism , Ion Transport , Mechanoreceptors/metabolism , Membrane Proteins/physiology , Plants, Genetically Modified/metabolism , Signal TransductionABSTRACT
A fundamental function of an organ is the ability to perceive mechanical cues. Yet, how this is accomplished is not fully understood, particularly in plant roots. In plants, the majority of studies dealing with the effects of mechanical stress have investigated the aerial parts. However, in natural conditions roots are also subjected to mechanical cues, for example when the root encounters a hard obstacle during its growth or when the soil settles. To investigate root cellular responses to root compression, we developed a microfluidic system associated with a microvalve allowing the delivery of controlled and reproducible mechanical stimulations to the root. In this study, examining plants expressing the R-GECO1-mTurquoise calcium reporter, we addressed the root cell deformation and calcium increase induced by the mechanical stimulation. Lateral pressure applied on the root induced a moderate elastic deformation of root cortical cells and elicited a multicomponent calcium signal at the onset of the pressure pulse, followed by a second one at the release of the pressure. This indicates that straining rather than stressing of tissues is relevant to trigger the calcium signal. Although the intensity of the calcium response increases with the pressure applied, successive pressure stimuli led to a remarkable attenuation of the calcium signal. The calcium elevation was restricted to the tissue under pressure and did not propagate. Strain sensing, spatial restriction and habituation to repetitive stimulation represent the fundamental properties of root signalling in response to local mechanical stimulation. These data linking mechanical properties of root cells to calcium elevation contribute to elucidating the pathway allowing the root to adapt to the mechanical cues generated by the soil.
Subject(s)
Arabidopsis , Calcium/metabolism , Signal Transduction/physiology , Soil , Plant RootsABSTRACT
Plant and animals have evolved different strategies for their development. Whether this is linked to major differences in their cell mechanics remains unclear, mainly because measurements on plant and animal cells relied on independent experiments and setups, thus hindering any direct comparison. In this study we used the same micro-rheometer to compare animal and plant single cell rheology. We found that wall-less plant cells exhibit the same weak power law rheology as animal cells, with comparable values of elastic and loss moduli. Remarkably, microtubules primarily contributed to the rheological behavior of wall-less plant cells whereas rheology of animal cells was mainly dependent on the actin network. Thus, plant and animal cells evolved different molecular strategies to reach a comparable cytoplasmic mechanical core, suggesting that evolutionary convergence could include the internal biophysical properties of cells.
Subject(s)
Arabidopsis/cytology , Mechanical Phenomena , Animals , Biomechanical Phenomena , Cell Line , Mice , Microtubules/metabolism , Single-Cell Analysis , Species SpecificityABSTRACT
The study of mechanosensitive channels (MS) in living organisms has progressed considerably over the past two decades. The understanding of their roles in mechanosensation and mechanotransduction was consecrated by the awarding of the Nobel Prize in 2021 to A. Patapoutian for his discoveries on the role of MS channels in mechanoperception in humans. In this review, we first summarize the fundamental properties of MS channels and their mode of operation. Then in a second step, we provide an update on the knowledge on the families of MS channels identified in plants and the roles and functions that have been attributed to them.
Subject(s)
Ion Channels , Mechanotransduction, Cellular , Humans , Ion Channels/metabolism , Mechanotransduction, Cellular/physiology , Plants/metabolismABSTRACT
In bacterial and animal systems, mechanosensitive (MS) ion channels are thought to mediate the perception of pressure, touch, and sound [1-3]. Although plants respond to a wide variety of mechanical stimuli, and although many mechanosensitive channel activities have been characterized in plant membranes by the patch-clamp method, the molecular nature of mechanoperception in plant systems has remained elusive [4]. Likely candidates are relatives of MscS (Mechanosensitive channel of small conductance), a well-characterized MS channel that serves to protect E. coli from osmotic shock [5]. Ten MscS-Like (MSL) proteins are found in the genome of the model flowering plant Arabidopsis thaliana[4, 6, 7]. MSL2 and MSL3, along with MSC1, a MscS family member from green algae, are implicated in the control of organelle morphology [8, 9]. Here, we characterize MSL9 and MSL10, two MSL proteins found in the plasma membrane of root cells. We use a combined genetic and electrophysiological approach to show that MSL9 and MSL10, along with three other members of the MSL family, are required for MS channel activities detected in protoplasts derived from root cells. This is the first molecular identification and characterization of MS channels in plant membranes.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Mechanotransduction, Cellular , Membrane Proteins/metabolism , Plant Roots/metabolism , Amino Acid Sequence , Cell Membrane/metabolism , Molecular Sequence Data , Protoplasts/metabolismABSTRACT
During development, tissues are submitted to high variation of compression and tension forces. The roles of the cell wall, the cytoskeleton, the turgor pressure and the cell geometry during this process have received due attention. In contrast, apart from its role in the establishment of turgor pressure, the involvement of the plasma membrane as a transducer of mechanical forces during development has been under studied. Force-gated (FG) or Mechanosensitive (MS) ion channels embedded in the bilayer represent 'per se' archetypal mechanosensor able to directly and instantaneously transduce membrane forces into electrical and calcium signals. We discuss here how their fine-tuning, combined with their ability to detect micro-curvature and local membrane tension, allows FG channels to transduce mechanical cues into developmental signals.
