RESUMEN
Symbiotic nitrogen fixation boosts legume growth and production in nitrogen-poor soils. It has long been assumed that fixed nitrogen increases reproductive success, but until now, the regulatory mechanism was unknown. Here, we report a symbiotic flowering pathway that couples symbiotic and nutrient signals to the flowering induction pathway in legumes. We show that the symbiotic microRNA-microRNA172c (miR172c) and fixed nitrogen systemically and synergistically convey symbiotic and nutritional cues from roots to leaves to promote soybean (Glycine max) flowering. The combinations of symbiotic miR172c and local miR172c elicited by fixed nitrogen and development in leaves activate florigen-encoding FLOWERING LOCUS T (FT) homologs (GmFT2a/5a) by repressing TARGET OF EAT1-like 4a (GmTOE4a). Thus, FTs trigger reproductive development, which allows legumes to survive and reproduce under low-nitrogen conditions.
Asunto(s)
Glycine max , Fijación del Nitrógeno , Nitrógeno/metabolismo , Fijación del Nitrógeno/fisiología , Glycine max/genética , Simbiosis/fisiología , MicroARNs , ARN de Planta , Genes de PlantasRESUMEN
Calcium (Ca2+) signals are decoded by the Ca2+-sensor protein calmodulin (CaM) and are transduced to Ca2+/CaM-binding transcription factors to directly regulate gene expression necessary for acclimation responses in plants. The molecular mechanisms of Ca2+/CaM signal transduction processes and their functional significance remains enigmatic. Here we report a novel Ca2+/CaM signal transduction mechanism that allosterically regulates DNA-binding activity of GT2-LIKE 1 (GTL1), a transrepressor of STOMATAL DENSITY AND DISTRIBUTION 1 (SDD1), to repress stomatal development in response to water stress. We demonstrated that Ca2+/CaM interaction with the 2nd helix of the GTL1 N-terminal trihelix DNA-binding domain (GTL1N) destabilizes a hydrophobic core of GTL1N and allosterically inhibits 3rd helix docking to the SDD1 promoter, leading to osmotic stress-induced Ca2+/CaM-dependent activation (de-repression) of SDD1 expression. This resulted in GTL1-dependent repression of stomatal development in response to water-deficit stress. Together, our results demonstrate that a Ca2+/CaM-regulated transcriptional switch on a trihelix transrepressor directly transduces osmotic stress to repress stomatal development to improve plant water-use efficiency as an acclimation response.
Asunto(s)
Arabidopsis/metabolismo , Señalización del Calcio , Calcio/metabolismo , Calmodulina/metabolismo , Estomas de Plantas/crecimiento & desarrollo , Transcripción Genética , Agua/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/biosíntesis , Proteínas de Arabidopsis/genética , Calmodulina/genética , Estomas de Plantas/genética , Serina Endopeptidasas/biosíntesis , Serina Endopeptidasas/genéticaRESUMEN
Crop yield reduction as a consequence of increasingly severe climatic events threatens global food security. Genetic loci that ensure productivity in challenging environments exist within the germplasm of crops, their wild relatives and species that are adapted to extreme environments. Selective breeding for the combination of beneficial loci in germplasm has improved yields in diverse environments throughout the history of agriculture. An effective new paradigm is the targeted identification of specific genetic determinants of stress adaptation that have evolved in nature and their precise introgression into elite varieties. These loci are often associated with distinct regulation or function, duplication and/or neofunctionalization of genes that maintain plant homeostasis.
