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1.
Plant J ; 117(1): 264-279, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37844131

RESUMO

Soil water uptake by roots is a key component of plant water homeostasis contributing to plant growth and survival under ever-changing environmental conditions. The water transport capacity of roots (root hydraulic conductivity; Lpr ) is mostly contributed by finely regulated Plasma membrane Intrinsic Protein (PIP) aquaporins. In this study, we used natural variation of Arabidopsis for the identification of quantitative trait loci (QTLs) contributing to Lpr . Using recombinant lines from a biparental cross (Cvi-0 x Col-0), we show that the gene encoding class 2 Sucrose-Non-Fermenting Protein kinase 2.4 (SnRK2.4) in Col-0 contributes to >30% of Lpr by enhancing aquaporin-dependent water transport. At variance with the inactive and possibly unstable Cvi-0 SnRK2.4 form, the Col-0 form interacts with and phosphorylates the prototypal PIP2;1 aquaporin at Ser121 and stimulates its water transport activity upon coexpression in Xenopus oocytes and yeast cells. Activation of PIP2;1 by Col-0 SnRK2.4 in yeast also requires its protein kinase activity and can be counteracted by clade A Protein Phosphatases 2C. SnRK2.4 shows all hallmarks to be part of core abscisic acid (ABA) signaling modules. Yet, long-term (>3 h) inhibition of Lpr by ABA possibly involves a SnRK2.4-independent inhibition of PIP2;1. SnRK2.4 also promotes stomatal aperture and ABA-induced inhibition of primary root growth. The study identifies a key component of Lpr and sheds new light on the functional overlap and specificity of SnRK2.4 with respect to other ABA-dependent or independent SnRK2s.


Assuntos
Aquaporinas , Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas Quinases/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Saccharomyces cerevisiae/metabolismo , Ácido Abscísico/farmacologia , Ácido Abscísico/metabolismo , Fosforilação , Aquaporinas/genética , Aquaporinas/metabolismo , Água/metabolismo
2.
Plant Cell ; 32(2): 508-524, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31776233

RESUMO

Iron (Fe) is an essential micronutrient for plant growth and development. Any defects in the maintenance of Fe homeostasis will alter plant productivity and the quality of their derived products. In Arabidopsis (Arabidopsis thaliana), the transcription factor ILR3 plays a central role in controlling Fe homeostasis. In this study, we identified bHLH121 as an ILR3-interacting transcription factor. Interaction studies showed that bHLH121 also interacts with the three closest homologs of ILR3 (i.e., basic-helix-loop-helix 34 [bHLH34], bHLH104, and bHLH115). bhlh121 loss-of-function mutants displayed severe defects in Fe homeostasis that could be reverted by exogenous Fe supply. bHLH121 acts as a direct transcriptional activator of key genes involved in the Fe regulatory network, including bHLH38, bHLH39, bHLH100, bHLH101, POPEYE, BRUTUS, and BRUTUS LIKE1, as well as IRONMAN1 and IRONMAN2 In addition, bHLH121 is necessary for activating the expression of transcription factor gene FIT in response to Fe deficiency via an indirect mechanism. bHLH121 is expressed throughout the plant body, and its expression is not affected by Fe availability. By contrast, Fe availability affects the cellular localization of bHLH121 protein in roots. Altogether, these data show that bHLH121 is a regulator of Fe homeostasis that acts upstream of FIT in concert with ILR3 and its closest homologs.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Homeostase/fisiologia , Ferro/metabolismo , Fatores de Transcrição/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Fatores de Transcrição de Zíper de Leucina e Hélice-Alça-Hélix Básicos , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Regulação da Expressão Gênica de Plantas , Técnicas de Inativação de Genes , Redes Reguladoras de Genes , Homeostase/genética , Hidroponia , Proteínas Nucleares , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Fatores de Transcrição/genética , Transcriptoma , Ubiquitina-Proteína Ligases
3.
Physiol Rev ; 95(4): 1321-58, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26336033

