RESUMEN
Plant hormones coordinate responses to environmental cues with developmental programs1, and are fundamental for stress resilience and agronomic yield2. The core signalling pathways underlying the effects of phytohormones have been elucidated by genetic screens and hypothesis-driven approaches, and extended by interactome studies of select pathways3. However, fundamental questions remain about how information from different pathways is integrated. Genetically, most phenotypes seem to be regulated by several hormones, but transcriptional profiling suggests that hormones trigger largely exclusive transcriptional programs4. We hypothesized that protein-protein interactions have an important role in phytohormone signal integration. Here, we experimentally generated a systems-level map of the Arabidopsis phytohormone signalling network, consisting of more than 2,000 binary protein-protein interactions. In the highly interconnected network, we identify pathway communities and hundreds of previously unknown pathway contacts that represent potential points of crosstalk. Functional validation of candidates in seven hormone pathways reveals new functions for 74% of tested proteins in 84% of candidate interactions, and indicates that a large majority of signalling proteins function pleiotropically in several pathways. Moreover, we identify several hundred largely small-molecule-dependent interactions of hormone receptors. Comparison with previous reports suggests that noncanonical and nontranscription-mediated receptor signalling is more common than hitherto appreciated.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Mapas de Interacción de Proteínas , Transducción de Señal , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Unión Proteica , Mapeo de Interacción de Proteínas , Reproducibilidad de los Resultados , Transcripción GenéticaRESUMEN
An amendment to this paper has been published and can be accessed via a link at the top of the paper.
RESUMEN
Copper (Cu) and iron (Fe) are essential for plant growth and are often in short supply under natural conditions. Molecular responses to simultaneous lack of both metals (-Cu-Fe) differ from those seen in the absence of either alone. Metabolome profiling of plant leaves previously revealed that fumarate levels fall under -Cu-Fe conditions. We employed lines lacking cytosolic FUMARASE2 (FUM2) activity to study the impact of constitutive suppression of cytosolic fumarate synthesis on plant growth under Cu and/or Fe deficiency. In fum2 mutants, photosynthesis and growth were less impaired under -Cu-Fe conditions than in wild-type (WT) seedlings. In particular, levels of photosynthetic proteins, chloroplast ultrastructure, amino acid profiles and redox state were less perturbed by simultaneous Cu-Fe deficiency in lines that cannot produce fumarate in the cytosol. Although cytosolic fumarate has been reported to promote acclimation of photosynthesis to low temperatures when metal supplies are adequate, the photosynthetic efficiency of fum2 lines grown under Cu-Fe deficiency in the cold was higher than in WT. Uptake and contents of Cu and Fe are similar in WT and fum2 plants under control and -Cu-Fe conditions, and lack of FUM2 does not alter the ability to sense metal deficiency, as indicated by marker gene expression. Collectively, we propose that reduced levels of cytosolic fumarate synthesis ultimately increase the availability of Fe for incorporation into metalloproteins.
Asunto(s)
Proteínas de Arabidopsis/fisiología , Arabidopsis/crecimiento & desarrollo , Cobre/deficiencia , Fumarato Hidratasa/fisiología , Hierro/metabolismo , Fotosíntesis , Aminoácidos/metabolismo , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Fumarato Hidratasa/genética , Fumaratos/metabolismo , Plantones/crecimiento & desarrollo , Plantones/metabolismo , Tilacoides/metabolismoRESUMEN
Plant responses to coincident nutrient deficiencies cannot be predicted from the responses to individual deficiencies. Although copper (Cu) and iron (Fe) are essential micronutrients for plant growth that are often and concurrently limited in soils, the combinatorial response to Cu-Fe deficiency remains elusive. In the present study, we characterised the responses of Arabidopsis thaliana plants deprived of Cu, Fe or both (-Cu-Fe) at the level of plant development, mineral composition, and reconfiguration of transcriptomes, proteomes and metabolomes. Compared to single deficiencies, simultaneous -Cu-Fe leads to a distinct pattern in leaf physiology and microelement concentration characterised by lowered protein content and enhanced manganese and zinc levels. Conditional networking analysis of molecular changes indicates that biological processes also display different co-expression patterns among single and double deficiencies. Indeed, the interaction between Cu and Fe deficiencies causes distinct expression profiles for 15% of all biomolecules, leading to specific enhancement of general stress responses and protein homeostasis mechanisms, at the same time as severely arresting photosynthesis. Accordingly, central carbon metabolites, in particular photosynthates, decrease especially under -Cu-Fe conditions, whereas the pool of free amino acids increases. Further meta-analysis of transcriptomes and proteomes corroborated that protein biosynthesis and folding capacity were readjusted during the combinatorial response and unveiled important rearrangements in the metabolism of organic acids. Consequently, our results demonstrate that the response to -Cu-Fe imposes a distinct reconfiguration of large sets of molecules, not triggered by single deficiencies, resulting into a switch from autotrophy to heterotrophy and involving organic acids such as fumaric acid as central mediators of the response.
