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1. Functional Characterization of the Arabidopsis Abscisic Acid Transporters NPF4.5 and NPF4.6 in Xenopus Oocytes

Author

Sophie Léran, Mélanie Noguero, Claire Corratgé-Faillie, Yann Boursiac, Chantal Brachet, and Benoit Lacombe

Subjects
abscisic acid, transport, hormone, Km, pH, Plant culture, SB1-1110
Abstract

Few proteins have been characterized as abscisic acid transporters. Several of them are NRT1/PRT Family (NPF) transporters which have been characterized in yeast using reporter systems. Because several members of the NPF4 subfamily members were identified in yeast as ABA transporters, here, we screened for ABA transport activity the seven members of the NPF4 subfamily in Xenopus oocytes using cRNA injection and 3H-ABA accumulation. The ABA transport capacities of NPF4.2, NPF4.5, NPF4.6, and NPF4.7 were confirmed. The transport properties of NPF4.5 and NPF4.6 were studied in more detail. Both ABA transporter activities are pH-dependent and slightly pH-dependent apparent Km around 500 μM. There is no competitive inhibition of the ABA-analogs pyrabactin and quinabactin on ABA accumulation demonstrating a different selectivity compared to the ABA receptors. Functional expression of these ABA transporters in Xenopus oocyte is an opportunity to start structure–function studies and also to identify partner proteins of these hormone transporters.

Published
2020
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2. Arbuscular mycorrhizal fungusRhizophagus irregularisexpresses an outwardly Shaker-like channel involved in potassium nutrition of rice (Oryza sativaL.)

Author

Claire Corratgé-Faillie, Layla Chmaiss, Houssein Zhour, Jean-Pierre Lolivier, Pierre-Alexandre Audebert, Xuan Thai Bui, Maguette Seck, Kawiporn Chinachanta, Cécile Fizames, Daniel Wipf, Hervé Sentenac, Anne-Aliénor Very, Pierre-Emmanuel Courty, and Doan Trung Luu

Abstract

Potassium (K+) plays crucial roles in many physiological, molecular and cellular processes in plants. Direct uptake of this nutrient by root cells has been extensively investigated, however, indirect uptake of K+mediated by the interactions of the roots with fungi in the frame of a mutualistic symbiosis, also called mycorrhizal nutrient uptake pathway, is much less known. We identified an ion channel in the arbuscular mycorrhizal (AM) fungusRhizophagus irregularis. This channel exhibits the canonical features of Shaker-like channel shared in other living kingdoms and is named RiSKC3. Transcriptionally expressed in hyphae and in arbuscules of colonized rice roots, RiSKC3 has been shown to be located in the plasma membrane. Voltage-clamp functional characterization inXenopusoocytes revealed that RiSKC3 is endowed with outwardly-rectifying voltage-gated activity with a high selectivity for K+over sodium ions. RiSKC3 may have a role in the AM K+pathway for rice nutrition in normal and salt stress conditions. The current working model proposes that K+ions taken up by peripheral hyphae ofR. irregularisare secreted towards the host root into periarbuscular space by RiSKC3.Significance StatementMutualistic symbiosis with arbuscular mycorrhizal (AM) fungi are beneficial for about 80% of land plants thanks to an exchange of nutrients. The AM pathway responsible for potassium (K+) nutrition of the plant is not known. Here we uncovered a key step of this phenomenon, by functionally characterizing the first transport system in the AM fungusRhizophagus irregularis, and we univocally demonstrated that RiSKC3 is an K+outwardly-rectifying voltage-gated Shaker-like channel.

Published
2022
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3. Recherche de systèmes de transport de potassium impliqués dans le transfert de K+ de la mycorhize arbusculaire au riz lors d'interactions symbiotiques

Author

Luu Doan Trung, Pierre-Emmanuel Courty, claire corratgé-faillie, Layla Chmaiss, Zhour, Houssein, Jean-Pierre Lolivier, Pierre-Alexandre Audebert, Bui, Xuan Thai, Maguette Seck, Hervé Sentenac, Anne-Aliénor Véry, and EL Mjiyad, Noureddine

Subjects
[SDV] Life Sciences [q-bio], Potassium, Rhizophagus irregularis, salinité, riz
Abstract

Les champignons mycorhiziens à arbuscules (CMA) développent des connexions interdépendantes avec les racines d'environ 90% des espèces végétales. Ces interactions augmentent la disponibilité ainsi que la translocation des nutriments (en particulier N et P), et améliorent ainsi la nutrition et la croissance des plantes. De plus, la résistance à une variété de stress, parmi lesquels le stress salin, s'est avérée améliorée par les interactions CMA-plante, par exemple chez le riz. Des recherches intenses pour expliquer les mécanismes moléculaires des interactions bénéfiques CMA-plante ont conduit à l'identification de transporteurs de phosphate et d'ammonium impliqués dans les échanges de nutriments du CMA vers la plante, chez plusieurs espèces végétales. Malgré l'importance du potassium (K+) pour la physiologie des plantes, la contribution de la symbiose mycorhizienne à arbuscule à la nutrition en K+ des plantes a été peu documentée. La surexpression des transporteurs de K+ végétaux a été décrite chez Lotus japonicus et la tomate en condition de symbiose mycorhizienne à arbuscule. Ici, la nutrition en K+ du riz colonisé par Rhizophagus irregulis a été analysée aux niveaux moléculaire et physiologique. Étonnamment, les principaux systèmes de transport de K+ dans le riz étaient régulés à la baisse lors des interactions AMF, suggérant une forte augmentation de la disponibilité de K+ pour l'absorption par les cellules racinaires dans des conditions symbiotiques. De plus, des systèmes de transport de K+ dans le CMA R. irregularis ont été identifiés in silico. La fonction de l’un d’entre eux a été analysée. Le rôle de K+ dans les relations entre le riz et R. irregularis sera également discuté.

Published
2022

4. Tolérance des plantes à la salinité des sols

Author

Luu Doan Trung, Jing Zhou, Dorsaf Hmidi, Madani, Ikram, Zhour, Houssein, Layla Chmaiss, claire corratgé-faillie, Hervé Sentenac, Anne-Aliénor Véry, and LUU, DOAN TRUNG

Subjects
[SDV] Life Sciences [q-bio]
Published
2022

5. The Arabidopsis protein NPF6.2/NRT1.4 is a plasma membrane nitrate transporter and a target of protein kinase CIPK23

Author

Alexis De Angeli, Claire Corratgé-Faillie, Francisco J. Quintero, Marika Lindahl, José M. Pardo, Laura Morales de los Ríos, Benoît Lacombe, Imelda Mendoza, Natalia Raddatz, Agencia Estatal de Investigación (España), and European Research Council

Subjects
Physiology, Potassium, Anion Transport Proteins, Arabidopsis, chemistry.chemical_element, Plant Science, Nitrate, Plant Roots, chemistry.chemical_compound, Genetics, Arabidopsis thaliana, Plant nutrition, Protein kinase A, Ion transporter, Ion transport, Plant Proteins, Nitrates, biology, Chemistry, Arabidopsis Proteins, Cell Membrane, Nitrate Transporters, biology.organism_classification, Biochemistry, Mutation, Heterologous expression, Protein Kinases, Plant Shoots
Abstract