RESUMO

Aquaporins are membrane channels that facilitate the transport of water and small neutral molecules across biological membranes of most living organisms. In plants, aquaporins occur as multiple isoforms reflecting a high diversity of cellular localizations, transport selectivity, and regulation properties. Plant aquaporins are localized in the plasma membrane, endoplasmic reticulum, vacuoles, plastids and, in some species, in membrane compartments interacting with symbiotic organisms. Plant aquaporins can transport various physiological substrates in addition to water. Of particular relevance for plants is the transport of dissolved gases such as carbon dioxide and ammonia or metalloids such as boron and silicon. Structure-function studies are developed to address the molecular and cellular mechanisms of plant aquaporin gating and subcellular trafficking. Phosphorylation plays a central role in these two processes. These mechanisms allow aquaporin regulation in response to signaling intermediates such as cytosolic pH and calcium, and reactive oxygen species. Combined genetic and physiological approaches are now integrating this knowledge, showing that aquaporins play key roles in hydraulic regulation in roots and leaves, during drought but also in response to stimuli as diverse as flooding, nutrient availability, temperature, or light. A general hydraulic control of plant tissue expansion by aquaporins is emerging, and their role in key developmental processes (seed germination, emergence of lateral roots) has been established. Plants with genetically altered aquaporin functions are now tested for their ability to improve plant tolerance to stresses. In conclusion, research on aquaporins delineates ever expanding fields in plant integrative biology thereby establishing their crucial role in plants.


Assuntos
Aquaporinas/metabolismo , Plantas/metabolismo , Animais , Transporte Biológico/fisiologia , Humanos , Concentração de Íons de Hidrogênio , Estresse Fisiológico/fisiologia
4.
Plant Physiol ; 187(4): 2056-2070, 2021 12 04.
Artigo em Inglês | MEDLINE | ID: mdl-35235672

RESUMO

Plant water transport and its molecular components including aquaporins are responsive, across diverse time scales, to an extremely wide array of environmental and hormonal signals. These include water deficit and abscisic acid (ABA) but also more recently identified stimuli such as peptide hormones or bacterial elicitors. The present review makes an inventory of corresponding signalling pathways. It identifies some main principles, such as the central signalling role of ROS, with a dual function of aquaporins in water and hydrogen peroxide transport, the importance of aquaporin phosphorylation that is targeted by multiple classes of protein kinases, and the emerging role of lipid signalling. More studies including systems biology approaches are now needed to comprehend how plant water transport can be adjusted in response to combined stresses.


Assuntos
Aquaporinas/metabolismo , Transporte Biológico/efeitos dos fármacos , Membrana Celular/metabolismo , Reguladores de Crescimento de Plantas/metabolismo , Fenômenos Fisiológicos Vegetais/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Água/metabolismo , Redes e Vias Metabólicas
5.
Plant Cell ; 31(2): 417-429, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30674691

RESUMO

The circadian clock regulates plant tissue hydraulics to synchronize water supply with environmental cycles and thereby optimize growth. The circadian fluctuations in aquaporin transcript abundance suggest that aquaporin water channels play a role in these processes. Here, we show that hydraulic conductivity (K ros) of Arabidopsis (Arabidopsis thaliana) rosettes displays a genuine circadian rhythmicity with a peak around midday. Combined immunological and proteomic approaches revealed that phosphorylation at two C-terminal sites (Ser280, Ser283) of PLASMA MEMBRANE INTRINSIC PROTEIN 2;1 (AtPIP2;1), a major plasma membrane aquaporin in rosettes, shows circadian oscillations and is correlated with K ros Transgenic expression of phosphodeficient and phosphomimetic forms of this aquaporin indicated that AtPIP2;1 phosphorylation is necessary but not sufficient for K ros regulation. We investigated the supporting role of 14-3-3 proteins, which are known to interact with and regulate phosphorylated proteins. Individual knockout plants for five 14-3-3 protein isoforms expressed in rosettes lacked circadian activation of K ros Two of these [GRF4 (14-3-3Phi); GRF10 (14-3-3Epsilon)] showed direct interactions with AtPIP2;1 in the plant and upon coexpression in Xenopus laevis oocytes and activated AtPIP2;1, preferentially when the latter was phosphorylated at its two C-terminal sites. We propose that this regulatory mechanism assists in the activation of phosphorylated AtPIP2;1 during circadian regulation of K ros.