Asunto(s)
Arabidopsis/metabolismo , Cobre/deficiencia , Deficiencias de Hierro , Arabidopsis/crecimiento & desarrollo , Perfilación de la Expresión Génica , Metabolómica , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/metabolismo , Estrés Fisiológico , Biología de SistemasRESUMEN
BACKGROUND: Cold stress causes dynamic changes in gene expression that are partially caused by small non-coding RNAs since they regulate protein coding transcripts and act in epigenetic gene silencing pathways. Thus, a detailed analysis of transcriptional changes of small RNAs (sRNAs) belonging to all known sRNA classes such as microRNAs (miRNA) and small interfering RNA (siRNAs) in response to cold contributes to an understanding of cold-related transcriptome changes. RESULT: We subjected A. thaliana plants to cold acclimation conditions (4 °C) and analyzed the sRNA transcriptomes after 3 h, 6 h and 2 d. We found 93 cold responsive differentially expressed miRNAs and only 14 of these were previously shown to be cold responsive. We performed miRNA target prediction for all differentially expressed miRNAs and a GO analysis revealed the overrepresentation of miRNA-targeted transcripts that code for proteins acting in transcriptional regulation. We also identified a large number of differentially expressed cis- and trans-nat-siRNAs, as well as sRNAs that are derived from long non-coding RNAs. By combining the results of sRNA and mRNA profiling with miRNA target predictions and publicly available information on transcription factors, we reconstructed a cold-specific, miRNA and transcription factor dependent gene regulatory network. We verified the validity of links in the network by testing its ability to predict target gene expression under cold acclimation. CONCLUSION: In A. thaliana, miRNAs and sRNAs derived from cis- and trans-NAT gene pairs and sRNAs derived from lncRNAs play an important role in regulating gene expression in cold acclimation conditions. This study provides a fundamental database to deepen our knowledge and understanding of regulatory networks in cold acclimation.