Subject(s)
Calcium , Ion Channels , Cell Membrane , Cytoskeleton , Mechanotransduction, CellularABSTRACT
BACKGROUND: The root is an important organ for water and nutrient uptake, and soil anchorage. It is equipped with root hairs (RHs) which are elongated structures increasing the exchange surface with the soil. RHs are also studied as a model for plant cellular development, as they represent a single cell with specific and highly regulated polarized elongation. For these reasons, it is useful to be able to accurately quantify RH length employing standardized procedures. Methods commonly employed rely on manual steps and are therefore time consuming and prone to errors, restricting analysis to a short segment of the root tip. Few partially automated methods have been reported to increase measurement efficiency. However, none of the reported methods allow an accurate and standardized definition of the position along the root for RH length measurement, making data comparison difficult. RESULTS: We developed an image analysis algorithm that semi-automatically detects RHs and measures their length along the whole differentiation zone of roots. This method, implemented as a simple automated script in ImageJ/Fiji software that we termed Root Hair Sizer, slides a rectangular window along a binarized and straightened image of root tips to estimate the maximal RH length in a given measuring interval. This measure is not affected by heavily bent RHs and any bald spots. RH length data along the root are then modelled with a sigmoidal curve, generating several biologically significant parameters such as RH length, positioning of the root differentiation zone and, under certain conditions, RH growth rate. CONCLUSIONS: Image analysis with Root Hair Sizer and subsequent sigmoidal modelling of RH length data provide a simple and efficient way to characterize RH growth in different conditions, equally suitable to small and large scale phenotyping experiments.
ABSTRACT
Mechanosensitive (MS) channels behave as microprobes that transduce mechanical tension into electric and ion signals. The plasma membrane anion-permeable channel AtMSL10 belongs to the first family of MS channels (MscS-LIKE) that has been characterized in Arabidopsis thaliana. In the same membrane, a rapidly activated calcium MS channel activity (RMA) associated with the presence of the DEFECTIVE KERNEL1 (AtDEK1) protein has been recently described. In this Review, based on the comparison of the specific properties of AtMSL10 and RMA, we put forward hypotheses on the mechanism of activation of these two channels, their respective roles in signalling and also raise the question of the molecular identity of RMA. Finally, we propose functions for these two channels within the context of plant mechanotransduction.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/metabolism , Calpain/metabolism , Membrane Proteins/metabolism , Arabidopsis Proteins/chemistry , Calcium/metabolism , Calpain/chemistry , Cytosol/metabolism , Mechanotransduction, Cellular , Membrane Proteins/chemistry , Protein DomainsABSTRACT
Anion channels/transporters appear as key players in signaling pathways leading to the adaptation of plant cells to abiotic and biotic environmental stresses, in the control of metabolism and in the maintenance of electrochemical gradients. Focusing on the most recent advances, this review aims at providing a description of the role of these channels in various physiological functions such as control of stomatal movements, plant-pathogen interaction, xylem loading, compartmentalization of metabolites and coupling with proton gradients. These functions have been demonstrated by a combination of electrophysiology, pharmacology and genetics approaches, the key issue being to identify the corresponding proteins and genes.