Assuntos
Proteínas 14-3-3/metabolismo , Aquaporinas/metabolismo , Folhas de Planta/metabolismo , Plantas Geneticamente Modificadas/metabolismo , Proteínas 14-3-3/genética , Aquaporinas/genética , Regulação da Expressão Gênica de Plantas/genética , Regulação da Expressão Gênica de Plantas/fisiologia , Fosforilação/genética , Fosforilação/fisiologia , Folhas de Planta/genética , Plantas Geneticamente Modificadas/genética , Proteômica/métodos
6.
Int J Mol Sci ; 23(4)2022 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-35216074

RESUMO

Osmotic stress can be detrimental to plants, whose survival relies heavily on proteomic plasticity. Protein ubiquitination is a central post-translational modification in osmotic-mediated stress. In this study, we used the K-Ɛ-GG antibody enrichment method integrated with high-resolution mass spectrometry to compile a list of 719 ubiquitinated lysine (K-Ub) residues from 450 Arabidopsis root membrane proteins (58% of which are transmembrane proteins), thereby adding to the database of ubiquitinated substrates in plants. Although no ubiquitin (Ub) motifs could be identified, the presence of acidic residues close to K-Ub was revealed. Our ubiquitinome analysis pointed to a broad role of ubiquitination in the internalization and sorting of cargo proteins. Moreover, the simultaneous proteome and ubiquitinome quantification showed that ubiquitination is mostly not involved in membrane protein degradation in response to short osmotic treatment but that it is putatively involved in protein internalization, as described for the aquaporin PIP2;1. Our in silico analysis of ubiquitinated proteins shows that two E2 Ub-conjugating enzymes, UBC32 and UBC34, putatively target membrane proteins under osmotic stress. Finally, we revealed a positive role for UBC32 and UBC34 in primary root growth under osmotic stress.


Assuntos
Arabidopsis/metabolismo , Arabidopsis/fisiologia , Pressão Osmótica/fisiologia , Raízes de Plantas/metabolismo , Raízes de Plantas/fisiologia , Ubiquitinação/fisiologia , Lisina/metabolismo , Proteínas de Membrana/metabolismo , Processamento de Proteína Pós-Traducional/fisiologia , Proteoma/metabolismo , Proteômica/métodos , Ubiquitina/metabolismo , Proteínas Ubiquitinadas/metabolismo
7.
J Biol Chem ; 295(6): 1727-1742, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-31911438

RESUMO

Proteins incorporating iron-sulfur (Fe-S) co-factors are required for a plethora of metabolic processes. Their maturation depends on three Fe-S cluster assembly machineries in plants, located in the cytosol, mitochondria, and chloroplasts. After de novo formation on scaffold proteins, transfer proteins load Fe-S clusters onto client proteins. Among the plastidial representatives of these transfer proteins, NFU2 and NFU3 are required for the maturation of the [4Fe-4S] clusters present in photosystem I subunits, acting upstream of the high-chlorophyll fluorescence 101 (HCF101) protein. NFU2 is also required for the maturation of the [2Fe-2S]-containing dihydroxyacid dehydratase, important for branched-chain amino acid synthesis. Here, we report that recombinant Arabidopsis thaliana NFU1 assembles one [4Fe-4S] cluster per homodimer. Performing co-immunoprecipitation experiments and assessing physical interactions of NFU1 with many [4Fe-4S]-containing plastidial proteins in binary yeast two-hybrid assays, we also gained insights into the specificity of NFU1 for the maturation of chloroplastic Fe-S proteins. Using bimolecular fluorescence complementation and in vitro Fe-S cluster transfer experiments, we confirmed interactions with two proteins involved in isoprenoid and thiamine biosynthesis, 1-hydroxy-2-methyl-2-(E)-butenyl-4-diphosphate synthase and 4-amino-5-hydroxymethyl-2-methylpyrimidine phosphate synthase, respectively. An additional interaction detected with the scaffold protein SUFD enabled us to build a model in which NFU1 receives its Fe-S cluster from the SUFBC2D scaffold complex and serves in the maturation of specific [4Fe-4S] client proteins. The identification of the NFU1 partner proteins reported here more clearly defines the role of NFU1 in Fe-S client protein maturation in Arabidopsis chloroplasts among other SUF components.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Cloroplastos/metabolismo , Proteínas Ferro-Enxofre/metabolismo , Plastídeos/metabolismo , Mapas de Interação de Proteínas , Complexo de Proteína do Fotossistema I/metabolismo , Ligação Proteica
8.
New Phytol ; 228(3): 1038-1054, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32463943

