RESUMEN
Homeostasis of the essential micronutrient manganese (Mn) is crucially determined through availability and uptake efficiency in all organisms. Mn deficiency of plants especially occurs in alkaline and calcareous soils, seriously restricting crop yield. However, the mechanisms underlying the sensing and signaling of Mn availability and conferring regulation of Mn uptake await elucidation. Here, we uncover that Mn depletion triggers spatiotemporally defined long-lasting Ca2+ oscillations in Arabidopsis roots. These Ca2+ signals initiate in individual cells, expand, and intensify intercellularly to transform into higher-order multicellular oscillations. Furthermore, through an interaction screen we identified the Ca2+-dependent protein kinases CPK21 and CPK23 as Ca2+ signal-decoding components that bring about translation of these signals into regulation of uptake activity of the high-affinity Mn transporter natural resistance associated macrophage proteins 1 (NRAMP1). Accordingly, a cpk21/23 double mutant displays impaired growth and root development under Mn-limiting conditions, while kinase overexpression confers enhanced tolerance to low Mn supply to plants. In addition, we define Thr498 phosphorylation within NRAMP1 as a pivot mechanistically determining NRAMP1 activity, as revealed by biochemical assays and complementation of yeast Mn uptake and Arabidopsis nramp1 mutants. Collectively, these findings delineate the Ca2+-CPK21/23-NRAMP1 axis as key for mounting plant Mn homeostasis.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Calcio , Proteínas de Transporte de Catión , Manganeso , Proteínas Quinasas , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Calcio/metabolismo , Proteínas de Transporte de Catión/genética , Proteínas de Transporte de Catión/metabolismo , Homeostasis , Manganeso/metabolismo , Micronutrientes/metabolismo , Fosforilación , Raíces de Plantas/metabolismo , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Saccharomyces cerevisiae/metabolismo , SueloRESUMEN
Signaling cross talks between auxin, a regulator of plant development, and Ca2+, a universal second messenger, have been proposed to modulate developmental plasticity in plants. However, the underlying molecular mechanisms are largely unknown. Here, we report that in Arabidopsis roots, auxin elicits specific Ca2+ signaling patterns that spatially coincide with the expression pattern of auxin-regulated genes. We have identified the single EF-hand Ca2+-binding protein Ca2+-dependent modulator of ICR1 (CMI1) as an interactor of the Rho of plants (ROP) effector interactor of constitutively active ROP (ICR1). CMI1 expression is directly up-regulated by auxin, whereas the loss of function of CMI1 associates with the repression of auxin-induced Ca2+ increases in the lateral root cap and vasculature, indicating that CMI1 represses early auxin responses. In agreement, cmi1 mutants display an increased auxin response including shorter primary roots, longer root hairs, longer hypocotyls, and altered lateral root formation. Binding to ICR1 affects subcellular localization of CMI1 and its function. The interaction between CMI1 and ICR1 is Ca2+-dependent and involves a conserved hydrophobic pocket in CMI1 and calmodulin binding-like domain in ICR1. Remarkably, CMI1 is monomeric in solution and in vitro changes its secondary structure at cellular resting Ca2+ concentrations ranging between 10-9 and 10-8 M. Hence, CMI1 is a Ca2+-dependent transducer of auxin-regulated gene expression, which can function in a cell-specific fashion at steady-state as well as at elevated cellular Ca2+ levels to regulate auxin responses.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Unión al Calcio/metabolismo , Calcio/metabolismo , Proteínas Portadoras/metabolismo , Ácidos Indolacéticos/metabolismo , Raíces de Plantas/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Proteínas de Unión al Calcio/genética , Proteínas Portadoras/genética , Perfilación de la Expresión Génica/métodos , Regulación del Desarrollo de la Expresión Génica/efectos de los fármacos , Regulación del Desarrollo de la Expresión Génica/genética , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/genética , Ácidos Indolacéticos/farmacología , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , Plantas Modificadas Genéticamente , Unión Proteica , Transducción de Señal/efectos de los fármacos , Transducción de Señal/genéticaRESUMEN
In plants, potassium (K+ ) homeostasis is tightly regulated and established against a concentration gradient to the environment. Despite the identification of Ca2+ -regulated kinases as modulators of K+ channels, the immediate signaling and adaptation mechanisms of plants to low-K+ conditions are only partially understood. To assess the occurrence and role of Ca2+ signals in Arabidopsis thaliana roots, we employed ratiometric analyses of Ca2+ dynamics in plants expressing the Ca2+ reporter YC3.6 in combination with patch-clamp analyses of root cells and two-electrode voltage clamp (TEVC) analyses in Xenopus laevis oocytes. K+ deficiency triggers two successive and distinct Ca2+ signals in roots exhibiting spatial and temporal specificity. A transient primary Ca2+ signature arose within 1 min in the postmeristematic stelar tissue of the elongation zone, while a secondary Ca2+ response occurred after several hours as sustained Ca2+ elevation in defined tissues of the elongation and root hair differentiation zones. Patch-clamp and TEVC analyses revealed Ca2+ dependence of the activation of the K+ channel AKT1 by the CBL1-CIPK23 Ca2+ sensor-kinase complex. Together, these findings identify a critical role of cell group-specific Ca2+ signaling in low K+ responses and indicate an essential and direct role of Ca2+ signals for AKT1 K+ channel activation in roots.