Nitrate and potassium nutrition is tightly coordinated in vascular plants. Physiological and molecular genetics studies have demonstrated that several NPF/NRT1 nitrate transporters have a significant impact on both uptake and the root-shoot partition of these nutrients. However, how these traits are biochemically connected remain controversial since some NPF proteins, e.g. NPF7.3/NRT1.5, have been suggested to mediate K+/H+ exchange instead of nitrate fluxes. Here we show that NPF6.2/NRT1.4, a protein that gates nitrate accumulation at the leaf petiole of Arabidopsis thaliana, also affects the root/shoot distribution of potassium. We demonstrate that NPF6.2/NRT1.4 is a plasma membrane nitrate transporter phosphorylated at threonine-98 by the CIPK23 protein kinase that is a regulatory hub for nitrogen and potassium nutrition. Heterologous expression of NPF6.2/NRT1.4 and NPF7.3/NRT1.5 in yeast mutants with altered potassium uptake and efflux systems showed no evidence of nitrate-dependent potassium transport by these proteins

Published
2021

6. Search for potassium transport systems involved in arbuscular mycorrhiza-rice symbiotic interactions

Author

Luu, Truong Luu, Pierre-Emmanuel Courty, claire corratgé-faillie, Jean-Pierre Lolivier, Pierre-Alexandre Audebert, Bui, Xuan Thai, Maguette Seck, Hervé Sentenac, Anne-Aliénor Véry, Zhour, Houssein, and EL Mjiyad, Noureddine

Subjects
[SDV] Life Sciences [q-bio]
Abstract

Arbuscular mycorrhizal fungi (AMF) develop interdependent connections with roots of about90% of plant species. These interactions increase availability as well as translocation ofnutrients (especially N and P), and thereby improve plant nutrition and growth. Moreover,resistance to a variety of stresses, among which salt stress, has been shown to be improved byAMF-plant interactions, for example in rice. Intense research to explain the molecularmechanisms of AMF-plant beneficial interactions led to the identification of phosphate andammonium transporters involved in nutrient exchanges from AMF to the plant, in several plantspecies. In spite of the importance of potassium (K+) for plant physiology, the contribution ofAMF symbiosis to plant K+ nutrition has been little documented. Over-expression of plant K+transporters has been described in Lotus japonicus and tomato in condition of AMF symbiosis.Furthermore, K+ transport systems in the AMF Rhizophagus irregularis have been identified insilico. Here, K+ nutrition in rice colonized by R. irregularis has been analyzed at molecular andphysiological levels. Surprisingly, major K+ transport systems in rice were down-regulated uponAMF interactions, suggesting strong increase in K+ availability for uptake by root cells insymbiotic conditions. Role of K+ in the relationships between rice and R. irregularis will also bediscussed

Published
2021

7. Ca

Author

Elsa, Ronzier, Claire, Corratgé-Faillie, Frédéric, Sanchez, Christian, Brière, and Tou Cheu, Xiong

Subjects
shaker channel, Models, Molecular, post-translational regulation, Patch-Clamp Techniques, Arabidopsis thaliana, Arabidopsis Proteins, Optical Imaging, Arabidopsis, In Vitro Techniques, Plants, Genetically Modified, calcium signaling, Recombinant Proteins, Article, Plant Leaves, CPK, Gene Knockout Techniques, Xenopus laevis, Potassium Channels, Voltage-Gated, Calcium-Calmodulin-Dependent Protein Kinases, Oocytes, Potassium, Fluorescence Resonance Energy Transfer, Animals, Female, Phosphorylation
Abstract

Post-translational regulations of Shaker-like voltage-gated K+ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, a Shaker channel cloned in the model plant Arabidopsis thaliana, where is it expressed namely in the vascular tissues of leaves. After co-expression of KAT2 with AtCPK6 in Xenopus laevis oocytes, voltage-clamp recordings demonstrated that AtCPK6 stimulates the activity of KAT2 in a calcium-dependent manner. A physical interaction between these two proteins has also been shown by Förster resonance energy transfer by fluorescence lifetime imaging (FRET-FLIM). Peptide array assays support that AtCPK6 phosphorylates KAT2 at several positions, also in a calcium-dependent manner. Finally, K+ fluorescence imaging in planta suggests that K+ distribution is impaired in kat2 knock-out mutant leaves. We propose that the AtCPK6/KAT2 couple plays a role in the homeostasis of K+ distribution in leaves.

Published
2020

8. Homology modeling identifies crucial amino-acid residues that confer higher Na+ transport capacity of OcHKT1;5 from Oryza coarctata Roxb

Author

Ramanathan Sowdhamini, Kavitha Kumaresan, Sergey Shabala, Claire Corratgé-Faillie, Gayatri Venkataraman, Suji Somasundaram, Shalini Pulipati, Lana Shabala, Anne-Aliénor Véry, Kumkum Kumari, Tetsuya Ishikawa, Rithvik S. Vinekar, Ajay Parida, Plant Molecular Biology Laboratory, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), National Centre for Biological Sciences (NCBS), NCBS, Tasmanian Institute of Agriculture, University of Tasmania [Hobart, Australia] (UTAS), Krishi Vigyan Kendra, National Centre for Biological Sciences [TIFR] (NCBS), Tata Institute for Fundamental Research (TIFR), Institute of Life Sciences (ILS), and School of Agricultural Sciences

Subjects
0106 biological sciences, Physiology, Sodium, Xenopus, homology modeling, [SDV]Life Sciences [q-bio], Halophyte, chemistry.chemical_element, Plant Science, yeast, 01 natural sciences, 03 medical and health sciences, Xylem, Animals, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, HKT1, Amino Acids, Cation Transport Proteins, 030304 developmental biology, Plant Proteins, Xenopus oocytes, chemistry.chemical_classification, 0303 health sciences, biology, Organisms, Genetically Modified, Sequence Homology, Amino Acid, Na+transporter, Cell Membrane, food and beverages, Transporter, Oryza, Salt-Tolerant Plants, Cell Biology, General Medicine, Oryza coarctata, biology.organism_classification, simulation, Amino acid, Salinity, chemistry, Biochemistry, Shoot, Oocytes, 010606 plant biology & botany
Abstract

