The group is interested in plant cell signaling during abiotic stress, and more specifically calcium and electric signals. We address this question targeting a family of poorly selective ion channels permeable to calcium and that are involved in both electric and calcium signaling pathways: the Glutamate receptor-like channels (GLRs). They are paralogue to animal ionotropic glutamate receptors involved in synaptic transmission and complex neurophysiological processes such as long term potentialization and associated memory and learning. Plant GLRs are involved in cell growth and organ development, as well as in biotic and abiotic stress sensing and response, including calcium and electrical long distance signal propagation. Our work includes molecular genetics, calcium and electric signal imaging, patch-clamp in protoplasts and in heterologous systems (COS and HEK cells, xenopus oocytes). We work in parallel with flowering plants (tomato and Arabidopsis thaliana) and mosses (Physcomitrela patens and Sanionia uncinata). The use of Bryophytes for experimental models has the advantage of genetic and morphological simplicity, and allows to perform simple cell biology studies which results help to understand physiological processes in much more complex angiosperm systems. S. uncinata is an extremophile plant that plays a major role in Antarctica coastal ecosystem. We are trying to understand the cellular processes conferring to this plant remarkable resistance to several extreme conditions including freezing,high salt concentration, high UV, oxidative stress and dark.
Institute of Biological Sciences, University of Talca, Chile.
Transport processes across membranes play central roles in any biological system. They are essential for homeostasis, cell nutrition, and signaling. Fluxes across membranes are governed by fundamental thermodynamic rules and are influenced by electrical potentials and concentration gradients. Transmembrane transport processes have been largely studied on single membranes. However, several important cellular or subcellular structures consist of two closely spaced membranes that form a membrane sandwich. Such a dual membrane structure results in remarkable properties for the transport processes that are not present in isolated membranes. At the core of membrane sandwich properties, a small intermembrane volume is responsible for efficient coupling between the transport systems at the two otherwise independent membranes. Here, we present the physicochemical principles of transport coupling at two adjacent membranes and illustrate this concept with three examples. In the supplementary material, we provide animated PowerPoint presentations that visualize the relationships. They could be used for teaching purposes, as has already been completed successfully at the University of Talca.
Sensing of external mineral nutrient concentrations is essential for plants to colonize environments with a large spectrum of nutrient availability. Here, we analyzed transporter networks in computational cell biology simulations to understand better the initial steps of this sensing process. The networks analyzed were capable of translating the information of changing external nutrient concentrations into cytosolic H+ and Ca2+ signals, two of the most ubiquitous cellular second messengers. The concept emerging from the computational simulations was confirmed in wet-lab experiments. We document in guard cells that alterations in the external KCl concentration were translated into cytosolic H+ and Ca2+ transients as predicted. We show that transporter networks do not only serve their primary task of transport, but can also take on the role of a receptor without requiring conformational changes of a transporter protein. Such transceptor-like phenomena may be quite common in plants.
Glutamate receptor-like channels (GLRs) play vital roles in various physiological processes in plants, such as wound response, stomatal aperture control, seed germination, root development, innate immune response, pollen tube growth, and morphogenesis. Despite the importance of GLRs, knowledge about their molecular organization is limited. Here we employ X-ray crystallography and single-particle cryo-EM to solve structures of the Arabidopsis thaliana GLR3.4. Our structures reveal the tetrameric assembly of GLR3.4 subunits into a three-layer domain architecture, reminiscent of animal ionotropic glutamate receptors (iGluRs). However, the non-swapped arrangement between layers of GLR3.4 domains, binding of glutathione through S-glutathionylation of cysteine C205 inside the amino-terminal domain clamshell, unique symmetry, inter-domain interfaces, and ligand specificity distinguish GLR3.4 from representatives of the iGluR family and suggest distinct features of the GLR gating mechanism. Our work elaborates on the principles of GLR architecture and symmetry and provides a molecular template for deciphering GLR-dependent signaling mechanisms in plants.
This article comments on: "Melatonin improves rice salinity stress tolerance by NADPH oxidase-dependent control of the plasma membrane K+ transporters and K+ homeostasis".
Animal ionotropic glutamate receptors (iGluRs) are ligand-gated channels whose evolution is intimately linked to the one of the nervous system, where the agonist glutamate and co-agonists glycine/D-serine act as neuro-transmitters or -modulators. While iGluRs are specialized in neuronal communication, plant glutamate receptor-like (GLR) homologues have evolved many plant-specific physiological functions, such as sperm signaling in moss, pollen tube growth, root meristem proliferation, innate immune and wound responses. GLRs have been associated with Ca2+ signaling by directly channeling its extracellular influx into the cytosol. Nevertheless, very limited information on functional properties of GLRs is available, and we mostly rely on structure/function data obtained for animal iGluRs to interpret experimental results obtained for plant GLRs. Yet, a deeper characterization and better understanding of plant GLRs is progressively unveiling original and different mode of functions when compared to their mammalian counterparts. Here, we review the function of plant GLRs comparing their predicted structure and physiological roles to the well-documented ones of iGluRs. We conclude that interpreting GLR function based on comparison to their animal counterparts calls for caution, especially when presuming physiological roles and mode of action for plant GLRs from comparison to iGluRs in peripheral, non-neuronal tissues.