Asunto(s)
Adaptación Fisiológica/genética , Productos Agrícolas/química , Productos Agrícolas/genética , Ingeniería Genética , Plantas Modificadas Genéticamente/genética , Sitios de Carácter Cuantitativo , Estrés Fisiológico/genéticaRESUMEN
Arabidopsis (Arabidopsis thaliana) CYCLIN-DEPENDENT KINASE Ds (CDKDs) phosphorylate the C-terminal domain of the largest subunit of RNA polymerase II. Arabidopsis CYCLIN H;1 (CYCH;1) interacts with and activates CDKDs; however, the physiological function of CYCH;1 has not been determined. Here, we report that CYCH;1, which is localized to the nucleus, positively regulates blue light-induced stomatal opening. Reduced-function cych;1 RNA interference (cych;1 RNAi) plants exhibited a drought tolerance phenotype. CYCH;1 is predominantly expressed in guard cells, and its expression was substantially down-regulated by dehydration. Transpiration of intact leaves was reduced in cych;1 RNAi plants compared with the wild-type control in light but not in darkness. CYCH;1 down-regulation impaired blue light-induced stomatal opening but did not affect guard cell development or abscisic acid-mediated stomatal closure. Microarray and real-time polymerase chain reaction analyses indicated that CYCH;1 did not regulate the expression of abscisic acid-responsive genes or light-induced stomatal opening signaling determinants, such as MYB60, MYB61, Hypersensitive to red and blue1, and Protein phosphatase7. CYCH;1 down-regulation induced the expression of redox homeostasis genes, such as LIPOXYGENASE3 (LOX3), LOX4, ARABIDOPSIS GLUTATHIONE PEROXIDASE 7 (ATGPX7), EARLY LIGHT-INDUCIBLE PROTEIN1 (ELIP1), and ELIP2, and increased hydrogen peroxide production in guard cells. Furthermore, loss-of-function mutations in CDKD;2 or CDKD;3 did not affect responsiveness to drought stress, suggesting that CYCH;1 regulates the drought stress response in a CDKD-independent manner. We propose that CYCH;1 regulates blue light-mediated stomatal opening by controlling reactive oxygen species homeostasis.
Asunto(s)
Arabidopsis/fisiología , Ciclina H/metabolismo , Estomas de Plantas/fisiología , Especies Reactivas de Oxígeno/metabolismo , Estrés Fisiológico , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Ciclina H/genética , Quinasas Ciclina-Dependientes/genética , Quinasas Ciclina-Dependientes/metabolismo , Sequías , Regulación de la Expresión Génica de las Plantas , Luz , Mutación , Transpiración de Plantas , Plantas Modificadas Genéticamente , Interferencia de ARNRESUMEN
Transpiration and gas exchange occur through stomata. Thus, the control of stomatal aperture is important for the efficiency and regulation of water use, and for the response to drought. Here, we demonstrate that SIZ1-mediated endogenous salicylic acid (SA) accumulation plays an important role in stomatal closure and drought tolerance. siz1 reduced stomatal apertures. The reduced stomatal apertures of siz1 were inhibited by the application of peroxidase inhibitors, salicylhydroxamic acid and azide, which inhibits SA-dependent reactive oxygen species (ROS) production, but not by an NADPH oxidase inhibitor, diphenyl iodonium chloride, which inhibits ABA-dependent ROS production. Furthermore, the introduction of nahG into siz1, which reduces SA accumulation, restored stomatal opening. Stomatal closure is generally induced by water deficit. The siz1 mutation caused drought tolerance, whereas nahG siz1 suppressed the tolerant phenotype. Drought stresses also induced expression of SA-responsive genes, such as PR1 and PR2. Furthermore, other SA-accumulating mutants, cpr5 and acd6, exhibited stomatal closure and drought tolerance, and nahG suppressed the phenotype of cpr5 and acd6, as did siz1 and nahG siz1. Together, these results suggest that SIZ1 negatively affects stomatal closure and drought tolerance through the accumulation of SA.