RESUMO

In Arabidopsis thaliana, NRT2.1 codes for a main component of the root nitrate high-affinity transport system. Previous studies revealed that post-translational regulation of NRT2.1 plays an important role in the control of root nitrate uptake and that one mechanism could correspond to NRT2.1 C-terminus processing. To further investigate this hypothesis, we produced transgenic plants with truncated forms of NRT2.1. This revealed an essential sequence for NRT2.1 activity, located between the residues 494 and 513. Using a phospho-proteomic approach, we found that this sequence contains one phosphorylation site, at serine 501, which can inactivate NRT2.1 function when mimicking the constitutive phosphorylation of this residue in transgenic plants. This phenotype could neither be explained by changes in abundance of NRT2.1 and NAR2.1, a partner protein of NRT2.1, nor by a lack of interaction between these two proteins. Finally, the relative level of serine 501 phosphorylation was found to be increased by ammonium nitrate in wild-type plants, leading to the inactivation of NRT2.1 and to a decrease in high affinity nitrate transport into roots. Altogether, these observations reveal a new and essential mechanism for the regulation of NRT2.1 activity.


Assuntos
Proteínas de Transporte de Ânions , Proteínas de Arabidopsis , Arabidopsis , Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Nitratos/metabolismo , Fosforilação , Proteínas de Plantas/metabolismo , Raízes de Plantas/metabolismo , Proteômica
9.
Plant Cell Environ ; 43(3): 760-774, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31759334

RESUMO

The mechanisms underlying the response and adaptation of plants to excess of trace elements are not fully described. Here, we analysed the importance of protein lysine methylation for plants to cope with cadmium. We analysed the effect of cadmium on lysine-methylated proteins and protein lysine methyltransferases (KMTs) in two cadmium-sensitive species, Arabidopsis thaliana and A. lyrata, and in three populations of A. halleri with contrasting cadmium accumulation and tolerance traits. We showed that some proteins are differentially methylated at lysine residues in response to Cd and that a few genes coding KMTs are regulated by cadmium. Also, we showed that 9 out of 23 A. thaliana mutants disrupted in KMT genes have a tolerance to cadmium that is significantly different from that of wild-type seedlings. We further characterized two of these mutants, one was knocked out in the calmodulin lysine methyltransferase gene and displayed increased tolerance to cadmium, and the other was interrupted in a KMT gene of unknown function and showed a decreased capacity to cope with cadmium. Together, our results showed that lysine methylation of non-histone proteins is impacted by cadmium and that several methylation events are important for modulating the response of Arabidopsis plants to cadmium stress.


Assuntos
Adaptação Fisiológica , Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Arabidopsis/fisiologia , Cádmio/toxicidade , Lisina/metabolismo , Estresse Fisiológico , Adaptação Fisiológica/efeitos dos fármacos , Adaptação Fisiológica/genética , Sequência de Aminoácidos , Arabidopsis/efeitos dos fármacos , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , Mutação/genética , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Raízes de Plantas/crescimento & desenvolvimento , Estresse Fisiológico/efeitos dos fármacos , Estresse Fisiológico/genética
10.
J Exp Bot ; 71(14): 4171-4187, 2020 07 06.
Artigo em Inglês | MEDLINE | ID: mdl-32240305

RESUMO

Iron-sulfur (Fe-S) proteins have critical functions in plastids, notably participating in photosynthetic electron transfer, sulfur and nitrogen assimilation, chlorophyll metabolism, and vitamin or amino acid biosynthesis. Their maturation relies on the so-called SUF (sulfur mobilization) assembly machinery. Fe-S clusters are synthesized de novo on a scaffold protein complex and then delivered to client proteins via several transfer proteins. However, the maturation pathways of most client proteins and their specificities for transfer proteins are mostly unknown. In order to decipher the proteins interacting with the Fe-S cluster transfer protein NFU2, one of the three plastidial representatives found in Arabidopsis thaliana, we performed a quantitative proteomic analysis of shoots, roots, and seedlings of nfu2 plants, combined with NFU2 co-immunoprecipitation and binary yeast two-hybrid experiments. We identified 14 new targets, among which nine were validated in planta using a binary bimolecular fluorescence complementation assay. These analyses also revealed a possible role for NFU2 in the plant response to desiccation. Altogether, this study better delineates the maturation pathways of many chloroplast Fe-S proteins, considerably extending the number of NFU2 clients. It also helps to clarify the respective roles of the three NFU paralogs NFU1, NFU2, and NFU3.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Proteínas Ferro-Enxofre , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Cloroplastos/metabolismo , Proteínas Ferro-Enxofre/genética , Proteômica
11.
Int J Mol Sci ; 21(21)2020 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-33143294