Asunto(s)
Aclimatación/genética , Arabidopsis/genética , ARN de Planta/fisiología , ARN Pequeño no Traducido/fisiología , Arabidopsis/fisiología , Frío , Regulación de la Expresión Génica de las Plantas , Ontología de Genes , Redes Reguladoras de Genes , Genes de Plantas , TranscriptomaRESUMEN
KEY MESSAGE: Copper deficiency and excess differentially affect iron homeostasis in rice and overexpression of the Arabidopsis high-affinity copper transporter COPT1 slightly increases endogenous iron concentration in rice grains. Higher plants have developed sophisticated mechanisms to efficiently acquire and use micronutrients such as copper and iron. However, the molecular mechanisms underlying the interaction between both metals remain poorly understood. In the present work, we study the effects produced on iron homeostasis by a wide range of copper concentrations in the growth media and by altered copper transport in Oryza sativa plants. Gene expression profiles in rice seedlings grown under copper excess show an altered expression of genes involved in iron homeostasis compared to standard control conditions. Thus, ferritin OsFER2 and ferredoxin OsFd1 mRNAs are down-regulated whereas the transcriptional iron regulator OsIRO2 and the nicotianamine synthase OsNAS2 mRNAs rise under copper excess. As expected, the expression of OsCOPT1, which encodes a high-affinity copper transport protein, as well as other copper-deficiency markers are down-regulated by copper. Furthermore, we show that Arabidopsis COPT1 overexpression (C1 OE ) in rice causes root shortening in high copper conditions and under iron deficiency. C1 OE rice plants modify the expression of the putative iron-sensing factors OsHRZ1 and OsHRZ2 and enhance the expression of OsIRO2 under copper excess, which suggests a role of copper transport in iron signaling. Importantly, the C1 OE rice plants grown on soil contain higher endogenous iron concentration than wild-type plants in both brown and white grains. Collectively, these results highlight the effects of rice copper status on iron homeostasis, which should be considered to obtain crops with optimized nutrient concentrations in edible parts.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cobre/farmacología , Homeostasis , Hierro/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Oryza/genética , Oryza/metabolismo , Transportador de Cobre 1 , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Homeostasis/efectos de los fármacos , Oryza/efectos de los fármacos , Oryza/crecimiento & desarrollo , Fenotipo , Proteínas de Plantas , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/metabolismo , Brotes de la Planta/efectos de los fármacos , Brotes de la Planta/genética , Plantas Modificadas Genéticamente , Transcriptoma/genéticaRESUMEN
In natural environments, plants often experience different stresses simultaneously, and adverse abiotic conditions can weaken the plant immune system. Interactome mapping revealed that the LOW SULPHUR UPREGULATED (LSU) proteins are hubs in an Arabidopsis protein interaction network that are targeted by virulence effectors from evolutionarily diverse pathogens. Here we show that LSU proteins are up-regulated in several abiotic and biotic stress conditions, such as nutrient depletion or salt stress, by both transcriptional and post-translational mechanisms. Interference with LSU expression prevents chloroplastic reactive oxygen species (ROS) production and proper stomatal closure during sulphur stress. We demonstrate that LSU1 interacts with the chloroplastic superoxide dismutase FSD2 and stimulates its enzymatic activity in vivo and in vitro. Pseudomonas syringae virulence effectors interfere with this interaction and preclude re-localization of LSU1 to chloroplasts. We demonstrate that reduced LSU levels cause a moderately enhanced disease susceptibility in plants exposed to abiotic stresses such as nutrient deficiency, high salinity, or heavy metal toxicity, whereas LSU1 overexpression confers significant disease resistance in several of these conditions. Our data suggest that the network hub LSU1 plays an important role in co-ordinating plant immune responses across a spectrum of abiotic stress conditions.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Proteínas Nucleares/genética , Pseudomonas syringae/fisiología , Superóxido Dismutasa/genética , Arabidopsis/inmunología , Arabidopsis/microbiología , Proteínas de Arabidopsis/metabolismo , Resistencia a la Enfermedad/inmunología , Proteínas Nucleares/metabolismo , Enfermedades de las Plantas/inmunología , Enfermedades de las Plantas/microbiología , Estrés Fisiológico , Azufre/metabolismo , Superóxido Dismutasa/metabolismoRESUMEN
Cadmium toxicity interferes with essential metal homeostasis, which is a problem for both plant nutrition and the consumption of healthy food by humans. Copper uptake is performed by the members of the Arabidopsis high affinity copper transporter (COPT) family. One of the members, COPT5, is involved in copper recycling from the vacuole toward the cytosolic compartment. We show herein that copt5 mutants are more sensitive to cadmium stress than wild-type plants, as indicated by reduced growth. Exacerbated cadmium toxicity in copt5 mutants is due specifically to altered copper traffic through the COPT5 transporter. Three different processes which have been shown to affect cadmium tolerance are altered in copt5 mutants. First, ethylene biosynthesis diminishes under copper deficiency and, in the presence of cadmium, ethylene production diminishes further. Copper deficiency responses are also attenuated under cadmium treatment. Remarkably, while copt5 roots present higher oxidative stress toxicity symptoms than controls, aerial copt5 parts display lower oxidative stress, as seen by reduced cadmium delivery to shoots. Taken together, these results demonstrate that copper transport plays a key role in cadmium resistance, and suggest that oxidative stress triggers an NADPH oxidase-mediated signaling pathway, which contributes to cadmium translocation and basal plant resistance. The slightly lower cadmium levels that reach aerial parts in the copt5 mutants, irrespective of the copper content in the media, suggest a new biotechnological approach to minimize toxic cadmium entry into food chains.