Asunto(s)
Arabidopsis/fisiología , Ligasas/deficiencia , Reguladores del Crecimiento de las Plantas/fisiología , Estomas de Plantas/fisiología , Especies Reactivas de Oxígeno/metabolismo , Ácido Salicílico/metabolismo , Adaptación Fisiológica/genética , Adaptación Fisiológica/fisiología , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/fisiología , Deshidratación/fisiopatología , Ligasas/genética , Ligasas/fisiología , Datos de Secuencia MolecularRESUMEN
Diminishing global fresh water availability has focused research to elucidate mechanisms of water use in poplar, an economically important species. A GT-2 family trihelix transcription factor that is a determinant of water use efficiency (WUE), PtaGTL1 (GT-2 like 1), was identified in Populus tremula × P. alba (clone 717-IB4). Like other GT-2 family members, PtaGTL1 contains both N- and C-terminal trihelix DNA binding domains. PtaGTL1 expression, driven by the Arabidopsis thaliana AtGTL1 promoter, suppressed the higher WUE and drought tolerance phenotypes of an Arabidopsis GTL1 loss-of-function mutation (gtl1-4). Genetic suppression of gtl1-4 was associated with increased stomatal density due to repression of Arabidopsis STOMATAL DENSITY AND DISTRIBUTION1 (AtSDD1), a negative regulator of stomatal development. Electrophoretic mobility shift assays (EMSA) indicated that a PtaGTL1 C-terminal DNA trihelix binding fragment (PtaGTL1-C) interacted with an AtSDD1 promoter fragment containing the GT3 box (GGTAAA), and this GT3 box was necessary for binding. PtaGTL1-C also interacted with a PtaSDD1 promoter fragment via the GT2 box (GGTAAT). PtaSDD1 encodes a protein with 60% primary sequence identity with AtSDD1. In vitro molecular interaction assays were used to determine that Ca(2+)-loaded calmodulin (CaM) binds to PtaGTL1-C, which was predicted to have a CaM-interaction domain in the first helix of the C-terminal trihelix DNA binding domain. These results indicate that, in Arabidopsis and poplar, GTL1 and SDD1 are fundamental components of stomatal lineage. In addition, PtaGTL1 is a Ca(2+)-CaM binding protein, which infers a mechanism by which environmental stimuli can induce Ca(2+) signatures that would modulate stomatal development and regulate plant water use.
Asunto(s)
Calcio/química , Proteínas de Unión a Calmodulina/fisiología , Calmodulina/química , Genes de Plantas/genética , Proteínas de Plantas/metabolismo , Factores de Transcripción/fisiología , Arabidopsis/genética , Proteínas de Unión a Calmodulina/genética , Núcleo Celular/metabolismo , Sequías , Modelos Genéticos , Mutación , Fenotipo , Hojas de la Planta/genética , Plantas Modificadas Genéticamente/genética , Regiones Promotoras Genéticas , Unión Proteica , Estructura Terciaria de Proteína , ARN Mensajero/metabolismo , Factores de Transcripción/genética , Agua/químicaRESUMEN
Female gametophyte is the multicellular haploid structure that can produce embryo and endosperm after fertilization, which has become an attractive model system for investigating molecular mechanisms in nuclei migration, cell specification, cell-to-cell communication and many other processes. Previous reports found that the small ubiquitin-like modifier (SUMO) E3 ligase, SIZ1, participated in many processes depending on particular target substrates and suppression of salicylic acid (SA) accumulation. Here, we report that SIZ1 mediates the reproductive process. SIZ1 showed enhanced expression in female organs, but was not detected in the anther or pollen. A defect in the siz1-2 maternal source resulted in reduced seed-set regardless of high SA concentration within the plant. Moreover, aniline blue staining and scanning electron microscopy revealed that funicular and micropylar pollen tube guidance was arrested in siz1-2 plants. Some of the embryo sacs of ovules in siz1-2 were also disrupted quickly after stage FG7. There was no significant affects of the siz1-2 mutation on expression of genes involved in female gametophyte development- or pollen tube guidance in ovaries. Together, our results suggest that SIZ1 sustains the stability and normal function of the mature female gametophyte which is necessary for pollen tube guidance.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/enzimología , Arabidopsis/genética , Ligasas/genética , Mutación/genética , Óvulo Vegetal/enzimología , Óvulo Vegetal/crecimiento & desarrollo , Ubiquitina-Proteína Ligasas/genética , Arabidopsis/embriología , Arabidopsis/ultraestructura , Gametogénesis en la Planta/genética , Regulación del Desarrollo de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Proteínas Fluorescentes Verdes/metabolismo , Homocigoto , Óvulo Vegetal/citología , Fenotipo , Tubo Polínico/genética , Tubo Polínico/crecimiento & desarrollo , Tubo Polínico/ultraestructura , Semillas/embriología , Semillas/ultraestructuraRESUMEN