RESUMO

Iron-sulfur (Fe-S) proteins play critical functions in plants. Most Fe-S proteins are synthetized in the cytosol as apo-proteins and the subsequent Fe-S cluster incorporation relies on specific protein assembly machineries. They are notably formed by a scaffold complex, which serves for the de novo Fe-S cluster synthesis, and by transfer proteins that insure cluster delivery to apo-targets. However, scarce information is available about the maturation pathways of most plastidial Fe-S proteins and their specificities towards transfer proteins of the associated SUF machinery. To gain more insights into these steps, the expression and protein localization of the NFU1, NFU2, and NFU3 transfer proteins were analyzed in various Arabidopsis thaliana organs and tissues showing quite similar expression patterns. In addition, quantitative proteomic analysis of an nfu3 loss-of-function mutant allowed to propose novel potential client proteins for NFU3 and to show that the protein accumulation profiles and thus metabolic adjustments differ substantially from those established in the nfu2 mutant. By clarifying the respective roles of the three plastidial NFU paralogs, these data allow better delineating the maturation process of plastidial Fe-S proteins.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Proteínas Ferro-Enxofre/metabolismo , Plastídeos/metabolismo , Proteoma/metabolismo , Arabidopsis/crescimento & desenvolvimento , Proteoma/análise
12.
Plant Cell Environ ; 42(6): 1788-1801, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30767240

RESUMO

The absorption of soil water by roots allows plants to maintain their water status. At the endodermis, water transport can be affected by initial formation of a Casparian strip and further deposition of suberin lamellas and regulated by the function of aquaporins. Four Casparian strip membrane domain protein-like (CASPL; CASPL1B1, CASPL1B2, CASPL1D1, and CASPL1D2) were previously shown to interact with PIP2;1. The present work shows that CASPL1B1, CASPL1B2, and CASPL1D2 are exclusively expressed in suberized endodermal cells, suggesting a cell-specific role in suberization and/or water transport regulation. When compared with wild-type plants, and by contrast to caspl1b1*caspl1b2 double loss of function, caspl1d1*caspl1d2 double mutants showed, in some control or NaCl stress experiments and not upon abscisic acid (ABA) treatment, a weak enlargement of the continuous suberization zone. None of the mutants showed root hydraulic conductivity (Lpr ) phenotype, whether in control, NaCl, or ABA treatment conditions. The data suggest a slight negative role for CASPL1D1 and CASPL1D2 in suberization under control or salt stress conditions, with no major impact on whole root transport functions. At the molecular level, CASPL1B1 was able to physically interact with PIP2;1 and potentially could influence the regulation of aquaporins by acting on their phosphorylated form.


Assuntos
Aquaporinas/metabolismo , Transporte Biológico/fisiologia , Parede Celular/metabolismo , Ácido Abscísico/metabolismo , Animais , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis , Regulação da Expressão Gênica de Plantas , Lipídeos , Proteínas de Membrana , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Estresse Psicológico , Água/metabolismo , Xenopus/genética , Xenopus/metabolismo
13.
Mol Cell Proteomics ; 15(11): 3473-3487, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27609422

RESUMO

PIP1;2 and PIP2;1 are aquaporins that are highly expressed in roots and bring a major contribution to root water transport and its regulation by hormonal and abiotic factors. Interactions between cellular proteins or with other macromolecules contribute to forming molecular machines. Proteins that molecularly interact with PIP1;2 and PIP2;1 were searched to get new insights into regulatory mechanisms of root water transport. For that, a immuno-purification strategy coupled to protein identification and quantification by mass spectrometry (IP-MS) of PIPs was combined with data from the literature, to build thorough PIP1;2 and PIP2;1 interactomes, sharing about 400 interacting proteins. Such interactome revealed PIPs to behave as a platform for recruitment of a wide range of transport activities and provided novel insights into regulation of PIP cellular trafficking by osmotic and oxidative treatments. This work also pointed a role of lipid signaling in PIP function and enhanced our knowledge of protein kinases involved in PIP regulation. In particular we show that 2 members of the receptor-like kinase (RLK) family (RKL1 (At1g48480) and Feronia (At3g51550)) differentially modulate PIP activity through distinct molecular mechanisms. The overall work opens novel perspectives in understanding PIP regulatory mechanisms and their role in adjustment of plant water status.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Fosfotransferases/metabolismo , Proteínas Quinases/metabolismo , Bases de Dados de Proteínas , Regulação da Expressão Gênica de Plantas , Espectrometria de Massas , Raízes de Plantas/metabolismo , Mapas de Interação de Proteínas
14.
Plant Cell ; 25(3): 1029-39, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23532070