Asunto(s)
Adaptación Fisiológica/efectos de los fármacos , Proteínas de Arabidopsis/genética , Arabidopsis/metabolismo , Cadmio/toxicidad , Proteínas de Transporte de Catión/genética , Cobre/metabolismo , Mutación/genética , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico/efectos de los fármacos , Biomarcadores/metabolismo , Proteínas de Transporte de Catión/metabolismo , Etilenos/farmacología , Etiolado/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Hipocótilo/efectos de los fármacos , Hipocótilo/crecimiento & desarrollo , Hierro/toxicidad , Peroxidación de Lípido/efectos de los fármacos , Modelos Biológicos , Estrés Oxidativo/efectos de los fármacos , Desarrollo de la Planta/efectos de los fármacos , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Proteínas SLC31 , Plantones/efectos de los fármacos , Plantones/crecimiento & desarrollo , Estrés Fisiológico/efectos de los fármacos , Estrés Fisiológico/genéticaRESUMEN
Proper copper (Cu) homeostasis is required by living organisms to maintain essential cellular functions. In the model plant Arabidopsis (Arabidopsis thaliana), the SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE7 (SPL7) transcription factor participates in reprogramming global gene expression during Cu insufficiency in order to improve the metal uptake and prioritize its distribution to Cu proteins of major importance. As a consequence, spl7 null mutants show morphological and physiological disorders during Cu-limited growth, resulting in lower fresh weight, reduced root elongation, and chlorosis. On the other hand, the Arabidopsis KIN17 homolog belongs to a well-conserved family of essential eukaryotic nuclear proteins known to be stress activated and involved in DNA and possibly RNA metabolism in mammals. In the study presented here, we uncovered that Arabidopsis KIN17 participates in promoting the Cu deficiency response by means of a direct interaction with SPL7. Moreover, the double mutant kin17-1 spl7-2 displays an enhanced Cu-dependent phenotype involving growth arrest, oxidative stress, floral bud abortion, and pollen inviability. Taken together, the data presented here provide evidence for SPL7 and KIN17 protein interaction as a point of convergence in response to both Cu deficiency and oxidative stress.
Asunto(s)
Adaptación Fisiológica/efectos de los fármacos , Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Secuencia Conservada , Cobre/toxicidad , Proteínas de Unión al ADN/metabolismo , Factores de Transcripción/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/fisiología , Secuencia de Bases , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Modelos Biológicos , Datos de Secuencia Molecular , Mutación/genética , Proteínas Nucleares/metabolismo , Estrés Oxidativo/efectos de los fármacos , Fenotipo , Desarrollo de la Planta/efectos de los fármacos , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/metabolismo , Polen/efectos de los fármacos , Polen/metabolismo , Unión Proteica/efectos de los fármacos , Nicotiana/efectos de los fármacos , Nicotiana/metabolismoRESUMEN
SPL8 and miR156-targeted SPL genes are known to play an essential role in Arabidopsis anther development. Here we show that these SPL genes are also expressed within the developing gynoecium, where they redundantly control development of the female reproductive tract. Whereas the gynoecium morphology in the spl8 single mutant is largely normal, additional down-regulation of miR156-targeted SPL genes results in a shortened style and an apically swollen ovary narrowing onto an elongated gynophore. In particular, the septum does not form properly and lacks a transmitting tract. Loss of SPL8 function enhances the mutant phenotypes of ett, crc and spt, indicating a functional overlap between SPL8 and these genes in regulating gynoecium development. Furthermore, gynoecium development of 35S:MIR156b spl8-1 double mutants shows enhanced sensitivity to a polar auxin transport inhibitor, and the expression pattern of the auxin biosynthesis gene YUCCA4 is altered compared to wild-type. Our observations imply that SPL8 and miR156-targeted SPL genes control gynoecium patterning through interference with auxin homeostasis and signalling.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Flores/genética , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/metabolismo , MicroARNs/genética , Arabidopsis/citología , Arabidopsis/efectos de los fármacos , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/metabolismo , Tipificación del Cuerpo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Regulación hacia Abajo , Flores/citología , Flores/efectos de los fármacos , Flores/crecimiento & desarrollo , Homeostasis , MicroARNs/metabolismo , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Familia de Multigenes , Mutación , Especificidad de Órganos , Fenotipo , Ftalimidas/farmacología , Hojas de la Planta/citología , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Plantas Modificadas Genéticamente , Plantones/citología , Plantones/efectos de los fármacos , Plantones/genética , Plantones/crecimiento & desarrollo , Transducción de SeñalRESUMEN