A stoma (pl. stomata) is the pore formed by two guard cells found predominantly in the leaf epidermis. Plants control stomatal aperture (opening and closing) and/or number (density) to regulate carbon dioxide (CO2) uptake and water loss (transpiration), which is necessary to optimize plant growth, development, and fitness in response to various environmental conditions. Recently, we identified Arabidopsis GT2-LIKE 1 (GTL1) as a transcriptional repressor of STOMATAL DENSITY AND DISTRIBUTION 1 (SDD1), a negative regulator of stomatal density. GTL1 directly interacts with the SDD1 promoter regulating stomatal density, transpiration, and water use efficiency (WUE). Here we discuss potential GTL1 orthologs in other plant species.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Estomas de Plantas/metabolismo , Factores de Transcripción/metabolismo , Agua/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Estomas de Plantas/genética , Serina Endopeptidasas/genética , Serina Endopeptidasas/metabolismo , Transducción de Señal , Factores de Transcripción/genéticaRESUMEN
ICE1, a MYC-type transcription factor, has an important role in the induction of CBF3/DREB1A for regulation of cold signaling and tolerance. Here we reveal that serine 403 of ICE1 is involved in regulating the transactivation and stability of the ICE1 protein. Substitution of serine 403 by alanine enhanced the transactivational activity of ICE1 in Arabidopsis protoplasts. Over-expression of ICE1(S403A) conferred more freezing tolerance than ICE1(WT) in Arabidopsis, and the expression of cold-regulated genes such as CBF3/DREB1A, COR47 and KIN1 was enhanced in plants over-expressing ICE1(S403A). Furthermore, the ICE1(S403A) protein level was not changed after cold treatment, whereas the ICE1(WT) protein level was reduced. Interestingly, polyubiquitylation of the ICE1(S403A) protein in vivo was apparently blocked. These results demonstrate that serine 403 of ICE1 has roles in both transactivation and cold-induced degradation of ICE1 via the ubiquitin/26S proteasome pathway, suggesting that serine 403 is a key residue for the attenuation of cold-stress responses by HOS1-mediated degradation of ICE1.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Respuesta al Choque por Frío , Serina/metabolismo , Factores de Transcripción/metabolismo , Sustitución de Aminoácidos , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Frío , Competencia de la Transformación por ADN , Regulación de la Expresión Génica de las Plantas , Complejo de la Endopetidasa Proteasomal/metabolismo , ARN de Planta/genética , Factores de Transcripción/genética , Activación Transcripcional , Ubiquitina/metabolismo , UbiquitinaciónRESUMEN
Phosphate (Pi) limitation causes plants to modulate the architecture of their root systems to facilitate the acquisition of Pi. Previously, we reported that the Arabidopsis (Arabidopsis thaliana) SUMO E3 ligase SIZ1 regulates root architecture remodeling in response to Pi limitation; namely, the siz1 mutations cause the inhibition of primary root (PR) elongation and the promotion of lateral root (LR) formation. Here, we present evidence that SIZ1 is involved in the negative regulation of auxin patterning to modulate root system architecture in response to Pi starvation. The siz1 mutations caused greater PR growth inhibition and LR development of seedlings in response to Pi limitation. Similar root phenotypes occurred if Pi-deficient wild-type seedlings were supplemented with auxin. N-1-Naphthylphthalamic acid, an inhibitor of auxin efflux activity, reduced the Pi starvation-induced LR root formation of siz1 seedlings