HKT1;5 loci/alleles are important determinants of crop salinity tolerance. HKT1;5s encode plasmalemma-localized Na+ transporters, which move xylem Na+ into xylem parenchyma cells, reducing shoot Na+ accumulation. Allelic variation in rice OsHKT1;5 sequence in specific landraces (Nona Bokra OsHKT1;5-NB/Nipponbare OsHKT1;5-Ni) correlates with variation in salt tolerance. Oryza coarctata, a halophytic wild rice, grows in fluctuating salinity at the seawater–estuarine interface in Indian and Bangladeshi coastal regions. The distinct transport characteristics of the shoots and roots expressing the O. coarctata OcHKT1;5 transporter are reported vis-à-vis OsHKT1;5-Ni. Yeast sodium extrusion-deficient cells expressing OcHKT1;5 are sensitive to increasing Na+ (10–100 mM). Electrophysiological measurements in Xenopus oocytes expressing O. coarctata or rice HKT1;5 transporters indicate that OcHKT1;5, like OsHKT1;5-Ni, is a Na+-selective transporter, but displays 16-fold lower affinity for Na+ and 3.5-fold higher maximal conductance than OsHKT1;5-Ni. For Na+ concentrations >10 mM, OcHKT1;5 conductance is higher than that of OsHKT1;5-Ni, indicating the potential of OcHKT1;5 for increasing domesticated rice salt tolerance. Homology modeling/simulation suggests that four key amino-acid changes in OcHKT1;5 (in loops on the extracellular side; E239K, G207R, G214R, L363V) account for its lower affinity and higher Na+ conductance vis-à-vis OsHKT1;5-Ni. Of these, E239K in OcHKT1;5 confers lower affinity for Na+ transport, as evidenced by Na+ transport assays of reciprocal site-directed mutants for both transporters (OcHKT1;5-K239E, OsHKT1;5-Ni-E270K) in Xenopus oocytes. Both transporters have likely analogous roles in xylem sap desalinization, and differences in xylem sap Na+ concentrations in both species are attributed to differences in Na+ transport affinity/conductance between the transporters.

Published
2020
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9. Functional Characterization of the Arabidopsis Abscisic Acid Transporters NPF4.5 and NPF4.6 in Xenopus Oocytes

Author

Mélanie Noguero, Sophie Léran, Claire Corratgé-Faillie, Chantal Brachet, Yann Boursiac, Benoît Lacombe, UMR - Interactions Plantes Microorganismes Environnement (UMR IPME), Institut de Recherche pour le Développement (IRD [France-Sud])-Université de Montpellier (UM)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), ANR-11-JSV6-0002,NUTSE,Perception des nutriments chez les plantes(2011), ANR-14-CE34-0007,HONIT,Hormone cross-talk drives nutrient dependent plant development(2014), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Université de Montpellier (UM)-Institut de Recherche pour le Développement (IRD [France-Sud]), and Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)

Subjects
0106 biological sciences, 0301 basic medicine, Subfamily, Arabidopsis thaliana, F60 - Physiologie et biochimie végétale, membrane transport, acide abscisique, hormone, Xenopus, F62 - Physiologie végétale - Croissance et développement, Plant Science, yeast, lcsh:Plant culture, 01 natural sciences, Km, abscisic acid, 03 medical and health sciences, chemistry.chemical_compound, Arabidopsis, abscissins, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, lcsh:SB1-1110, ovocyte de xénope, transport membranaire, Receptor, Abscisic acid, levure, Original Research, Pyrabactin, biology, pH, organic chemicals, fungi, food and beverages, Transporter, Membrane transport, biology.organism_classification, Cell biology, arabidopsis, 030104 developmental biology, chemistry, ABA, transport, 010606 plant biology & botany
Abstract

Dynamic reprogramming of gene regulatory networks (GRNs) enables organisms to rapidlyrespond to environmental perturbation. However, the underlying transient interactionsbetween transcription factors (TFs) and genome-wide targets typically elude biochemicaldetection. Here, we capture both stable and transient TF-target interactions genome-widewithin minutes after controlled TF nuclear import using time-series chromatin immunoprecipitation(ChIP-seq) and/or DNA adenine methyltransferase identification (DamID-seq).The transient TF-target interactions captured uncover the early mode-of-action of NIN-LIKEPROTEIN 7 (NLP7), a master regulator of the nitrogen signaling pathway in plants. Thesetransient NLP7 targets captured in root cells using temporal TF perturbation account for 50%of NLP7-regulated genes not detectably bound by NLP7 in planta. Rapid and transient NLP7binding activates early nitrogen response TFs, which we validate to amplify the NLP7-initiatedtranscriptional cascade. Our approaches to capture transient TF-target interactions genomewidecan be applied to validate dynamic GRN models for any pathway or organism of interest.

Published
2020
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10. BCL2-ASSOCIATED ATHANOGENE4 Regulates the KAT1 Potassium Channel and Controls Stomatal Movement

Author

Antonella Locascio, Lynne Yenush, Claire Corratgé-Faillie, Nuria Andrés-Colás, Maria Carmen Marques, José A. Fernández, Lourdes Rubio, José M. Mulet, Anne-Aliénor Véry, Guillermo García-Martínez, Instituto de Biología Molecular y Celular de Plantas, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Facultad de Ciencias (UDELAR), Universidad de Málaga (Málaga), and Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0106 biological sciences, Patch-Clamp Techniques, approche génétique, Physiology, [SDV]Life Sciences [q-bio], stomata, Regulator, Arabidopsis, K+ 59 channel regulator, KAT2, Expression, KAT1, Plant Science, stomate, 01 natural sciences, chemistry.chemical_compound, Guard cell, Arabidopsis thaliana, Abscisic acid, Research Articles, Trafficking, biology, Chemistry, BAG4, Signal transducing adaptor protein, système hétérologue, Potassium channel, Cell biology, Potassium Channels, Voltage-Gated, heterologous system, BCL-2-Associated athanogene, Abscisic-Acid, Plasma-Membrane, stomata regulation, Genetics, BIOQUIMICA Y BIOLOGIA MOLECULAR, Saccharomyces-Cerevisiae, Family, Patch clamp, Potassium Channels, Inwardly Rectifying, Endoplasmic-Reticulum, Adaptor Proteins, Signal Transducing, shaker potassium channel, Arabidopsis Proteins, Cell Membrane, biology.organism_classification, Plant Stomata, Potassium, Plant K+ channel, canaux potassiques shaker, contrôle biologique, 010606 plant biology & botany
Abstract

[EN] Potassium (K+) is a key monovalent cation necessary for multiple aspects of cell growth and survival. In plants, this cation also plays a key role in the control of stomatal movement. KAT1 and its homolog KAT2 are the main inward rectifying channels present in guard cells, mediating K+ influx into these cells, resulting in stomatal opening. To gain further insight into the regulation of these channels, we performed a split-ubiquitin protein-protein interaction screen searching for KAT1 interactors in Arabidopsis (Arabidopsis thaliana). We characterized one of these candidates, BCL2-ASSOCIATED ATHANOGENE4 (BAG4), in detail using biochemical and genetic approaches to confirm this interaction and its effect on KAT1 activity. We show that BAG4 improves KAT1-mediated K+ transport in two heterologous systems and provide evidence that in plants, BAG4 interacts with KAT1 and favors the arrival of KAT1 at the plasma membrane. Importantly, lines lacking or overexpressing the BAG4 gene show altered KAT1 plasma membrane accumulation and alterations in stomatal movement. Our data allowed us to identify a KAT1 regulator and define a potential target for the plant BAG family. The identification of physiologically relevant regulators of K+ channels will aid in the design of approaches that may impact drought tolerance and pathogen susceptibility., The authors would like to thank Elena Moreno, María José Falaguer, Alejandro Mossi, Sara Aljama, José Antonio Navarro, Vicente Pallás, Daniel Franco-Aragón, Jorge Lozano, and Stan Gelvin for assistance in the completion of this work and for providing reagents. This work was supported by the Spanish Ministry of Economy and Competitiveness (BIO201677776-P and BIO2016-81957-REDT) and the Valencian Government (AICO/2018/300).