Subject(s)
Anion Transport Proteins/metabolism , Ion Channels/metabolism , Plants/metabolism , Anions/metabolism , Cell Membrane/metabolism , Malates/metabolism , Models, Biological , Nitrate Transporters , Plant Cells , Proton Pumps/metabolism , Signal Transduction , Xylem/metabolismABSTRACT
Responses of cells to mechanical stress are thought to be critical in coordinating growth and development. Consistent with this idea, mechanically activated channels play important roles in animal development. For example, the PIEZO1 channel controls cell division and epithelial-layer integrity and is necessary for vascular development in mammals. In plants, the actual contribution of mechanoperception to development remains questionable because very few putative mechanosensors have been identified and the phenotypes of the corresponding mutants are rather mild. Here, we show that the Arabidopsis Defective Kernel 1 (DEK1) protein, which is essential for development beyond early embryogenesis, is associated with a mechanically activated Ca2+ current in planta, suggesting that perception of mechanical stress plays a critical role in plant development.
Subject(s)
Arabidopsis Proteins/metabolism , Arabidopsis/chemistry , Arabidopsis/growth & development , Calcium/metabolism , Calpain/metabolism , Arabidopsis/genetics , Arabidopsis/metabolism , Arabidopsis Proteins/genetics , Calpain/genetics , Phenotype , Stress, MechanicalABSTRACT
Root hairs are involved in water and nutrient uptake, and thereby in plant autotrophy. In legumes, they also play a crucial role in establishment of rhizobial symbiosis. To obtain a holistic view of Medicago truncatula genes expressed in root hairs and of their regulation during the first hours of the engagement in rhizobial symbiotic interaction, a high throughput RNA sequencing on isolated root hairs from roots challenged or not with lipochitooligosaccharides Nod factors (NF) for 4 or 20 h was carried out. This provided a repertoire of genes displaying expression in root hairs, responding or not to NF, and specific or not to legumes. In analyzing the transcriptome dataset, special attention was paid to pumps, transporters, or channels active at the plasma membrane, to other proteins likely to play a role in nutrient ion uptake, NF electrical and calcium signaling, control of the redox status or the dynamic reprogramming of root hair transcriptome induced by NF treatment, and to the identification of papilionoid legume-specific genes expressed in root hairs. About 10% of the root hair expressed genes were significantly up- or down-regulated by NF treatment, suggesting their involvement in remodeling plant functions to allow establishment of the symbiotic relationship. For instance, NF-induced changes in expression of genes encoding plasma membrane transport systems or disease response proteins indicate that root hairs reduce their involvement in nutrient ion absorption and adapt their immune system in order to engage in the symbiotic interaction. It also appears that the redox status of root hair cells is tuned in response to NF perception. In addition, 1176 genes that could be considered as "papilionoid legume-specific" were identified in the M. truncatula root hair transcriptome, from which 141 were found to possess an ortholog in every of the six legume genomes that we considered, suggesting their involvement in essential functions specific to legumes. This transcriptome provides a valuable resource to investigate root hair biology in legumes and the roles that these cells play in rhizobial symbiosis establishment. These results could also contribute to the long-term objective of transferring this symbiotic capacity to non-legume plants.
ABSTRACT
Plants, like other organisms, are facing multiple mechanical constraints generated both in their tissues and by the surrounding environments. They need to sense and adapt to these forces throughout their lifetimes. To do so, different mechanisms devoted to force transduction have emerged. Here we focus on fascinating proteins: the mechanosensitive (MS) channels. Mechanosensing in plants has been described for centuries but the molecular identification of MS channels occurred only recently. This review is aimed at plant biologists and plant biomechanists who want to be introduced to MS channel identity, how they work and what they might do in planta? In this review, electrophysiological properties, regulations, and functions of well-characterized MS channels belonging to bacteria and animals are compared with those of plants. Common and specific properties are discussed. We deduce which tools and concepts from animal and bacterial fields could be helpful for improving our understanding of plant mechanotransduction. MS channels embedded in their plasma membrane are sandwiched between the cell wall and the cytoskeleton. The consequences of this peculiar situation are analyzed and discussed. We also stress how important it is to probe mechanical forces at cellular and subcellular levels in planta in order to reveal the intimate relationship linking the membrane with MS channel activity. Finally we will propose new tracks to help to reveal their physiological functions at tissue and plant levels.