to a level equivalent to that seen in the wild type. Monitoring of the auxin-responsive reporter DR5::uidA indicated that auxin accumulates in PR tips at early stages of the Pi starvation response. Subsequently, DR5::uidA expression was observed in the LR primordia, which was associated with LR elongation. The time-sequential patterning of DR5::uidA expression occurred earlier in the roots of siz1 as compared with the wild type. In addition, microarray analysis revealed that several other auxin-responsive genes, including genes involved in cell wall loosening and biosynthesis, were up-regulated in siz1 relative to wild-type seedlings in response to Pi starvation. Together, these results suggest that SIZ1 negatively regulates Pi starvation-induced root architecture remodeling through the control of auxin patterning.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/genética , Ácidos Indolacéticos/metabolismo , Ligasas/metabolismo , Fosfatos/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Arabidopsis/enzimología , Proteínas de Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Ligasas/genética , Mutación , Análisis de Secuencia por Matrices de Oligonucleótidos , Fosfatos/deficiencia , Ftalimidas/farmacología , Reguladores del Crecimiento de las Plantas/metabolismo , Raíces de Plantas/enzimología , Plantas Modificadas Genéticamente/enzimología , Plantas Modificadas Genéticamente/genética , ARN de Planta/genética , Plantones/crecimiento & desarrolloRESUMEN
A goal of modern agriculture is to improve plant drought tolerance and production per amount of water used, referred to as water use efficiency (WUE). Although stomatal density has been linked to WUE, the causal molecular mechanisms have yet to be determined. Arabidopsis thaliana GT-2 LIKE 1 (GTL1) loss-of-function mutations result in increased water deficit tolerance and higher integrated WUE by reducing daytime transpiration without a demonstrable reduction in biomass accumulation. gtl1 plants had higher instantaneous WUE that was attributable to ~25% lower transpiration and stomatal conductance but equivalent CO(2) assimilation. Lower transpiration was associated with higher STOMATAL DENSITY AND DISTRIBUTION1 (SDD1) expression and an ~25% reduction in abaxial stomatal density. GTL1 expression occurred in abaxial epidermal cells where the protein was localized to the nucleus, and its expression was downregulated by water stress. Chromatin immunoprecipitation analysis indicated that GTL1 interacts with a region of the SDD1 promoter that contains a GT3 box. An electrophoretic mobility shift assay was used to determine that the GT3 box is necessary for the interaction between GTL1 and the SDD1 promoter. These results establish that GTL1 negatively regulates WUE by modulating stomatal density via transrepression of SDD1.
Asunto(s)
Adaptación Fisiológica/fisiología , Proteínas de Arabidopsis/fisiología , Regulación hacia Abajo/fisiología , Sequías , Estomas de Plantas/metabolismo , Serina Endopeptidasas/genética , Agua/metabolismo , Proteínas de Arabidopsis/genética , Ensayo de Cambio de Movilidad Electroforética , Regulación de la Expresión Génica de las Plantas/fisiología , Microscopía Fluorescente , Mutación , Regiones Promotoras Genéticas , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Post-translational modifications diversify proteome activity to mediate complex hierarchical regulatory processes that are crucial to eukaryotic cell function. Protein modification by Ub (ubiquitin) and Ubls (ubiquitin-like proteins) in plants, as in yeast and metazoans, is necessary for numerous cellular and developmental processes and for the genetic reprogramming that occurs in response to hormonal stimuli, host-pathogen interaction-related stimuli and environmental stimuli. Ub and Ubl modifications, such as sumoylation, facilitate molecular interaction with specific substrates. Recent evidence has permitted inference of the mechanisms by which Ubl modifications regulate physiological processes such as cell-cycle progression, abscisic acid signaling, development, and biotic and abiotic stress responses. This review presents our current understanding of sumoylation and other Ubl conjugation processes in plant biology.