RESUMO

The water status of plant leaves depends on the efficiency of the water supply, from the vasculature to inner tissues. This process is under hormonal and environmental regulation and involves aquaporin water channels. In Arabidopsis thaliana, the rosette hydraulic conductivity (Kros) is higher in darkness than it is during the day. Knockout plants showed that three plasma membrane intrinsic proteins (PIPs) sharing expression in veins (PIP1;2, PIP2;1, and PIP2;6) contribute to rosette water transport, and PIP2;1 can fully account for Kros responsiveness to darkness. Directed expression of PIP2;1 in veins of a pip2;1 mutant was sufficient to restore Kros. In addition, a positive correlation, in both wild-type and PIP2;1-overexpressing plants, was found between Kros and the osmotic water permeability of protoplasts from the veins but not from the mesophyll. Thus, living cells in veins form a major hydraulic resistance in leaves. Quantitative proteomic analyses showed that light-dependent regulation of Kros is linked to diphosphorylation of PIP2;1 at Ser-280 and Ser-283. Expression in pip2;1 of phosphomimetic and phosphorylation-deficient forms of PIP2;1 demonstrated that phosphorylation at these two sites is necessary for Kros enhancement under darkness. These findings establish how regulation of a single aquaporin isoform in leaf veins critically determines leaf hydraulics.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/efeitos da radiação , Regulação da Expressão Gênica de Plantas , Luz , Folhas de Planta/metabolismo , Aquaporinas/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Transporte Biológico , Membrana Celular/genética , Membrana Celular/metabolismo , Escuridão , Células do Mesofilo/metabolismo , Osmose , Fosforilação , Folhas de Planta/genética , Folhas de Planta/efeitos da radiação , Transpiração Vegetal , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/metabolismo , Plantas Geneticamente Modificadas/efeitos da radiação , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Água/metabolismo
15.
Biochim Biophys Acta ; 1840(5): 1574-82, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24246957

RESUMO

BACKGROUND: Aquaporins are membrane channels that facilitate the transport of water and small neutral molecules across biological membranes of most living organisms. SCOPE OF REVIEW: Here, we present comprehensive insights made on plant aquaporins in recent years, pointing to their molecular and physiological specificities with respect to animal or microbial counterparts. MAJOR CONCLUSIONS: In plants, aquaporins occur as multiple isoforms reflecting a high diversity of cellular localizations and various physiological substrates in addition to water. Of particular relevance for plants is the transport by aquaporins of dissolved gases such as carbon dioxide or metalloids such as boric or silicic acid. The mechanisms that determine the gating and subcellular localization of plant aquaporins are extensively studied. They allow aquaporin regulation in response to multiple environmental and hormonal stimuli. Thus, aquaporins play key roles in hydraulic regulation and nutrient transport in roots and leaves. They contribute to several plant growth and developmental processes such as seed germination or emergence of lateral roots. GENERAL SIGNIFICANCE: Plants with genetically altered aquaporin functions are now tested for their ability to improve plant resistance to stresses. This article is part of a Special Issue entitled Aquaporins.


Assuntos
Aquaporinas/fisiologia , Fenômenos Fisiológicos Vegetais , Plantas/metabolismo , Transporte Biológico , Água/metabolismo
16.
Plant Cell Environ ; 38(7): 1312-20, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25366820