ABSTRACT
Anion channels/transporters are key to a wide spectrum of physiological functions in plants, such as osmoregulation, cell signaling, plant nutrition and compartmentalization of metabolites, and metal tolerance. The recent identification of gene families encoding some of these transport systems opened the way for gene expression studies, structure-function analyses of the corresponding proteins, and functional genomics approaches toward further understanding of their integrated roles in planta. This review, based on a few selected examples, illustrates that the members of a given gene family exhibit a diversity of substrate specificity, regulation, and intracellular localization, and are involved in a wide range of physiological functions. It also shows that post-translational modifications of transport proteins play a key role in the regulation of anion transport activity. Key questions arising from the increasing complexity of networks controlling anion transport in plant cells (the existence of redundancy, cross talk, and coordination between various pathways and compartments) are also addressed.
Subject(s)
Antiporters/physiology , Arabidopsis Proteins/physiology , Arabidopsis/metabolism , Antiporters/genetics , Antiporters/metabolism , Arabidopsis/genetics , Arabidopsis Proteins/genetics , Arabidopsis Proteins/metabolism , Biological Transport/physiology , Cell Membrane/chemistry , Cell Membrane/metabolism , Gene Expression Regulation, Plant , Models, Biological , Multigene Family/physiology , Nitrates/metabolism , Phosphorylation , Protein Processing, Post-Translational , Substrate SpecificityABSTRACT
Plant genomes code for channels involved in the transport of cations, anions and uncharged molecules through membranes. Although the molecular identity of channels for cations and uncharged molecules has progressed rapidly in the recent years, the molecular identity of anion channels has lagged behind. Electrophysiological studies have identified S-type (slow) and R-type (rapid) anion channels. In this brief review, we summarize the proposed functions of the R-type anion channels which, like the S-type, were first characterized by electrophysiology over 20 years ago, but unlike the S-type, have still yet to be cloned. We show that the R-type channel can play multiple roles.
Subject(s)
Calcium Channels, R-Type/physiology , Gene Expression Regulation, Plant/physiology , Plant Proteins/physiology , Plants/metabolism , Calcium Signaling , Ion Channel Gating/physiologyABSTRACT
Plants need nitrate for growth and store the major part of it in the central vacuole of cells from root and shoot tissues. Based on few studies on the two model plants Arabidopsis thaliana and rice, members of the large ChLoride Channel (CLC) family have been proposed to encode anion channels/transporters involved in nitrate homeostasis. Proteins from the Arabidopsis CLC family (AtClC, comprising seven members) are present in various membrane compartments including the vacuolar membrane (AtClCa), Golgi vesicles (AtClCd and AtClCf) or chloroplast membranes (AtClCe). Through a combination of electrophysiological and genetic approaches, AtClCa was shown to function as a 2NO3-/1H+ exchanger that is able to accumulate specifically nitrate into the vacuole, in agreement with the main phenotypic trait of knockout mutant plants that accumulate 50 per cent less nitrate than their wild-type counterparts. The set-up of a functional complementation assay relying on transient expression of AtClCa cDNA in the mutant background opens the way for studies on structure-function relationships of the AtClCa nitrate transporter. Such studies will reveal whether important structural determinants identified in bacterial or mammalian CLCs are also crucial for AtClCa transport activity and regulation.
Subject(s)
Anions/metabolism , Chloride Channels/metabolism , Plant Cells , Plants/metabolism , Plant Proteins/metabolismABSTRACT
Plants are constantly exposed to environmental biotic and abiotic stresses. Plants cells perceive these factors and trigger early responses followed by delayed and complex adaptation processes. Using cell suspensions of Arabidopsis thaliana (L.) as a cellular model, we investigated the role of plasma membrane anion channels in Reactive Oxygen Species (ROS) production and in cell death which occurs during non-host pathogen infection. Protoplasts derived from Arabidopsis suspension cells display two anion currents with characteristics very similar to those of the slow nitrate-permeable (S-type) and rapid sulfate-permeable (R-type) channels previously characterised in hypocotyl cells and other cell types. Using seven inhibitors, we showed that the R-type channel and ROS formation in cell cultures present similar pharmacological profiles. The efficiency of anion channel blockers to inhibit ROS production was independent of the nature of the triggering signal (osmotic stress or general elicitors of plant defence), indicating that the R-type channel represents a crossroad in the signalling pathways leading to ROS production. In a second step, we show that treatment with R-type channel blockers accelerates cell death triggered by the non-specific plant pathogen Xanthomonas campestris. Finally, we discuss the hypothesis that the R-type channel is involved in innate immune response allowing cell defence via antibacterial ROS production.