Ion homeostasis is a crucial process in plants that is closely linked to the efficiency of nutrient uptake, stress tolerance and overall plant growth and development. Nevertheless, our understanding of the fundamental processes of ion homeostasis is still incomplete and highly fragmented. Especially at the mechanistic level, we are still in the process of dissecting physiological systems to analyze the different parts in isolation. However, modeling approaches have shown that it is not individual transporters but rather transporter networks (homeostats) that control membrane transport and associated homeostatic processes in plant cells. To facilitate access to such theoretical approaches, the modeling of the potassium homeostat is explained here in detail to serve as a blueprint for other homeostats. Based on a few, elementary knowledge about the thermodynamics of the different transport processes, it is possible to draw fundamental conclusions about the properties and physiology of the transporter network.
Compared to animals, evolution of plant calcium (Ca2+) physiology has led to a loss of proteins for influx and small ligand-operated control of cytosolic Ca2+, leaving many Ca2+ mechanisms unaccounted for. Here, we show a mechanism for sorting and activation of glutamate receptor-like channels (GLRs) by CORNICHON HOMOLOG (CNIH) proteins. Single mutants of pollen-expressed Arabidopsis thaliana GLRs (AtGLRs) showed growth and Ca2+ flux phenotypes expected for plasma membrane Ca2+ channels. However, higher-order mutants of AtGLR3.3 revealed phenotypes contradicting this assumption. These discrepancies could be explained by subcellular AtGLR localization, and we explored the implication of AtCNIHs in this sorting. We found that AtGLRs interact with AtCNIH pairs, yielding specific intracellular localizations. AtCNIHs further trigger AtGLR activity in mammalian cells without any ligand. These results reveal a regulatory mechanism underlying Ca2+ homeostasis by sorting and activation of AtGLRs by AtCNIHs. .
Perception of biotic and abiotic stresses often leads to stomatal closure in plants1,2. Rapid influx of calcium ions (Ca2+) across the plasma membrane has an important role in this response, but the identity of the Ca2+ channels involved has remained elusive3,4. Here we report that the Arabidopsis thaliana Ca2+-permeable channel OSCA1.3 controls stomatal closure during immune signalling. OSCA1.3 is rapidly phosphorylated upon perception of pathogen-associated molecular patterns (PAMPs). Biochemical and quantitative phosphoproteomics analyses reveal that the immune receptor-associated cytosolic kinase BIK1 interacts with and phosphorylates the N-terminal cytosolic loop of OSCA1.3 within minutes of treatment with the peptidic PAMP flg22, which is derived from bacterial flagellin. Genetic and electrophysiological data reveal that OSCA1.3 is permeable to Ca2+, and that BIK1-mediated phosphorylation on its N terminus increases this channel activity. Notably, OSCA1.3 and its phosphorylation by BIK1 are critical for stomatal closure during immune signalling, and OSCA1.3 does not regulate stomatal closure upon perception of abscisic acid—a plant hormone associated with abiotic stresses. This study thus identifies a plant Ca2+ channel and its activation mechanisms underlying stomatal closure during immune signalling, and suggests specificity in Ca2+ influx mechanisms in response to different stresses. .
Perception of biotic and abiotic stresses often leads to stomatal closure in plants1,2. Rapid influx of calcium ions (Ca2+) across the plasma membrane has an important role in this response, but the identity of the Ca2+ channels involved has remained elusive3,4. Here we report that the Arabidopsis thaliana Ca2+-permeable channel OSCA1.3 controls stomatal closure during immune signalling. OSCA1.3 is rapidly phosphorylated upon perception of pathogen-associated molecular patterns (PAMPs). Biochemical and quantitative phosphoproteomics analyses reveal that the immune receptor-associated cytosolic kinase BIK1 interacts with and phosphorylates the N-terminal cytosolic loop of OSCA1.3 within minutes of treatment with the peptidic PAMP flg22, which is derived from bacterial flagellin. Genetic and electrophysiological data reveal that OSCA1.3 is permeable to Ca2+, and that BIK1-mediated phosphorylation on its N terminus increases this channel activity. Notably, OSCA1.3 and its phosphorylation by BIK1 are critical for stomatal closure during immune signalling, and OSCA1.3 does not regulate stomatal closure upon perception of abscisic acid—a plant hormone associated with abiotic stresses. This study thus identifies a plant Ca2+ channel and its activation mechanisms underlying stomatal closure during immune signalling, and suggests specificity in Ca2+ influx mechanisms in response to different stresses.
Glutamate receptor-like channels (GLRs) play important roles in numerous plant physiological processes. GLRs are homologous to ionotropic glutamate receptors (iGluRs) that mediate neurotransmission in vertebrates. Here we determine crystal structures of Arabidopsis thaliana GLR3.2 ligand-binding domain (LBD) in complex with glycine and methionine to 1.58- and 1.75-Å resolution, respectively. Our structures show a fold similar to that of iGluRs, but with several secondary structure elements either missing or different. The closed clamshell conformation of GLR3.2 LBD suggests that both glycine and methionine act as agonists. The mutation R133A strongly increases the constitutive activity of the channel, suggesting that the LBD mutated at the residue critical for agonist binding produces a more stable closed clamshell conformation. Furthermore, our structures explain the promiscuity of GLR activation by different amino acids, confirm evolutionary conservation of structure between GLRs and iGluRs, and predict common molecular principles of their gating mechanisms driven by bilobed clamshell-like LBDs.
