RESUMEN
D6 PROTEIN KINASE (D6PK) is a polarly localized plasma-membrane-associated kinase from Arabidopsis thaliana that activates polarly distributed PIN-FORMED auxin transporters. D6PK moves rapidly to and from the plasma membrane, independent of its PIN-FORMED targets. The middle D6PK domain, an insertion between kinase subdomains VII and VIII, is required and sufficient for association and polarity of the D6PK plasma membrane. How D6PK polarity is established and maintained remains to be shown. Here we show that cysteines from repeated middle domain CXX(X)P motifs are S-acylated and required for D6PK membrane association. While D6PK S-acylation is not detectably regulated during intracellular transport, phosphorylation of adjacent serine residues, in part in dependence on the upstream 3-PHOSPHOINOSITIDE-DEPENDENT PROTEIN KINASE, promotes D6PK transport, controls D6PK residence time at the plasma membrane and prevents its lateral diffusion. We thus identify new mechanisms for the regulation of D6PK plasma membrane interaction and polarity.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Fosforilación , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Membrana Celular/metabolismoRESUMEN
There are growing doubts about the true role of the common mycorrhizal networks (CMN or wood wide web) connecting the roots of trees in forests. We question the claims of a substantial carbon transfer from 'mother trees' to their offspring and nearby seedlings through the CMN. Recent reviews show that evidence for the 'mother tree concept' is inconclusive or absent. The origin of this concept seems to stem from a desire to humanize plant life but can lead to misunderstandings and false interpretations and may eventually harm rather than help the commendable cause of preserving forests. Two recent books serve as examples: The Hidden Life of Trees and Finding the Mother Tree.
Asunto(s)
Micorrizas , Árboles , Humanos , Bosques , Hongos , Raíces de Plantas/microbiología , Plantas , SueloRESUMEN
GATAs are evolutionarily conserved zinc-finger transcription factors from eukaryotes. In plants, GATAs can be subdivided into four classes, A-D, based on their DNA-binding domain, and into further subclasses based on additional protein motifs. B-GATAs with a so-called leucine-leucine-methionine (LLM)-domain can already be found in algae. In angiosperms, the B-GATA family is expanded and can be subdivided in to LLM- or HAN-domain B-GATAs. Both, the LLM- and the HAN-domain are conserved domains of unknown biochemical function. Interestingly, the B-GATA family in the liverwort Marchantia polymorpha and the moss Physcomitrium patens is restricted to one and four family members, respectively. And, in contrast to vascular plants, the bryophyte B-GATAs contain a HAN- as well as an LLM-domain. Here, we characterise mutants of the single B-GATA from Marchantia polymorpha. We reveal that this mutant has defects in thallus growth and in gemma formation. Transcriptomic studies uncover that the B-GATA mutant displays a constitutive high-light (HL) stress response, a phenotype that we then also confirm in mutants of Arabidopsis thaliana LLM-domain B-GATAs, suggesting that the B-GATAs have a protective role towards HL stress.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Marchantia , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Factores de Transcripción GATA/genética , Factores de Transcripción GATA/metabolismo , Marchantia/genética , LeucinaRESUMEN
Plants perceive the direction of gravity during skotomorphogenic growth, and of gravity and light during photomorphogenic growth. Gravity perception occurs through the sedimentation of starch granules in shoot endodermal and root columella cells. In this study, we demonstrate that the Arabidopsis thaliana GATA factors GNC (GATA, NITRATE-INDUCIBLE, CARBON METABOLISM-INVOLVED) and GNL/CGA1 (GNC-LIKE/CYTOKININ-RESPONSIVE GATA1) repress starch granule growth and amyloplast differentiation in endodermal cells. In our comprehensive study, we analysed gravitropic responses in the shoot, root and hypocotyl. We performed an RNA-seq analysis, used advanced microscopy techniques to examine starch granule size, number and morphology and quantified transitory starch degradation patterns. Using transmission electron microscopy, we examined amyloplast development. Our results indicate that the altered gravitropic responses in hypocotyls, shoots and roots of gnc gnl mutants and GNL overexpressors are due to the differential accumulation of starch granules observed in the GATA genotypes. At the whole-plant level, GNC and GNL play a more complex role in starch synthesis, degradation and starch granule initiation. Our findings suggest that the light-regulated GNC and GNL help balance phototropic and gravitropic growth responses after the transition from skotomorphogenesis to photomorphogenesis by repressing the growth of starch granules.