RESUMO

The hydraulic conductivity of plant roots (Lp(r)) is determined in large part by the activity of aquaporins. Mechanisms occurring at the post-translational level, in particular phosphorylation of aquaporins of the plasma membrane intrinsic protein 2 (PIP2) subfamily, are thought to be of critical importance for regulating root water transport. However, knowledge of protein kinases and phosphatases acting on aquaporin function is still scarce. In the present work, we investigated the Lp(r) of knockout Arabidopsis plants for four Ca(2+)-dependent protein kinases. cpk7 plants showed a 30% increase in Lp(r) because of a higher aquaporin activity. A quantitative proteomic analysis of wild-type and cpk7 plants revealed that PIP gene expression and PIP protein quantity were not correlated and that CPK7 has no effect on PIP2 phosphorylation. In contrast, CPK7 exerts a negative control on the cellular abundance of PIP1s, which likely accounts for the higher Lp(r) of cpk7. In addition, this study revealed that the cellular amount of a few additional proteins including membrane transporters is controlled by CPK7. The overall work provides evidence for CPK7-dependent stability of specific membrane proteins.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimologia , Proteínas Quinases/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Aquaporinas/genética , Arabidopsis/genética , Arabidopsis/fisiologia , Proteínas de Arabidopsis/genética , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Técnicas de Inativação de Genes , Mutagênese Insercional , Fosforilação , Raízes de Plantas/enzimologia , Raízes de Plantas/genética , Raízes de Plantas/fisiologia , Transpiração Vegetal/fisiologia , Plantas Geneticamente Modificadas , Proteínas Quinases/genética , Proteínas Serina-Treonina Quinases/genética , Proteômica , Água/fisiologia
17.
Mol Cell Proteomics ; 12(12): 3886-97, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24056735

RESUMO

In plants, aquaporins play a crucial role in regulating root water transport in response to environmental and physiological cues. Controls achieved at the post-translational level are thought to be of critical importance for regulating aquaporin function. To investigate the general molecular mechanisms involved, we performed, using the model species Arabidopsis, a comprehensive proteomic analysis of root aquaporins in a large set of physiological contexts. We identified nine physiological treatments that modulate root hydraulics in time frames of minutes (NO and H2O2 treatments), hours (mannitol and NaCl treatments, exposure to darkness and reversal with sucrose, phosphate supply to phosphate-starved roots), or days (phosphate or nitrogen starvation). All treatments induced inhibition of root water transport except for sucrose supply to dark-grown plants and phosphate resupply to phosphate-starved plants, which had opposing effects. Using a robust label-free quantitative proteomic methodology, we identified 12 of 13 plasma membrane intrinsic protein (PIP) aquaporin isoforms, 4 of the 10 tonoplast intrinsic protein isoforms, and a diversity of post-translational modifications including phosphorylation, methylation, deamidation, and acetylation. A total of 55 aquaporin peptides displayed significant changes after treatments and enabled the identification of specific and as yet unknown patterns of response to stimuli. The data show that the regulation of PIP and tonoplast intrinsic protein abundance was involved in response to a few treatments (i.e. NaCl, NO, and nitrate starvation), whereas changes in the phosphorylation status of PIP aquaporins were positively correlated to changes in root hydraulic conductivity in the whole set of treatments. The identification of in vivo deamidated forms of aquaporins and their stimulus-induced changes in abundance may reflect a new mechanism of aquaporin regulation. The overall work provides deep insights into the in vivo post-translational events triggered by environmental constraints and their possible role in regulating plant water status.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Regulação da Expressão Gênica de Plantas , Raízes de Plantas/metabolismo , Processamento de Proteína Pós-Traducional , Água/metabolismo , Acetilação , Amidas/metabolismo , Sequência de Aminoácidos , Aquaporinas/genética , Arabidopsis/efeitos dos fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Escuridão , Variação Genética , Peróxido de Hidrogênio/farmacologia , Manitol/farmacologia , Metilação , Anotação de Sequência Molecular , Dados de Sequência Molecular , Óxido Nítrico/farmacologia , Fosfatos/farmacologia , Fosforilação , Raízes de Plantas/efeitos dos fármacos , Raízes de Plantas/genética , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Cloreto de Sódio/farmacologia , Estresse Fisiológico , Sacarose/farmacologia
18.
Proteomics ; 14(9): 1058-70, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24616185