Asunto(s)
Arabidopsis/metabolismo , Señalización del Calcio , Potasio/metabolismo , Adaptación Fisiológica/efectos de los fármacos , Animales , Arabidopsis/efectos de los fármacos , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Calcio/metabolismo , Señalización del Calcio/efectos de los fármacos , Citoplasma/efectos de los fármacos , Citoplasma/metabolismo , Citosol/efectos de los fármacos , Citosol/metabolismo , Electrodos , Activación del Canal Iónico/efectos de los fármacos , Lantano/farmacología , Mutación/genética , Oocitos/efectos de los fármacos , Oocitos/metabolismo , Técnicas de Placa-Clamp , Raíces de Plantas/citología , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/metabolismo , Dominios Proteicos , Protoplastos/efectos de los fármacos , Protoplastos/metabolismo , Factores de Tiempo , XenopusRESUMEN
Ca(2+) signatures are central to developmental processes and adaptive responses in plants. However, high-resolution studies of Ca(2+) dynamics using genetically encoded Ca(2+) indicators (GECIs) such as Yellow Cameleon (YC) proteins have so far not been conducted in important model crops such as rice (Oryza sativa). We conducted a comparative study of 35S and ubiquitin-10 (UBQ10) promoter functionality in Arabidopsis thaliana and O. sativa plants expressing the Ca(2+) indicator Yellow Cameleon 3.6 (YC3.6) under control of the UBQ10 or 35S promoter. Ca(2+) signatures in roots of both species were analyzed during exposure to hyperpolarization/depolarization cycles or in response to application of the amino acid glutamate. We found a superior performance of the UBQ10 promoter with regard to expression pattern, levels and expression stabilities in both species. We observed remarkable differences between the two species in the spatiotemporal parameters of the observed Ca(2+) signatures. Rice appeared in general to respond with a lower maximal signal amplitude but greatly increased signal duration when compared with Arabidopsis. Our results identify important advantages to using the UBQ10 promoter in Arabidopsis and rice and in T-DNA mutant backgrounds. Moreover, the observed differences in Ca(2+) signaling in the two species underscore the need for comparative studies to achieve a comprehensive understanding of Ca(2+) signaling in plants.
Asunto(s)
Arabidopsis/genética , Arabidopsis/metabolismo , Calcio/análisis , Calmodulina/metabolismo , Proteínas Luminiscentes/metabolismo , Oryza/metabolismo , Regiones Promotoras Genéticas/genética , Proteínas Recombinantes de Fusión/metabolismo , Arabidopsis/citología , Calcio/metabolismo , Señalización del Calcio , Calmodulina/genética , Citoplasma/metabolismo , Expresión Génica , Genes Reporteros , Ácido Glutámico/metabolismo , Procesamiento de Imagen Asistido por Computador , Proteínas Luminiscentes/genética , Microscopía Confocal , Oryza/citología , Oryza/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente , Proteínas Recombinantes de Fusión/genética , TransgenesRESUMEN
Excessive Na+ in soils inhibits plant growth. Here, we report that Na+ stress triggers primary calcium signals specifically in a cell group within the root differentiation zone, thus forming a "sodium-sensing niche" in Arabidopsis. The amplitude of this primary calcium signal and the speed of the resulting Ca2+ wave dose-dependently increase with rising Na+ concentrations, thus providing quantitative information about the stress intensity encountered. We also delineate a Ca2+-sensing mechanism that measures the stress intensity in order to mount appropriate salt detoxification responses. This is mediated by a Ca2+-sensor-switch mechanism, in which the sensors SOS3/CBL4 and CBL8 are activated by distinct Ca2+-signal amplitudes. Although the SOS3/CBL4-SOS2/CIPK24-SOS1 axis confers basal salt tolerance, the CBL8-SOS2/CIPK24-SOS1 module becomes additionally activated only in response to severe salt stress. Thus, Ca2+-mediated translation of Na+ stress intensity into SOS1 Na+/H+ antiporter activity facilitates fine tuning of the sodium extrusion capacity for optimized salt-stress tolerance.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Calcio/metabolismo , Estrés Salino , Sodio/metabolismo , Intercambiadores de Sodio-Hidrógeno/genéticaRESUMEN
Organismal homeostasis of the essential ion K+ requires sensing of its availability, efficient uptake, and defined distribution. Understanding plant K+ nutrition is essential to advance sustainable agriculture, but the mechanisms underlying K+ sensing and the orchestration of downstream responses have remained largely elusive. Here, we report where plants sense K+ deprivation and how this translates into spatially defined ROS signals to govern specific downstream responses. We define the organ-scale K+ pattern of roots and identify a postmeristematic K+-sensing niche (KSN) where rapid K+ decline and Ca2+ signals coincide. Moreover, we outline a bifurcating low-K+-signaling axis of CIF peptide-activated SGN3-LKS4/SGN1 receptor complexes that convey low-K+-triggered phosphorylation of the NADPH oxidases RBOHC, RBOHD, and RBOHF. The resulting ROS signals simultaneously convey HAK5 K+ uptake-transporter induction and accelerated Casparian strip maturation. Collectively, these mechanisms synchronize developmental differentiation and transcriptome reprogramming for maintaining K+ homeostasis and optimizing nutrient foraging by roots.