ABSTRACT
Plant cells, like those of animals and bacteria, are able to sense physical deformation of the plasma membrane. Mechanosensitive (MS) channels are proteins that transduce mechanical force into ion flux, providing a mechanism for the perception of mechanical stimuli such as sound, touch and osmotic pressure. We recently identified AtMSL9 and AtMSL10, two mechanosensitive channels in Arabidopsis thaliana, as molecular candidates for mechanosensing in higher plants.1 AtMSL9 and AtMSL10 are members of a family of proteins in Arabidopsis that are related to the bacterial MS channel MscS, termed MscS-Like (or MSL).2 MscS (Mechanosensitive channel of Small conductance) is one of the best-characterized MS channels, first identified as an electrophysiological activity in the plasma membrane (PM) of giant E. coli spheroplasts.3,4 Activation of MscS is voltage-independent, but responds directly to tension applied to the membrane and does not require other cellular proteins for this regulation.5,6 MscS family members are widely distributed throughout bacterial and archaeal genomes, are present in all plant genomes yet examined, and are found in selected fungal genomes.2,7,8 MscS homolgues have not yet been identified in animals.
ABSTRACT
Variations in both intracellular and extracellular pH are known to be involved in a wealth of physiological responses. Using the patch-clamp technique on Arabidopsis hypocotyl cells, it is shown that rapid-type and slow-type anion channels at the plasma membrane are both regulated by pH via distinct mechanisms. Modifications of pH modulate the voltage-dependent gating of the rapid channel. While intracellular alkalinization facilitates channel activation by shifting the voltage gate towards negative potentials, extracellular alkalinization shifts the activation threshold to more positive potentials, away from physiological resting membrane potentials. By contrast, pH modulates slow anion channel activity in a voltage-independent manner. Intracellular acidification and extracellular alkalinization increase slow anion channel currents. The possible role of these distinct modulations in physiological processes involving anion efflux and modulation of extracellular and/or intracellular pH, such as elicitor and ABA signalling, are discussed.
Subject(s)
Anions/metabolism , Arabidopsis/metabolism , Cell Membrane/metabolism , Ion Channels/metabolism , Arabidopsis Proteins/metabolism , Hydrogen-Ion Concentration , Hypocotyl/metabolism , Membrane PotentialsABSTRACT
Potassium (K+) channels play multiple roles in higher plants, and have been characterized electrophysiologically in various subcellular membranes. The K+ channel AtKCO1 from Arabidopsis thaliana is the prototype of a new family of plant K+ channels. In a previous study the protein has been functionally characterized after heterologous expression in Baculovirus-infected insect cells. In order to obtain further information on the physiological function of AtKCO1, the gene expression pattern and subcellular localization of the protein in plants were investigated. The regulatory function of the 5' region of the AtKCO1 gene was examined in transgenic A. thaliana plants carrying beta-glucuronidase (GUS) fusion constructs. Our analysis demonstrates that the AtKCO1 promoter is active in various tissues and cell types, and the highest GUS activity could be detected in mitotically active tissues of the plant. Promoter activity was strongly dependent on the presence of a 5' leader intron. The same overall structure was identified in two genes encoding AtKCO1-like K+ channels from Solanum tuberosum (StKCO1alpha and StKCO1beta). To investigate the subcellular localization of AtKCO1, the channel protein, as well as a fusion protein of AtKCO1 with green fluorescence protein (GFP), were expressed in transgenic tobacco BY2 cells. In sucrose density gradients, both proteins co-fractionate with tonoplast markers (Nt-TIPa, vATPase). In fluorescence images from transgenic AtKCO1-GFP BY2 cells fluorescence was exclusively detected in the tonoplast. Thus AtKCO1 is the first cloned K+ channel demonstrated to be a vacuolar K+ channel.