RESUMO

An excess of NaCl in the soil is detrimental for plant growth. It interferes with mineral nutrition and water uptake and leads to accumulation of toxic ions in the plant. Understanding the response of roots to NaCl stress may facilitate the development of crops with increased tolerance to this and other stresses. Since controls achieved at the posttranslational level are of critical importance for regulating protein function, the present work used a robust label-free quantitative proteomic methodology to quantify phosphorylation events that affect root membrane proteins in Arabidopsis, in response to short-term (up to 2 h) NaCl treatments. This work identified 302 proteotypic phosphopeptides including 77 novel phosphorylated sites. NaCl treatment significantly altered the abundance of 74 phosphopeptides, giving novel insights into the regulation of major classes of membrane proteins, including ATPases, sodium transporters, and aquaporins. The data provide a unique access to phosphorylation reprogramming of ionic equilibrium in plant cells under NaCl stress. The use of predictive bioinformatic tools for kinase motifs suggested that root membrane proteins are substrates of cAMP-dependent protein kinase, cGMP-dependent protein kinase, and protein kinase C families, also called AGC kinases, arguing for an important role of lipid signaling in abiotic stress responses. It also pointed to cross-talks between protein kinase families during NaCl stress.


Assuntos
Proteínas de Arabidopsis/análise , Proteínas de Membrana/análise , Fosfoproteínas/análise , Raízes de Plantas/efeitos dos fármacos , Cloreto de Sódio/farmacologia , Estresse Fisiológico/efeitos dos fármacos , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Arabidopsis/fisiologia , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Fosforilação , Raízes de Plantas/química , Raízes de Plantas/metabolismo , Proteoma/análise , Proteoma/química , Proteoma/efeitos dos fármacos , Proteoma/metabolismo , Proteômica
19.
Traffic ; 12(4): 473-82, 2011 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-21182578

RESUMO

Plasma membrane intrinsic proteins (PIPs) are aquaporins that mediate water transport across the plant plasma membrane (PM). The present work addresses, using Arabidopsis AtPIP2;1 as a model, the mechanisms and significance of trafficking of newly synthesized PIPs from the endoplasmic reticulum (ER) to the Golgi apparatus. A functional diacidic export motif (Asp4-Val5-Glu6) was identified in the N-terminal tail of AtPIP2;1, using expression in transgenic Arabidopsis of site-directed mutants tagged with the green fluorescent protein (GFP). Confocal fluorescence imaging and a novel fluorescence recovery after photobleaching application based on the distinct diffusion of PM and intracellular AtPIP2;1-GFP forms revealed a retention in the ER of diacidic mutated forms, but with quantitative differences. Thus, the individual role of the two acidic Asp4 and Glu6 residues was established. In addition, expression in transgenic Arabidopsis of ER-retained AtPIP2;1-GFP constructs reduced the root hydraulic conductivity. Co-expression of AtPIP2;1-GFP and AtPIP1;4-mCherry constructs suggested that ER-retained AtPIP2;1-GFP may interact with other PIPs to hamper their trafficking to the PM, thereby contributing to inhibition of root cell hydraulic conductivity.


Assuntos
Aquaporinas/metabolismo , Proteínas de Arabidopsis/metabolismo , Retículo Endoplasmático/metabolismo , Motivos de Aminoácidos , Aquaporinas/química , Aquaporinas/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Membrana Celular/genética , Membrana Celular/metabolismo , Retículo Endoplasmático/genética , Regulação da Expressão Gênica de Plantas , Complexo de Golgi/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Proteínas de Membrana/química , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mutagênese Sítio-Dirigida , Raízes de Plantas/genética , Raízes de Plantas/metabolismo , Plantas Geneticamente Modificadas , Transporte Proteico , Água/metabolismo
20.
Annu Rev Plant Biol ; 59: 595-624, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-18444909

RESUMO

Aquaporins are channel proteins present in the plasma and intracellular membranes of plant cells, where they facilitate the transport of water and/or small neutral solutes (urea, boric acid, silicic acid) or gases (ammonia, carbon dioxide). Recent progress was made in understanding the molecular bases of aquaporin transport selectivity and gating. The present review examines how a wide range of selectivity profiles and regulation properties allows aquaporins to be integrated in numerous functions, throughout plant development, and during adaptations to variable living conditions. Although they play a central role in water relations of roots, leaves, seeds, and flowers, aquaporins have also been linked to plant mineral nutrition and carbon and nitrogen fixation.


Assuntos
Aquagliceroporinas/fisiologia , Desenvolvimento Vegetal , Fenômenos Fisiológicos Vegetais , Proteínas de Plantas/fisiologia , Transporte Biológico , Carbono/metabolismo , Membrana Celular/fisiologia , Temperatura Baixa , Hipóxia , Luz , Nitrogênio/metabolismo , Água/metabolismo
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