Asunto(s)
Adaptación Fisiológica , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Homeostasis , Nutrientes/metabolismo , Raíces de Plantas/metabolismo , Potasio/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Complejo del Señalosoma COP9/genética , Complejo del Señalosoma COP9/metabolismo , Calcio/metabolismo , Regulación de la Expresión Génica de las Plantas , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , TranscriptomaRESUMEN
Cells maintain integrity despite changes in their mechanical properties elicited during growth and environmental stress. How cells sense their physical state and compensate for cell-wall damage is poorly understood, particularly in plants. Here we report that FERONIA (FER), a plasma-membrane-localized receptor kinase from Arabidopsis, is necessary for the recovery of root growth after exposure to high salinity, a widespread soil stress. The extracellular domain of FER displays tandem regions of homology with malectin, an animal protein known to bind di-glucose in vitro and important for protein quality control in the endoplasmic reticulum. The presence of malectin-like domains in FER and related receptor kinases has led to widespread speculation that they interact with cell-wall polysaccharides and can potentially serve a wall-sensing function. Results reported here show that salinity causes softening of the cell wall and that FER is necessary to sense these defects. When this function is disrupted in the fer mutant, root cells explode dramatically during growth recovery. Similar defects are observed in the mur1 mutant, which disrupts pectin cross-linking. Furthermore, fer cell-wall integrity defects can be rescued by treatment with calcium and borate, which also facilitate pectin cross-linking. Sensing of these salinity-induced wall defects might therefore be a direct consequence of physical interaction between the extracellular domain of FER and pectin. FER-dependent signaling elicits cell-specific calcium transients that maintain cell-wall integrity during salt stress. These results reveal a novel extracellular toxicity of salinity, and identify FER as a sensor of damage to the pectin-associated wall.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/fisiología , Señalización del Calcio/genética , Fosfotransferasas/genética , Estrés Salino/fisiología , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Pared Celular/metabolismo , Fosfotransferasas/metabolismoRESUMEN
The significance of cell wall invertase (cwINV) for plant defense was investigated by comparing wild-type tobacco (Nicotiana tabacum) Samsun NN (SNN) with plants with RNA interference (RNAi)-mediated repression of cwINV (SNNcwINV). In source leaves of SNNcwINV, the activity of cwINV was repressed by about 90%. Sucrose export and apoplastic carbohydrate levels were significantly reduced, while photosynthesis and dark respiration exhibited little or no change. Activities of sucrose synthase and phosphofructokinase were depressed moderately, while ADP-glucose pyrophosphorylase was diminished greatly. Yet, the content of cytosolic/vacuolar carbohydrates was not significantly lower, which correlated with the absence of phenotypic effects in SNNcwINV under normal growing conditions. By contrast, defense-related processes in primary metabolism and hypersensitive cell death were impaired and delayed in correlation with repression of cwINV. The increase in cwINV observed in source leaves of the resistant wild type following infection with Phytophthora nicotianae was absent in SNNcwINV. Also, defense-related callose deposition at cell-to-cell interfaces, the related decline in sugar export, and accumulation of apoplastic carbohydrates were reduced and delayed. Expression of pathogenesis-related proteins and increase in phenylalanine ammonia-lyase and glucose-6-phosphate dehydrogenase activities were alleviated. Formation of hydrogen peroxide and development of hypersensitive lesions were weak and heterogeneous, and the pathogen was able to sporulate. We conclude that in photosynthetically active leaves of the apoplastic phloem loader, tobacco cwINV plays an essential role for acquisition of carbohydrates during plant-pathogen interactions and that the availability of these carbohydrates supports the onset of the hypersensitive reaction and ensures successful defense.