Published
2019
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11. Ca2+-Dependent Protein Kinase 6 Enhances KAT2 Shaker Channel Activity in Arabidopsis thaliana

Author

Elsa Ronzier, Tou Cheu Xiong, Claire Corratgé-Faillie, Christian Brière, Frédéric Sanchez, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Biologie intégrative des organismes marins (BIOM), Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Observatoire océanologique de Banyuls (OOB), Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Recherche en Sciences Végétales (LRSV), Université Toulouse III - Paul Sabatier (UT3), and Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)

Subjects
0106 biological sciences, 0301 basic medicine, Arabidopsis thaliana, [SDV]Life Sciences [q-bio], Mutant, calcium signaling, 01 natural sciences, Catalysis, lcsh:Chemistry, Inorganic Chemistry, 03 medical and health sciences, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Shaker, Physical and Theoretical Chemistry, Protein kinase A, lcsh:QH301-705.5, Molecular Biology, ComputingMilieux_MISCELLANEOUS, Spectroscopy, Calcium signaling, shaker channel, post-translational regulation, biology, Kinase, Chemistry, Organic Chemistry, General Medicine, biology.organism_classification, Computer Science Applications, Cell biology, CPK, 030104 developmental biology, Förster resonance energy transfer, lcsh:Biology (General), lcsh:QD1-999, Phosphorylation, 010606 plant biology & botany
Abstract

Post-translational regulations of Shaker-like voltage-gated K+ channels were reported to be essential for rapid responses to environmental stresses in plants. In particular, it has been shown that calcium-dependent protein kinases (CPKs) regulate Shaker channels in plants. Here, the focus was on KAT2, a Shaker channel cloned in the model plant Arabidopsis thaliana, where is it expressed namely in the vascular tissues of leaves. After co-expression of KAT2 with AtCPK6 in Xenopuslaevis oocytes, voltage-clamp recordings demonstrated that AtCPK6 stimulates the activity of KAT2 in a calcium-dependent manner. A physical interaction between these two proteins has also been shown by Förster resonance energy transfer by fluorescence lifetime imaging (FRET-FLIM). Peptide array assays support that AtCPK6 phosphorylates KAT2 at several positions, also in a calcium-dependent manner. Finally, K+ fluorescence imaging in planta suggests that K+ distribution is impaired in kat2 knock-out mutant leaves. We propose that the AtCPK6/KAT2 couple plays a role in the homeostasis of K+ distribution in leaves.

Published
2021
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12. Unravelling nutrient exchange in ectomycorrhizal symbiosis contributing to plant potassium nutrition

Author

Gabriella Houdinet, Carmen Guerrero-Galán, Kevin Garcia, Amandine Delteil, claire corratgé-faillie, Geneviève Conejero, Isabelle Gaillard, Bruno Touraine, Herve Sentenac, Sabine Zimmermann, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), and Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
symbiose ectomycorhizienne, [SDV]Life Sciences [q-bio], fungi, education, food and beverages, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, nutrition potassique, social sciences, health care economics and organizations
Abstract

Unravelling nutrient exchange in ectomycorrhizal symbiosis contributing to plant potassium nutrition. 12th Congress of the International Plant Molecular Biology

Published
2018

13. Substrate (un)specificity of Arabidopsis NRT1/PTR FAMILY (NPF) proteins

Author

Claire Corratgé-Faillie, Benoît Lacombe, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)

Subjects
0106 biological sciences, 0301 basic medicine, Physiology, acide abscisique, Anion Transport Proteins, hormone, Peptide, Plant Science, plant hormone, 01 natural sciences, Substrate Specificity, 03 medical and health sciences, chemistry.chemical_compound, Nitrate, Auxin, nitrate, transporteur de nitrate, Arabidopsis, abscissins, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, spécificité du substrat, nitrite, Abscisic acid, chemistry.chemical_classification, biology, auxine, Arabidopsis Proteins, Substrate (chemistry), food and beverages, Glucosinolate, Transporter, biology.organism_classification, peptide, jasmonate, arabidopsis, 030104 developmental biology, chemistry, Biochemistry, transporter, Gibberellin, auxin, 010606 plant biology & botany, hormone végétale
Abstract

article accepté mais pas encore paru; The conventional approach to categorizing transporters has been to class them according to their sequence homology, defining a ‘family’ (or a ‘superfamily’ if they are numerous), and according to their substrate specificity or selectivity. This general view is still relevant for some transporters, but it is being increasingly challenged. Here, we take the NRT1/PTR FAMILY (NPF) as one such example. NPF members do indeed display sequence and structural homologies with peptide transporter (PTR) proteins involved in the uptake of di- and tri-peptides in most other organisms. And in plants they were initially characterized as nitrate or peptide transporters. However, in recent years several other substrates have been identified, namely nitrite, chloride, glucosinolates, auxin (IAA), abscisic acid (ABA), jasmonates (JAs), and gibberellins (GAs). Some of these transporters are even capable of transporting more than one different substrate (e.g. nitrate/auxin, nitrate/ABA, nitrate/glucosinolates, or GA/JA). In this review, we give an overview of the substrate-specificity of the Arabidopsis NPF.

Published
2017
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14. The Arabidopsis guard cell outward potassium channel GORK is regulated by CPK33

Author

Nathalie Leonhardt, Frédéric Sanchez, Tou Cheu Xiong, Jeong-Hyeon Kim, Elsa Ronzier, Claire Corratgé-Faillie, Benoît Lacombe, Sophie Lanciano, Karine Prado, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Signalisation de l'Adaptation des Végétaux à l'Environnement (SAVE), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), and Plant Environmental Physiology and Stress Signaling (PEPSS)

Subjects
0301 basic medicine, oeuf de xenopus, clamp, Potassium Channels, Voltage clamp, Movement, Phosphatase, guard cells, Biophysics, Xenopus, Biochemistry, cellule de garde, CPKs, 03 medical and health sciences, Gene Knockout Techniques, shaker K+ channels, Structural Biology, Arabidopsis, Guard cell, canal à potassium, Botany, Genetics, oocytes, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Shaker, voltage clamp, Potassium Channels, Inwardly Rectifying, Molecular Biology, biology, Kinase, Chemistry, Arabidopsis Proteins, urogenital system, Cell Biology, biology.organism_classification, Potassium channel, Cell biology, arabidopsis, 030104 developmental biology, Potassium Channels, Voltage-Gated, voltage, Mutation, Plant Stomata, Calcium, guard cell, transporteur ionique, Protein Kinases
Abstract

A complex signaling network involving voltage-gated potassium channels from the Shaker family contributes to the regulation of stomatal aperture. Several kinases and phosphatases have been shown to be crucial for ABA-dependent regulation of the ion transporters. To date, the Ca(2+) -dependent regulation of Shaker channels by Ca(2+) -dependent protein kinases (CPKs) is still elusive. A functional screen in Xenopus oocytes was launched to identify such CPKs able to regulate the three main guard cell Shaker channels KAT1, KAT2, and GORK. Seven guard cell CPKs were tested and multiple CPK/Shaker couples were identified. Further work on CPK33 indicates that GORK activity is enhanced by CPK33 and unaffected by a nonfunctional CPK33 (CPK33-K102M). Furthermore, Ca(2+) -induced stomatal closure is impaired in two cpk33 mutant plants.