Asunto(s)
Plantas/metabolismo , Procesamiento Proteico-Postraduccional/fisiología , Ubiquitinas/metabolismo , Animales , Arabidopsis/citología , Arabidopsis/genética , Arabidopsis/metabolismo , Ciclo Celular , Regulación de la Expresión Génica de las Plantas/fisiología , Respuesta al Choque Térmico , Humanos , Células Vegetales , Plantas/genética , Ubiquitinas/genéticaRESUMEN
The post-translational conjugation of small ubiquitin-related modifiers (SUMOs) to other proteins is involved in regulation of many processes in eukaryotic development; although its role in plant development is beginning to be dissected. Previously, we demonstrated that the siz1 mutant, which is impaired in SUMO E3 ligase, showed a dwarf-like shoot phenotype with accumulation of salicylic acid (SA), and the expression of nahG, a bacterial salicylate hydroxylase that catabolizes SA, in siz1 reduced the SA level and suppressed dwarfism. Herein, we provide evidence that the SIZ1 gene controls cell division and elongation through regulation of the SA level. Mature siz1-2 and siz1-3 plants exhibited a dwarf-like shoot phenotype that is attributable to decreased leaf cell volume and number relative to the wild type. Cell division and expansion defects caused by siz1 were also suppressed by the expression of nahG. Expression of XTH8 and XTH31, encoding xyloglucan endotransglycosylase/hydrolase, which are thought to facilitate leaf cell expansion, was down-regulated in siz1 leaves. However, reduced XTH8 and XTH31 expression in siz1 plants was restored in nahG siz1-2 plants. These results indicate that SIZ1 regulates cell growth and plant development with regulation of SA accumulation. Also, XTH8 and XTH31 genes may be responsible for reduced leaf cell expansion.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Aumento de la Célula , Procesos de Crecimiento Celular/fisiología , Glicósido Hidrolasas/metabolismo , Ligasas/metabolismo , Ácido Salicílico/metabolismo , Arabidopsis/citología , Proteínas de Arabidopsis/genética , Diferenciación Celular/fisiología , Proliferación Celular , Tamaño de la Célula , Enanismo/genética , Enanismo/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Regulación de la Expresión Génica de las Plantas/fisiología , Glicósido Hidrolasas/genética , Glicosiltransferasas/genética , Glicosiltransferasas/metabolismo , Ligasas/genética , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Hojas de la Planta/citología , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/metabolismo , Brotes de la Planta/citología , Brotes de la Planta/crecimiento & desarrollo , Brotes de la Planta/metabolismoRESUMEN
NHX-type antiporters in the tonoplast have been reported to increase the salt tolerance of various plants species, and are thought to mediate the compartmentation of Na(+) in vacuoles. However, all isoforms characterized so far catalyze both Na(+)/H(+) and K(+)/H(+) exchange. Here, we show that AtNHX1 has a critical involvement in the subcellular partitioning of K(+), which in turn affects plant K(+) nutrition and Na(+) tolerance. Transgenic tomato plants overexpressing AtNHX1 had larger K(+) vacuolar pools in all growth conditions tested, but no consistent enhancement of Na(+) accumulation was observed under salt stress. Plants overexpressing AtNHX1 have a greater capacity to retain intracellular K(+) and to withstand salt-shock. Under K(+)-limiting conditions, greater K(+) compartmentation in the vacuole occurred at the expense of the cytosolic K(+) pool, which was lower in transgenic plants. This caused the early activation of the high-affinity K(+) uptake system, enhanced K(+) uptake by roots, and increased the K(+) content in plant tissues and the xylem sap of transformed plants. Our results strongly suggest that NHX proteins are likely candidates for the H(+)-linked K(+) transport that is thought to facilitate active K(+) uptake at the tonoplast, and the partitioning of K(+) between vacuole and cytosol.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Transporte de Catión/metabolismo , Potasio/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismo , Solanum lycopersicum/metabolismo , Vacuolas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Transporte de Catión/genética , Regulación de la Expresión Génica de las Plantas , Solanum lycopersicum/genética , Plantas Modificadas Genéticamente , Intercambiadores de Sodio-Hidrógeno/genéticaRESUMEN