BACKGROUND: The Arabidopsis SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE (SPL) transcription factor SPL7 reprograms cellular gene expression to adapt plant growth and cellular metabolism to copper (Cu) limited culture conditions. Plant cells require Cu to maintain essential processes, such as photosynthesis, scavenging reactive oxygen species, cell wall lignification and hormone sensing. More specifically, SPL7 activity promotes a high-affinity Cu-uptake system and optimizes Cu (re-)distribution to essential Cu-proteins by means of specific miRNAs targeting mRNA transcripts for those dispensable. However, the functional mechanism underlying SPL7 activation is still to be elucidated. As SPL7 transcript levels are largely non-responsive to Cu availability, post-translational modification seems an obvious possibility. Previously, it was reported that the SPL7 SBP domain does not bind to DNA in vitro in the presence of Cu ions and that SPL7 interacts with a kin17 domain protein to raise SPL7-target gene expression upon Cu deprivation. Here we report how additional conserved SPL7 protein domains may contribute to the Cu deficiency response in Arabidopsis. RESULTS: Cytological and biochemical approaches confirmed an operative transmembrane domain (TMD) and uncovered a dual localisation of SPL7 between the nucleus and an endomembrane system, most likely the endoplasmic reticulum (ER). This new perspective unveiled a possible link between Cu deficit and ER stress, a metabolic dysfunction found capable of inducing SPL7 targets in an SPL7-dependent manner. Moreover, in vivo protein-protein interaction assays revealed that SPL7 is able to homodimerize, probably mediated by the IRPGC domain. These observations, in combination with the constitutive activation of SPL7 targets, when ectopically expressing the N-terminal part of SPL7 including the SBP domain, shed some light on the mechanisms governing SPL7 function. CONCLUSIONS: Here, we propose a revised model of SPL7 activation and regulation. According to our results, SPL7 would be initially located to endomembranes and activated during ER stress as a result of Cu deficiency. Furthermore, we added the SPL7 dimerization in the presence of Cu ions as an additional regulatory mechanism to modulate the Cu deficiency response.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Cobre , Proteínas de Unión al ADN , Regulación de la Expresión Génica de las Plantas , Factores de Transcripción , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Núcleo Celular/metabolismo , Cobre/deficiencia , Cobre/metabolismo , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Estrés del Retículo Endoplásmico/fisiología , Estructura Terciaria de Proteína , Factores de Transcripción/química , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Copper and iron are essential micronutrients for most living organisms because they participate as cofactors in biological processes, including respiration, photosynthesis, and oxidative stress protection. In many eukaryotic organisms, including yeast (Saccharomyces cerevisiae) and mammals, copper and iron homeostases are highly interconnected; yet, such interdependence is not well established in higher plants. Here, we propose that COPT2, a high-affinity copper transport protein, functions under copper and iron deficiencies in Arabidopsis (Arabidopsis thaliana). COPT2 is a plasma membrane protein that functions in copper acquisition and distribution. Characterization of the COPT2 expression pattern indicates a synergic response to copper and iron limitation in roots. We characterized a knockout of COPT2, copt2-1, that leads to increased resistance to simultaneous copper and iron deficiencies, measured as reduced leaf chlorosis and improved maintenance of the photosynthetic apparatus. We propose that COPT2 could play a dual role under iron deficiency. First, COPT2 participates in the attenuation of copper deficiency responses driven by iron limitation, possibly to minimize further iron consumption. Second, global expression analyses of copt2-1 versus wild-type Arabidopsis plants indicate that low-phosphate responses increase in the mutant. These results open up new biotechnological approaches to fight iron deficiency in crops.
Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Proteínas de Transporte de Catión/genética , Regulación de la Expresión Génica de las Plantas , Deficiencias de Hierro , Fosfatos/metabolismo , Arabidopsis/fisiología , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Proteínas de Transporte de Catión/metabolismo , Cobre/metabolismo , Homeostasis , Hierro/metabolismo , Modelos Biológicos , Mutación , Fenotipo , Hojas de la Planta/genética , Hojas de la Planta/fisiología , Raíces de Plantas/genética , Raíces de Plantas/fisiología , Plantas Modificadas Genéticamente , Proteínas Recombinantes de Fusión , Proteínas SLC31 , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Plantones/genética , Plantones/fisiología , Análisis de Secuencia de ADN , Transducción de Señal , Regulación hacia ArribaRESUMEN
Understanding plant immune responses is complex because of the high interdependence among biological processes in homeostatic networks. Hence, the integration of environmental cues causes network rewiring that interferes with defense responses. Similarly, plants retain molecular signatures configured under abiotic stress periods to rapidly respond to recurrent stress, and these can alter immunity. Metabolome changes imposed by abiotic stressors are persistent, although their impact on defense remains to be clarified. In this study, we profiled metabolomes of Arabidopsis plants under several abiotic stress treatments applied individually or simultaneously to capture temporal trajectories in metabolite composition during adverse conditions and recovery. Further systemic analysis was performed to address the relevance of metabolome changes and extract central features to be tested in planta. Our results demonstrate irreversibility in major fractions of metabolome changes as a general pattern in response to abiotic stress periods. Functional analysis of metabolomes and co-abundance networks points to convergence in the reconfiguration of organic acid and secondary metabolite metabolism. Arabidopsis mutant lines for components related to these metabolic pathways showed altered defense capacities against different pathogens. Collectively, our data suggest that sustained metabolome changes configured in adverse environments can act as modulators of immune responses and provide evidence for a new layer of regulation in plant defense.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Metaboloma , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Estrés Fisiológico , Redes y Vías Metabólicas , Plantas/metabolismoRESUMEN
Copper (Cu), an essential redox active cofactor, participates in fundamental biological processes, but it becomes highly cytotoxic when present in excess. Therefore, living organisms have established suitable mechanisms to balance cellular and systemic Cu levels. An important strategy to maintain Cu homeostasis consists of regulating uptake and mobilization via the conserved family of CTR/COPT Cu transport proteins. In the model plant Arabidopsis thaliana, COPT1 protein mediates root Cu acquisition, whereas COPT5 protein functions in Cu mobilization from intracellular storage organelles. The function of these transporters becomes critical when environmental Cu bioavailability diminishes. However, little is know about the mechanisms that mediate plant Cu distribution. In this report, we present evidence supporting an important role for COPT6 in Arabidopsis Cu distribution. Similarly to COPT1 and COPT2, COPT6 fully complements yeast mutants defective in high-affinity Cu uptake and localizes to the plasma membrane of Arabidopsis cells. Whereas COPT2 mRNA is only up-regulated upon severe Cu deficiency, COPT6 transcript is expressed under Cu excess conditions and displays a more gradual increase in response to decreases in environmental Cu levels. Consistent with COPT6 expression in aerial vascular tissues and reproductive organs, copt6 mutant plants exhibit altered Cu distribution under Cu-deficient conditions, including increased Cu in rosette leaves but reduced Cu levels in seeds. This altered Cu distribution is fully rescued when the wild-type COPT6 gene is reintroduced into the copt6 mutant line. Taken together, these findings highlight the relevance of COPT6 in shoot Cu redistribution when environmental Cu is limited.