Published
2017
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15. Nitrate sensing and uptake in Arabidopsis are enhanced by ABI2, a phosphatase inactivated by the stress hormone abscisic acid

Author

Pascal Tillard, Kai H. Edel, Alain Gojon, Jan Niklas Offenborn, Sophie Léran, Benoît Lacombe, Kenji Hashimoto, Claire Corratgé-Faillie, Marjorie Pervent, Jörg Kudla, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster (WWU), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)

Subjects
[SDV]Life Sciences [q-bio], Phosphatase, Anion Transport Proteins, Arabidopsis, chemistry.chemical_element, Biological Transport, Active, Calcium, engineering.material, Nitrate, Biochemistry, nitrogen, abscisic acid, chemistry.chemical_compound, Xenopus laevis, nitrate sensing, Stress, Physiological, Phosphoprotein Phosphatases, Arabidopsis thaliana, Animals, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, nitrate transport, Molecular Biology, Abscisic acid, nitrate signaling, ComputingMilieux_MISCELLANEOUS, Plant Proteins, Nitrates, biology, Cell Biology, biology.organism_classification, chemistry, Nitrate transport, engineering, Fertilizer
Abstract

International audience; Living organisms sense and respond to changes in nutrient availability to cope with diverse environmental conditions. Nitrate (NO3-) is the main source of nitrogen for plants and is a major component in fertilizer. Unraveling the molecular basis of nitrate sensing and regulation of nitrate uptake should enable the development of strategies to increase the efficiency of nitrogen use and maximize nitrate uptake by plants, which would aid in reducing nitrate pollution. NPF6.3 (also known as NRT1.1), which functions as a nitrate sensor and transporter; the kinase CIPK23; and the calcium sensor CBL9 form a complex that is crucial for nitrate sensing in Arabidopsis thaliana. We identified two additional components that regulate nitrate transport, sensing, and signaling: the calcium sensor CBL1 and protein phosphatase 2C family member ABI2, which is inhibited by the stress-response hormone abscisic acid. Bimolecular fluorescence complementation assays and in vitro kinase assays revealed that ABI2 interacted with and dephosphorylated CIPK23 and CBL1. Coexpression studies in Xenopus oocytes and analysis of plants deficient in ABI2 indicated that ABI2 enhanced NPF6.3-dependent nitrate transport, nitrate sensing, and nitrate signaling. These findings suggest that ABI2 may functionally link stress-regulated control of growth and nitrate uptake and utilization, which are energy-expensive processes.

Published
2015
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16. AtNPF5.5, a nitrate transporter affecting nitrogen accumulation in Arabidopsis embryo

Author

Sophie Léran, Claire Corratgé-Faillie, Chantal Brachet, Benoît Lacombe, Pascal Tillard, Alain Gojon, Bharti Garg, Yann Boursiac, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Léran, Sophie, Garg, Bharti, Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut National de la Recherche Agronomique (CJS PhD Fellowship) [BAP2013-33-NITSE], Indo-French Centre for the Promotion of Advance Research (CEFIPRA) [4609-1], Agence Nationale de la Recherche [ANR-11-JSV6-002-01-NUTSE], Agropolis Fondation (RHIZOPOLIS) [07024], and Region Languedoc-Roussillon (Chercheur d'Avenir)

Subjects
transport de nitrate, transport nitrate, Nitrogen, Xenopus, transporter AtNPF8.1 and AtNPF8.3, [SDV]Life Sciences [q-bio], Anion Transport Proteins, Molecular Sequence Data, Saccharomyces cerevisiae, Arabidopsis, Dipeptide (Leu-Leu), Biology, Article, Gene Knockout Techniques, chemistry.chemical_compound, Gene Expression Regulation, Plant, nitrate, Animals, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Amino Acid Sequence, RNA, Messenger, Nitrates, Multidisciplinary, Dipeptide, Arabidopsis Proteins, Biological Transport, Nitrate Transporters, Embryo, Dipeptides, biology.organism_classification, Transport protein, Dipeptide transport, chemistry, Biochemistry, Nitrate transport, Seeds, Oocytes, NPF5.5, Xenopus laevis oocytes, AtNPF5.5a and AtNPF5.5b, dipeptide
Abstract

International audience; Dipeptide (Leu-Leu) and nitrate transport activities of 26 Arabidopsis NPF (NRT1/PTR Family) proteins were screened in Saccharomyces cerevisiae and Xenopus laevis oocytes, respectively. Dipeptide transport activity has been confirmed for 2 already known dipeptide transporters (AtNPF8.1 and AtNPF8.3) but none of the other tested NPFs displays dipeptide transport. The nitrate transport screen resulted in the identification of two new nitrate transporters, AtNPF5.5 and AtNPF5.10. The localization of the mRNA coding for NPF5.5 demonstrates that it is the first NPF transporter reported to be expressed in Arabidopsis embryo. Two independent homozygous npf5.5 KO lines display reduced total nitrogen content in the embryo as compared to WT plants, demonstrating an effect of NPF5.5 function on the embryo nitrogen content. Finally, NPF5.5 gene produces two different transcripts (AtNPF5.5a and AtNPF5.5b) encoding proteins with different N-terminal ends. Both proteins are able to transport nitrate in xenopus oocytes.

Published
2014
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17. CPK13, a noncanonical Ca2+-dependent protein kinase, specifically inhibits KAT2 and KAT1 shaker K+ channels and reduces stomatal opening

Author

Christian Brière, Claire Corratgé-Faillie, Jean-Baptiste Thibaud, Nathalie Leonhardt, Frédéric Sanchez, Tou Cheu Xiong, Elsa Ronzier, Karine Prado, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Recherche en Sciences Végétales (LRSV), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Biologie végétale et microbiologie environnementale - UMR7265 (BVME), Institut de Biosciences et Biotechnologies d'Aix-Marseille (ex-IBEB) (BIAM), Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Agropolis Foundation (grant no. AA0803-022), European Project: 268393,EC:FP7:PEOPLE,FP7-PEOPLE-2010-RG,PLANTELEXIGNAL(2010), Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), and Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)-Direction de Recherche Fondamentale (CEA) (DRF (CEA))