Insertion mutations that disrupt the function of PHT4;6 (At5g44370) cause NaCl hypersensitivity of Arabidopsis seedlings that is characterized by reduced growth of the primary root, enhanced lateral branching, and swelling of root tips. Mutant phenotypes were exacerbated by sucrose, but not by equiosmolar concentrations of mannitol, and attenuated by low inorganic phosphate in the medium. Protein PHT4;6 belongs to the Major Facilitator Superfamily of permeases that shares significant sequence similarity to mammalian type-I Pi transporters and vesicular glutamate transporters, and is a member of the PHT4 family of putative intracellular phosphate transporters of plants. PHT4;6 localizes to the Golgi membrane and transport studies indicate that PHT4;6 facilitates the selective transport of Pi but not of chloride or inorganic anions. Phenotypic similarities with other mutants displaying root swelling suggest that PHT4;6 likely functions in protein N-glycosylation and cell wall biosynthesis, which are essential for salt tolerance. Together, our results indicate that PHT4;6 transports Pi out of the Golgi lumenal space for the re-cycling of the Pi released from glycosylation processes.
Asunto(s)
Arabidopsis/fisiología , Aparato de Golgi/fisiología , Proteínas de Transporte de Fosfato/fisiología , Tolerancia a la Sal/fisiología , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/fisiología , Transporte Biológico/fisiología , Proteínas de Transporte de Fosfato/genética , Proteínas de Transporte de Fosfato/metabolismo , Fosfatos/metabolismo , Fosfatos/fisiología , Raíces de Plantas/fisiología , Brotes de la Planta/fisiologíaRESUMEN
Mutants of the plant cation/H(+) antiporter AtNHX1 that confer greater halotolerance were generated by random mutagenesis and selected in yeast by phenotypic complementation. The amino acid substitutions that were selected were conservative and occurred in the second half of the membrane-associated N terminus. AtNHX1 complemented the lack of endogenous ScNHX1 in endosomal protein trafficking assays. Growth enhancement on hygromycin B and vanadate media agreed with a generally improved endosomal/prevacuolar function of the mutated proteins. In vivo measurements by (31)P NMR revealed that wild-type and mutant AtNHX1 transporters did not affect cytosolic or vacuolar pH. Surprisingly, when yeast cells were challenged with lithium, a tracer for sodium, the main effect of the mutations in AtNHX1 was a reduction in the amount of compartmentalized lithium. When purified and reconstituted into proteoliposomes or assayed in intact vacuoles isolated from yeast cells, a representative mutant transporter (V318I) showed a greater cation discrimination favoring potassium transport over that of sodium or lithium. Together, our data suggest that the endosome/prevacuolar compartment is a target for salt toxicity. Poisoning by toxic cations in the endosome/prevacuolar compartment is detrimental for cell functions, but it can be alleviated by improving the discrimination of transported alkali cations by the resident cation/H(+) antiporter.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Proteínas de Transporte de Catión/metabolismo , Endosomas/metabolismo , Mutación/genética , Saccharomyces cerevisiae/metabolismo , Tolerancia a la Sal/efectos de los fármacos , Cloruro de Sodio/toxicidad , Intercambiadores de Sodio-Hidrógeno/metabolismo , Vacuolas/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Transporte de Catión/química , Cationes , Compartimento Celular/efectos de los fármacos , Endosomas/efectos de los fármacos , Prueba de Complementación Genética , Concentración de Iones de Hidrógeno/efectos de los fármacos , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Espectroscopía de Resonancia Magnética , Modelos Biológicos , Fenotipo , Transporte de Proteínas/efectos de los fármacos , Proteolípidos/efectos de los fármacos , Proteolípidos/metabolismo , Saccharomyces cerevisiae/citología , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/crecimiento & desarrollo , Intercambiadores de Sodio-Hidrógeno/química , Vacuolas/efectos de los fármacosRESUMEN