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Factores de Transcripción GATA/genética , Factores de Transcripción GATA/metabolismo , Proteínas de Arabidopsis/metabolismo , Citocininas/metabolismo , Almidón/metabolismo , Gravitropismo/genética , Mutación/genética , Raíces de Plantas/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Plants are an indispensable cornerstone of sustainable global food supply. While immense progress has been made in decoding the genomes of crops in recent decades, the composition of their proteomes, the entirety of all expressed proteins of a species, is virtually unknown. In contrast to the model plant Arabidopsis thaliana, proteomic analyses of crop plants have often been hindered by the presence of extreme concentrations of secondary metabolites such as pigments, phenolic compounds, lipids, carbohydrates or terpenes. As a consequence, crop proteomic experiments have, thus far, required individually optimized protein extraction protocols to obtain samples of acceptable quality for downstream analysis by liquid chromatography tandem mass spectrometry (LC-MS/MS). In this article, we present a universal protein extraction protocol originally developed for gel-based experiments and combined it with an automated single-pot solid-phase-enhanced sample preparation (SP3) protocol on a liquid handling robot to prepare high-quality samples for proteomic analysis of crop plants. We also report an automated offline peptide separation protocol and optimized micro-LC-MS/MS conditions that enables the identification and quantification of ~10,000 proteins from plant tissue within 6 h of instrument time. We illustrate the utility of the workflow by analyzing the proteomes of mature tomato fruits to an unprecedented depth. The data demonstrate the robustness of the approach which we propose for use in upcoming large-scale projects that aim to map crop tissue proteomes.
Asunto(s)
Proteoma , Proteómica , Proteómica/métodos , Cromatografía Liquida/métodos , Proteoma/análisis , Espectrometría de Masas en Tándem/métodos , Productos AgrícolasRESUMEN
GATA factors are evolutionarily conserved transcription factors that are found in animals, fungi, and plants. Compared to that of animals, the size of the plant GATA family is increased. In angiosperms, four main GATA classes and seven structural subfamilies can be defined. In recent years, knowledge about the biological role and regulation of plant GATAs has substantially improved. Individual family members have been implicated in the regulation of photomorphogenic growth, chlorophyll biosynthesis, chloroplast development, photosynthesis, and stomata formation, as well as root, leaf, and flower development. In this review, we summarize the current knowledge of plant GATA factors. Using phylogenomic analysis, we trace the evolutionary origin of the GATA classes in the green lineage and examine their relationship to animal and fungal GATAs. Finally, we speculate about a possible conservation of GATA-regulated functions across the animal, fungal, and plant kingdoms.
Asunto(s)
Factores de Transcripción GATA , Factores de Transcripción , Animales , Biología , Evolución Molecular , Factores de Transcripción GATA/genética , Factores de Transcripción GATA/metabolismo , Regulación de la Expresión Génica de las Plantas , Filogenia , Hojas de la Planta/metabolismoRESUMEN
The directional transport of the phytohormone auxin is required for proper plant development and tropic growth. Auxin cell-to-cell transport gains directionality through the polar distribution of 'canonical' long PIN-FORMED (PIN) auxin efflux carriers. In recent years, AGC kinases, MAP kinases, Ca2+/CALMODULIN-DEPENDENT PROTEIN KINASE-RELATED KINASEs and receptor kinases have been implicated in the control of PIN activity, polarity and trafficking. In this review, we summarize the current knowledge in understanding the posttranslational regulation of PINs by these different protein kinase families. The proposed regulation of PINs by AGC kinases after salt stress and by the stress-activated MAP kinases suggest that abiotic and biotic stress factors may modulate auxin transport and thereby plant growth.
Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Fosforilación , Raíces de Plantas/metabolismo , Proteínas Quinasas/metabolismoRESUMEN
From embryogenesis to fruit formation, almost every aspect of plant development and differentiation is controlled by the cellular accumulation or depletion of auxin from cells and tissues. The respective auxin maxima and minima are generated by cell-to-cell auxin transport via transporter proteins. Differential auxin accumulation as a result of such transport processes dynamically regulates auxin distribution during differentiation. In this review, we introduce all auxin transporter (families) identified to date and discuss the knowledge on prominent family members, namely, the PIN-FORMED exporters, ATP-binding cassette B (ABCB)-type transporters, and AUX1/LAX importers. We then concentrate on the biochemical features of these transporters and their regulation by posttranslational modifications and interactors.
Asunto(s)
Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos , Transporte Biológico , Humanos , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismoRESUMEN