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Cobre/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Transporte de Membrana/metabolismo , Secuencia de Aminoácidos , Arabidopsis/citología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Transporte Biológico , Membrana Celular/metabolismo , Cobre/deficiencia , Prueba de Complementación Genética , Homeostasis , Proteínas de Transporte de Membrana/genética , Datos de Secuencia Molecular , Mutagénesis Insercional , Hojas de la Planta/citología , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Raíces de Plantas/citología , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Brotes de la Planta/citología , Brotes de la Planta/genética , Brotes de la Planta/metabolismo , Haz Vascular de Plantas/citología , Haz Vascular de Plantas/genética , Haz Vascular de Plantas/metabolismo , Plantas Modificadas Genéticamente , Proteínas SLC31 , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Semillas/citología , Semillas/genética , Semillas/metabolismo , Alineación de Secuencia , Regulación hacia ArribaRESUMEN
The sessile lifestyle of plants requires an immediate response to environmental stressors that affect photosynthesis, growth, and crop yield. Here, we showed that three abiotic perturbations-heat, cold, and high light-triggered considerable changes in the expression signatures of 42 epitranscriptomic factors (writers, erasers, and readers) with putative chloroplast-associated functions that formed clusters of commonly expressed genes in Arabidopsis. The expression changes under all conditions were reversible upon deacclimation, identifying epitranscriptomic players as modulators in acclimation processes. Chloroplast dysfunctions, particularly those induced by the oxidative stress-inducing norflurazon in a largely GENOME UNCOUPLED-independent manner, triggered retrograde signals to remodel chloroplast-associated epitranscriptomic expression patterns. N6-methyladenosine (m6A) is known as the most prevalent RNA modification and impacts numerous developmental and physiological functions in living organisms. During cold treatment, expression of components of the primary nuclear m6A methyltransferase complex was upregulated, accompanied by a significant increase in cellular m6A mRNA marks. In the cold, the presence of FIP37, a core component of the writer complex, played an important role in positive regulation of thylakoid structure, photosynthetic functions, and accumulation of photosystem I, the Cytb6f complex, cyclic electron transport proteins, and Curvature Thylakoid1 but not that of photosystem II components and the chloroplast ATP synthase. Downregulation of FIP37 affected abundance, polysomal loading, and translation of cytosolic transcripts related to photosynthesis in the cold, suggesting m6A-dependent translational regulation of chloroplast functions. In summary, we identified multifaceted roles of the cellular m6A RNA methylome in coping with cold; these were predominantly associated with chloroplasts and served to stabilize photosynthesis.
Asunto(s)
Arabidopsis , ARN , ARN/metabolismo , Epigenoma , Luz , Fotosíntesis/genética , Cloroplastos/genética , Cloroplastos/metabolismo , Arabidopsis/metabolismoRESUMEN
Copper is an essential micronutrient that functions as a redox cofactor in multiple plant processes, including photosynthesis. Arabidopsis thaliana possesses a conserved family of CTR-like high-affinity copper transport proteins denoted as COPT1-5. COPT1, the only family member that is functionally characterized, participates in plant copper acquisition. However, little is known about the function of the other Arabidopsis COPT proteins in the transport and distribution of copper. Here, we show that a functional fusion of COPT5 to the green fluorescent protein localizes in Arabidopsis cells to the prevacuolar compartment. Plants defective in COPT5 do not exhibit any significant phenotype under copper-sufficient conditions, but their growth is compromised under copper limitation. Under extreme copper deficiency, two independent copt5 knockout mutant lines exhibit severe defects in vegetative growth and root elongation, low chlorophyll content, and impairment in the photosynthetic electron transfer. All these phenotypes are rescued when the wild-type copy of the COPT5 gene is retransformed into a copt5 knockout line or when copper, but not other metals, are added to the medium. COPT5 is expressed in vascular tissues, with elevated levels in roots. Taken together, these results suggest that COPT5 plays an important role in the plant response to environmental copper scarcity, probably by remobilizing copper from prevacuolar vesicles, which could act as internal stores or recycling vesicles to provide the metal cofactor to key copper-dependent processes such as photosynthesis.