Subjects
0106 biological sciences, KAT2 channels, Physiology, Xenopus, KAT1, Plant Science, Ca 2+-dependent, 01 natural sciences, CPKs, 03 medical and health sciences, Arabidopsis, Guard cell, Botany, Genetics, Arabidopsis thaliana, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Patch clamp, Protein kinase A, 030304 developmental biology, Xenopus oocytes, 0303 health sciences, biology, fungi, protein kinase, biology.organism_classification, Plant cell, Potassium channel, Cell biology, 010606 plant biology & botany
Abstract

Ca2 +-dependent protein kinases (CPKs) form a large family of 34 genes in Arabidopsis (Arabidopsis thaliana). Based on their dependence on Ca2+, CPKs can be sorted into three types: strictly Ca2+-dependent CPKs, Ca2+-stimulated CPKs (with a significant basal activity in the absence of Ca2+), and essentially calcium-insensitive CPKs. Here, we report on the third type of CPK, CPK13, which is expressed in guard cells but whose role is still unknown. We confirm the expression of CPK13 in Arabidopsis guard cells, and we show that its overexpression inhibits light-induced stomatal opening. We combine several approaches to identify a guard cell-expressed target. We provide evidence that CPK13 (1) specifically phosphorylates peptide arrays featuring Arabidopsis K+ Channel KAT2 and KAT1 polypeptides, (2) inhibits KAT2 and/or KAT1 when expressed in Xenopus laevis oocytes, and (3) closely interacts in plant cells with KAT2 channels (Förster resonance energy transfer-fluorescence lifetime imaging microscopy). We propose that CPK13 reduces stomatal aperture through its inhibition of the guard cell-expressed KAT2 and KAT1 channels.

Published
2014
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18. Imaging long distance propagating calcium signals in intact plant leaves with the BRET-based GFP-aequorin reporter

Author

Elsa Ronzier, Claire Corratgé-Faillie, Frédéric Sanchez, Tou Cheu Xiong, Christian Mazars, Jean-Baptiste Thibaud, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Recherche en Sciences Végétales (LRSV), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), CNRS-INRA fellowship, Agropolis Fondation [0803-022], and European Commission [IRG 268393]

Subjects
cooled CCD camera, Arabidopsis thaliana, Aequorin, Plant Science, lcsh:Plant culture, 7. Clean energy, calcium imaging, GFP-aequorin, leaf, long distance calcium signaling, calcium waves, salt stress, Green fluorescent protein, Calcium imaging, Guard cell, Botany, Methods Article, Bioluminescence, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, lcsh:SB1-1110, calcium, biology, fungi, food and beverages, biology.organism_classification, Plant cell, Fluorescence, Aequorea victoria, biology.protein, Biophysics
Abstract

International audience; Calcium (Ca(2+)) is a second messenger involved in many plant signaling processes. Biotic and abiotic stimuli induce Ca(2+) signals within plant cells, which, when decoded, enable these cells to adapt in response to environmental stresses. Multiple examples of Ca(2+) signals from plants containing the fluorescent yellow cameleon sensor (YC) have contributed to the definition of the Ca(2+) signature in some cell types such as root hairs, pollen tubes and guard cells. YC is, however, of limited use in highly autofluorescent plant tissues, in particular mesophyll cells. Alternatively, the bioluminescent reporter aequorin enables Ca(2+) imaging in the whole plant, including mesophyll cells, but this requires specific devices capable of detecting the low amounts of emitted light. Another type of Ca(2+) sensor, referred to as GFP-aequorin (G5A), has been engineered as a chimeric protein, which combines the two photoactive proteins from the jellyfish Aequorea victoria, the green fluorescent protein (GFP) and the bioluminescent protein aequorin. The Ca(2+)-dependent light-emitting property of G5A is based on a bioluminescence resonance energy transfer (BRET) between aequorin and GFP. G5A has been used for over 10 years for enhanced in vivo detection of Ca(2+) signals in animal tissues. Here, we apply G5A in Arabidopsis and show that G5A greatly improves the imaging of Ca(2+) dynamics in intact plants. We describe a simple method to image Ca(2+) signals in autofluorescent leaves of plants with a cooled charge-coupled device (cooled CCD) camera. We present data demonstrating how plants expressing the G5A probe can be powerful tools for imaging of Ca(2+) signals. It is shown that Ca(2+) signals propagating over long distances can be visualized in intact plant leaves and are visible mainly in the veins.

Published
2014
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19. Promoter-dependent expression of the fungal transporter HcPT1.1 under Pi shortage and its spatial localization in ectomycorrhiza

Author

Kevin Garcia, Amandine Delteil, Muhammad Zulqurnain Haider, Claire Corratgé-Faillie, Claude Plassard, Laurie Amenc, Marie-Violaine Tatry, Hervé Sentenac, Sabine Zimmermann, Geneviève Conejero, Adeline Becquer, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Plateforme d'histocytologie et d'imagerie cellulaire végétale (PHIV), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Amélioration génétique et adaptation des plantes méditerranéennes et tropicales (UMR AGAP), Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro), Ecologie fonctionnelle et biogéochimie des sols et des agro-écosystèmes (UMR Eco&Sols), Institut National de la Recherche Agronomique (INRA)-Institut de Recherche pour le Développement (IRD)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Amélioration génétique et adaptation des plantes méditerranéennes et tropicales (UMR AGAP), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro), Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut de Recherche pour le Développement (IRD)-Institut National de la Recherche Agronomique (INRA), French Minister of Research and Technology, Higher Education Commission (HEC) of Pakistan, ANR project 'TRANSMUT' [2010 BLAN 1604 03], INRA, and ANR-10-BLAN-1604,TRANSMUT,Rôle du transportome aux interfaces biotrophes des interactions mutualistes plantes-champignons.(2010)

Subjects
0106 biological sciences, Hyphae, Hebeloma cylindrosporum phosphate transporter HcPT1.1, 01 natural sciences, Microbiology, Phosphates, Agrotransformation, Fungal Proteins, 03 medical and health sciences, Gene Expression Regulation, Fungal, Mycorrhizae, Translational regulation, Botany, Genetics, Hebeloma, Phosphate Transport Proteins, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Promoter Regions, Genetic, Symbiosis, GFP-promoter expression, 030304 developmental biology, 2. Zero hunger, 0303 health sciences, Hartig net, Fungal protein, Protein immunolocalization, biology, Ectomycorrhizal (ECM) symbiosis, Transporter, Membrane transport, Pinus, biology.organism_classification, Protein subcellular localization prediction, Cell biology, Transport protein, Ectomycorrhiza, Protein Transport, In situ hybridization, 010606 plant biology & botany
Abstract

International audience; Mycorrhizal exchange of nutrients between fungi and host plants involves a specialization and polarization of the fungal plasma membrane adapted for the uptake from the soil and for secretion of nutrient ions towards root cells. In addition to the current progress in identification of membrane transport systems of both symbiotic partners, data concerning the transcriptional and translational regulation of these proteins are needed to elucidate their role for symbiotic functions. To answer whether the formerly described Pi-dependent expression of the phosphate transporter HcPT1.1 from Hebeloma cylindrosporum is the result of its promoter activity, we introduced promoter-EGFP fusion constructs in the fungus by Agrotransformation. Indeed, HcPT1.1 expression in pure fungal cultures quantified and visualized by EGFP under control of the HcPT1.1 promoter was dependent on external Pi concentrations, low Pi stimulating the expression. Furthermore, to study expression and localization of the phosphate transporter HcPT1.1 in symbiotic conditions, presence of transcripts and proteins was analyzed by the in situ hybridization technique as well as by immunostaining of proteins. In ectomycorrhiza, expression of the phosphate transporter was clearly enhanced by Pi-shortage indicating its role in Pi nutrition in the symbiotic association. Transcripts were detected in external hyphae and in the hyphal mantle, proteins in addition also within the Hartig net. Exploiting the transformable fungus H. cylindrosporum, Pi-dependent expression of the fungal transporter HcPT1.1 as result from its promoter activity as well as transcript and protein localization in ectomycorrhizal symbiosis are shown.

Published
2013
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20. Arabidopsis WAT1 is a vacuolar auxin transport facilitator required for auxin homoeostasis

Author

Kris Morreel, Jean-Pierre Renou, Enrico Martinoia, Deborah Goffner, Urs Fischer, Stephanie Pfrunder, Benoît Lacombe, Philippe Ranocha, Yves Martinez, Claire Corratgé-Faillie, Jean-Baptiste Thibaud, Ondřej Novák, Judith Felten, Oana Dima, Karin Ljung, Réka Nagy, Wout Boerjan, Xu Jin, University of Zurich, Goffner, Deborah, Surfaces Cellulaires et Signalisation chez les Végétaux (SCSV), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS), Department of Plant Systems Biology, Ghent University [Belgium] (UGENT)-Department of Plant Biotechnology and Bioinformatics, Institut of Plant Biology, University of Zürich [Zürich] (UZH), Umea Plant Science Center, Swedish University of Agricultural Sciences (SLU)-Department of Forest Genetics and Plant Physiology, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Institut de Recherche en Horticulture et Semences (IRHS), Université d'Angers (UA)-Institut National de la Recherche Agronomique (INRA)-AGROCAMPUS OUEST, Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Universiteit Gent = Ghent University [Belgium] (UGENT)-Department of Plant Biotechnology and Bioinformatics, Universität Zürich [Zürich] = University of Zurich (UZH), Umea Plant Science Center (UPSC), Department of Forest Genetics and Plant Physiology, Swedish University of Agricultural Sciences (SLU)-Swedish University of Agricultural Sciences (SLU), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)

Subjects
0106 biological sciences, Arabidopsis, General Physics and Astronomy, Vacuole, 580 Plants (Botany), hormone auxin, plasma membrane, 01 natural sciences, Xenopus laevis, Plant Growth Regulators, 10126 Department of Plant and Microbial Biology, Cell Wall, Gene Expression Regulation, Plant, Homeostasis, Gene Regulatory Networks, heterocyclic compounds, évolution, DISSECTION, chemistry.chemical_classification, 0303 health sciences, Multidisciplinary, biology, BINDING PROTEIN-1, food and beverages, 3100 General Physics and Astronomy, Cell biology, Fiber cell, dissection, plante, endoplasmic reticulum membrane, Plant hormone, plant development, Secondary cell wall, Intracellular, WALLS ARE THIN1, 1600 General Chemistry, Saccharomyces cerevisiae, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, ROOT, Auxin, 1300 General Biochemistry, Genetics and Molecular Biology, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, Animals, PLANTS, binding protein 1, polar transport, plant, gene, identification, thaliana, root, transcriptome, evolution, TRANSCRIPTOME, 030304 developmental biology, Endoplasmic reticulum membrane, THALIANA, POLAR TRANSPORT, IDENTIFICATION, Indoleacetic Acids, Arabidopsis Proteins, fungi, arabidopsis thaliana, auxin transport, Biology and Life Sciences, Membrane Transport Proteins, Biological Transport, General Chemistry, biology.organism_classification, GENE, EVOLUTION, racine, chemistry, Mutation, Vacuoles, 010606 plant biology & botany
Abstract

The plant hormone auxin (indole-3-acetic acid, IAA) has a crucial role in plant development. Its spatiotemporal distribution is controlled by a combination of biosynthetic, metabolic and transport mechanisms. Four families of auxin transporters have been identified that mediate transport across the plasma or endoplasmic reticulum membrane. Here we report the discovery and the functional characterization of the first vacuolar auxin transporter. We demonstrate that WALLS ARE THIN1 (WAT1), a plant-specific protein that dictates secondary cell wall thickness of wood fibres, facilitates auxin export from isolated Arabidopsis vacuoles in yeast and in Xenopus oocytes. We unambiguously identify IAA and related metabolites in isolated Arabidopsis vacuoles, suggesting a key role for the vacuole in intracellular auxin homoeostasis. Moreover, local auxin application onto wat1 mutant stems restores fibre cell wall thickness. Our study provides new insight into the complexity of auxin transport in plants and a means to dissect auxin function during fibre differentiation., The plant hormone auxin is essential for plant development and growth and is transported across cellular membranes via specialized transporter proteins. In this study, Ranocha et al. identify the first vacuolar auxin transporter, WAT1, suggesting an involvement of the vacuole in auxin signalling.

Published
2013
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21. ABA transport and transporters

Author

Benoît Lacombe, Sophie Léran, Gabriel Krouk, Alain Gojon, Claire Corratgé-Faillie, Yann Boursiac, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut National de la Recherche Agronomique (CJS PhD Fellowship), Centre National de la Recherche Scientifique [PEPS Bio-Math-Info-2012-2013-Super-RegNet], Agence Nationale de la Recherche [ANR-11-JSV6-002-01-NUTSE, ANR-11-PDOC-020-01-NITRONET], Agropolis Fondation (RHIZOPOLIS grant) [07024], Region Languedoc-Roussillon (Chercheur d'Avenir), and Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0106 biological sciences, [SDV]Life Sciences [q-bio], Anion Transport Proteins, Arabidopsis, Biological Transport, Active, Plant Science, plasma membrane, 01 natural sciences, nitrogen, 03 medical and health sciences, chemistry.chemical_compound, Receptor, Abscisic acid, 030304 developmental biology, 0303 health sciences, biology, Arabidopsis Proteins, organic chemicals, Abscisic acid metabolism, fungi, food and beverages, Membrane Transport Proteins, Transporter, Nitrate Transporters, Metabolism, biology.organism_classification, Biochemistry, chemistry, transport, ATP-Binding Cassette Transporters, Signal transduction, 010606 plant biology & botany, Abscisic Acid, Signal Transduction
Abstract

International audience; Abscisic acid (ABA) metabolism, perception, and transport form a triptych allowing higher plants to use ABA as a signaling molecule. The molecular bases of ABA metabolism are now well described and, over the past few years, several ABA receptors have been discovered. Although ABA transport has long been demonstrated in planta, the first breakthroughs in identifying plasma membrane-localized ABA transporters came in 2010, with the identification of two ATP-binding cassette (ABC) proteins. More recently, two ABA transporters in the nitrate transporter 1/peptide transporter (NRT1/PTR) family have been identified. In this review, we discuss the role of these different ABA transporters and examine the scientific impact of their identification. Given that the NRT1/PTR family is involved in the transport of nitrogen (N) compounds, further work should determine whether an interaction between ABA and N signaling or nutrition occurs.