SUMO (small ubiquitin-related modifier) conjugation (i.e., sumoylation) to protein substrates is a reversible posttranslational modification that regulates signaling by modulating transcription factor activity. This paper presents evidence that the SUMO E3 ligase SIZ1 negatively regulates abscisic acid (ABA) signaling, which is dependent on the bZIP transcripton factor ABI5. Loss-of-function T-DNA insertion siz1-2 and siz1-3 mutations caused ABA hypersensitivity for seed germination arrest and seedling primary root growth inhibition. Furthermore, expression of genes that are ABA-responsive through ABI5-dependent signaling (e.g., RD29A, Rd29B, AtEm6, RAB18, ADH1) was hyperinduced by the hormone in siz1 seedlings. abi5-4 suppressed ABA hypersensitivity caused by siz1 (siz1-2 abi5-4), demonstrating an epistatic genetic interaction between SIZ1 and ABI5. A K391R substitution in ABI5 [ABI5(K391R)] blocked SIZ1-mediated sumoylation of the transcription factor in vitro and in Arabidopsis protoplasts, indicating that ABI5 is sumoylated through SIZ1 and that K391 is the principal site for SUMO conjugation. In abi5-4 plants, ABI5(K391R) expression caused greater ABA hypersensitivity (gene expression, seed germination arrest and primary root growth inhibition) compared with ABI5 expression. Together, these results establish that SIZ1-dependent sumoylation of ABI5 attenuates ABA signaling. The double mutant siz1-2 afp-1 exhibited even greater ABA sensitivity than the single mutant siz1, suggesting that SIZ1 represses ABI5 signaling function independent of AFP1.
Asunto(s)
Ácido Abscísico/metabolismo , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/fisiología , Arabidopsis/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Ligasas/fisiología , Transducción de Señal , Proteínas de Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Germinación , Ligasas/genética , Ligasas/metabolismo , Mutación , Raíces de Plantas/crecimiento & desarrollo , Proteína SUMO-1 , Ubiquitina-Proteína LigasasRESUMEN
The conjugation of small ubiquitin-related modifier (SUMO) to substrates (sumoylation) is one of posttranslational modification systems in eukaryotes. Sumoylation plays an important role in the regulation of environmental stress response, biotic stress response, and flowering control in plants. Covalent SUMO conjugation requires an E1/E2/E3 enzyme, and SUMO E3 ligase SIZ1 is essential for these regulations. This addendum summarizes our recent study in which it has been established that in Arabidopsis, SUMO E3 ligase SIZ1 negatively controls abscisic acid (ABA) signaling through the sumoylation of ABI5. The conjugation of SUMO to ABI5 represses its activity and also prevents ABI5 from undergoing degradation.
RESUMEN
Histone modification in chromatin is one of the key control points in gene regulation in eukaryotic cells. Protein complexes composed of histone acetyltransferase or deacetylase, WD40 repeat protein, and many other components have been implicated in this process. Here, we report the identification and functional characterization of HOS15, a WD40-repeat protein crucial for repression of genes associated with abiotic stress tolerance through histone deacetylation in Arabidopsis. HOS15 shares high sequence similarity with human transducin-beta like protein (TBL), a component of a repressor protein complex involved in histone deacetylation. Mutation of the HOS15 gene renders mutant plants hypersensitive to freezing temperatures. HOS15 is localized in the nucleus and specifically interacts with histone H4. The level of acetylated histone H4 is higher in the hos15 mutant than in WT plants. Moreover, the stress inducible RD29A promoter is hyperinduced and associated with a substantially higher level of acetylated histone H4 in the hos15 mutant under cold stress conditions. Our results suggest a critical role for gene activation/repression by histone acetylation/deacetylation in plant acclimation and tolerance to cold stress.