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Transporte de Catión/metabolismo , Cobre/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Secuencia de Bases , Proteínas de Transporte de Catión/genética , Cobre/deficiencia , ADN de Plantas/genética , Transporte de Electrón , Técnicas de Inactivación de Genes , Genes de Plantas , Prueba de Complementación Genética , Proteínas Fluorescentes Verdes/genética , Proteínas Fluorescentes Verdes/metabolismo , Fotosíntesis , Plantas Modificadas Genéticamente , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Proteínas SLC31 , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Vacuolas/metabolismoRESUMEN
Copper (Cu) and iron (Fe) constitute fundamental nutrients for plant biology but are often limited due to low bioavailability. Unlike responses to single Cu or Fe deprivation, the consequences of simultaneous Cu and Fe deficiency have not yet been fully deciphered. Previously, it was demonstrated that Cu and Fe deficiency applied in combination imposes transcriptome, proteome, and metabolome changes different from those triggered under each deficiency individually. Here, we evaluated the effect of alternative splicing (AS) on the transcriptome of rosette leaves under single and simultaneous Cu and Fe deficiency. Differentially spliced genes (DSGs) and differentially expressed genes (DEGs) coincided in number (2,600 approx.) although the overlapping fraction was minimal (15%). Functional annotation of changes exclusively detected under simultaneous Cu and Fe deficiency revealed that DEGs participated in general stress responses and translation, while DSGs were involved in metabolic reactions, especially amino acid biosynthesis. Interestingly, transcripts encoding central features for tryptophan (Trp) and asparagine (Asn) synthesis - two significantly altered metabolites under simultaneous Cu and Fe deficiency - underwent exclusive intron retention events under the double deficiency. However, transcript and protein amounts for these enzymes did not correlate with Trp and Asn concentration. In consequence, we propose that AS might act as a regulatory mechanism to modify the stability and/or functionality of the enzymes and therefore fine-tune amino acid production during the combinatorial response to simultaneous Cu and Fe deficiency.
RESUMEN
Faster onset of photoprotection could potentially increase biomass accumulation. Indeed, this has been realized in tobacco VPZ lines by overexpression of three photoprotective proteins in parallel. To explore the range of application of this approach, we generated Arabidopsis VPZ lines. These lines triggered photoprotection more rapidly, but growth rate and biomass accumulation were impaired under fluctuating light. This implies that the strategy might interfere with other mechanisms controlling excitation energy distribution, or with source-sink relationships or plastid signalling.
Asunto(s)
Arabidopsis/efectos de la radiación , Biomasa , Fotosíntesis/efectos de la radiación , Proteínas de Plantas/genética , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente/genética , Plantas Modificadas Genéticamente/crecimiento & desarrollo , Plantas Modificadas Genéticamente/metabolismo , Plantas Modificadas Genéticamente/efectos de la radiación , Nicotiana/genética , Transcripción GenéticaRESUMEN
Plant metabolism is broadly reprogrammed during acclimation to abiotic changes. Most previous studies have focused on transitions from standard to single stressful conditions. Here, we systematically analyze acclimation processes to levels of light, heat, and cold stress that subtly alter physiological parameters and assess their reversibility during de-acclimation. Metabolome and transcriptome changes were monitored at 11 different time points. Unlike transcriptome changes, most alterations in metabolite levels did not readily return to baseline values, except in the case of cold acclimation. Similar regulatory networks operate during (de-)acclimation to high light and cold, whereas heat and high-light responses exhibit similar dynamics, as determined by surprisal and conditional network analyses. In all acclimation models tested here, super-hubs in conditional transcriptome networks are enriched for components involved in translation, particularly ribosomes. Hence, we suggest that the ribosome serves as a common central hub for the control of three different (de-)acclimation responses.