Published
2012
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22. Calcium-dependent modulation and plasma membrane targeting of the AKT2 potassium channel by the CBL4/CIPK6 calcium sensor/protein kinase complex

Author

Jörg Kudla, Ingo Dreyer, Kenji Hashimoto, Claire Corratgé-Faillie, Jean-Baptiste Thibaud, Pawel Gajdanowicz, Jan Niklas Offenborn, François Pascaud, Christian Eckert, Katrin Held, Benoît Lacombe, Institut für Biologie und Biotechnologie der Pflanzen, Westfälische Wilhelms-Universität Münster (WWU), Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Institut für Biochemie und Biologie, Universität Potsdam, Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), and Universität Potsdam - Abteilung Molekularbiologie

Subjects
0106 biological sciences, Protein kinase complex, Potassium Channels, Arabidopsis, Biology, Endoplasmic Reticulum, 01 natural sciences, SK channel, 03 medical and health sciences, Palmitoylation, Gene Expression Regulation, Plant, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, ASK1, Phosphorylation, Protein kinase A, Molecular Biology, Institut für Biochemie und Biologie, 030304 developmental biology, Myristoylation, 0303 health sciences, Arabidopsis Proteins, Endoplasmic reticulum, Cell Membrane, protein kinase, Cell Biology, Potassium channel, Cell biology, Protein Transport, Phenotype, Calcium, Original Article, calcium sensor, Protein Kinases, signal transduction, 010606 plant biology & botany, potassium channel
Abstract

International audience; Potassium (K(+)) channel function is fundamental to many physiological processes. However, components and mechanisms regulating the activity of plant K(+) channels remain poorly understood. Here, we show that the calcium (Ca(2+)) sensor CBL4 together with the interacting protein kinase CIPK6 modulates the activity and plasma membrane (PM) targeting of the K(+) channel AKT2 from Arabidopsis thaliana by mediating translocation of AKT2 to the PM in plant cells and enhancing AKT2 activity in oocytes. Accordingly, akt2, cbl4 and cipk6 mutants share similar developmental and delayed flowering phenotypes. Moreover, the isolated regulatory C-terminal domain of CIPK6 is sufficient for mediating CBL4- and Ca(2+)-dependent channel translocation from the endoplasmic reticulum membrane to the PM by a novel targeting pathway that is dependent on dual lipid modifications of CBL4 by myristoylation and palmitoylation. Thus, we describe a critical mechanism of ion-channel regulation where a Ca(2+) sensor modulates K(+) channel activity by promoting a kinase interaction-dependent but phosphorylation-independent translocation of the channel to the PM.

Published
2011
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23. Two Ca2+-dependent protein kinases (cpks) oppositely regulate a ubiquitous k+ channel in arabidopsis thaliana

Author

Elsa Ronzier, claire corratgé-faillie, Frédéric Sanchez, Christian Brière, Jean-Baptiste Thibaud, Tou Cheu Xiong, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Equipe Transport ionique chez les céréales et adaptation à l’environnement (TICER), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Recherche en Sciences Végétales (LRSV), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Centre National de la Recherche Scientifique (CNRS), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées, Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), and Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)

Subjects
shaker potassium channel, xenopus oocyte, transport phloème, fungi, stomata, arabidopsis thaliana, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, stomate, canaux potassiques shaker, guard cell, cellule de garde
Abstract

In plant cells, as regards both the electrical polarization of the plasma-membrane and the osmotic homeostasis, voltage-gated K + channels participate in transduction chains, often downstream Ca 2+ signals. Thus, worth considering is, among the range of post-translational modifications, that potentially regulate their activity, the phosphorylation of these so-called Shaker channels by Ca 2+ -dependent protein kinases (CPKs). We investigated the interactions between these two families of proteins by a combination of techniques: heterologous expression in Xenopus oocytes followed by electrophysiology recordings, in planta interaction by FRET-FLIM imaging, in vitro phosphorylation study using channel-derived peptide arrays, looking for overlap of expression patterns and characterization of gain- or loss-of-function mutants. Here we have identified several CPKs targeting and regulating the Shaker channel activities. Interestingly, we found that CPK13, a Ca 2+ -insensitive CPK, and CPK6, a Ca 2+ -strictly dependent CPK, respectively inhibits and activates the same Shaker channel, KAT2. In the context of (i) the large expression pattern of KAT2 (e.g. in leaf vasculature and in guard cells) and (ii) the documented ability of KAT2 to form heteromeric channels with the Shaker sub‐units AKT2 and KAT1, we shall discuss the potential implication of KAT2 regulation by CPKs in phloem and stomata physiology.

24. Regulation by Calcium-dependent protein kinases of Skaker channels expressed in guard cells

Author

Elsa Ronzier, Claire Corratgé-Faillie, Frédéric Sanchez, Jean-Baptiste THIBAUD, Tou Cheu Xiong, Biochimie et Physiologie Moléculaire des Plantes (BPMP), Université de Montpellier (UM)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Centre international d'études supérieures en sciences agronomiques (Montpellier SupAgro)-Institut National de la Recherche Agronomique (INRA)-Institut national d’études supérieures agronomiques de Montpellier (Montpellier SupAgro), and Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)

Subjects
arabidopsis, protéine kinase, shaker potassium channel, xenopus oocyte, fungi, [SDV.BV]Life Sciences [q-bio]/Vegetal Biology, canaux potassiques shaker, guard cell, cellule de garde, absorption du calcium
Abstract

Stomatal movements result from osmotically-driven fluxes of water, which follow massive exchanges of solutes, including K+ ions, between guard cells and surrounding tissues. These exchanges involve a set of dedicated transport systems, controlled through signalling pathways, including Ca2+-dependent ones. For example, Ca2+ signals have been reported to lead to stomatal closure through the inhibition of inward and activation of outward K+ channels. This could be mediated in particular by Ca2+-dependent kinases targeting these channels.Here, we addressed the post-translational regulation of three K+-selective voltage-gated channels expressed in Arabidopsis guard cells (inwardly-rectifying KAT1 and KAT2, and outwardly-rectifying GORK) by Ca2+-dependent protein kinases (CPKs). Of the 34 CPK genes inArabidopsis thaliana, some CPK are thought to be expressed in guard cells. Seven of these CPKs were co-expressed in Xenopus oocytes with the three studied Shaker channels, and their influence on K+ transport was examined with a classical voltage-clamp technique. Pinpointed interactions were checked by on-chip phosphorylation assays on peptide arrays designed from the channel primary sequence. This enabled us to list a number of candidate phosphorylation sites and, subsequently, to design mutant channels expected to display CPK-insensitive of CPKphosphorylated phenotype. The functional characterisation of these mutant channels and the phenotype of plants overexpressing CPKs will be presented.

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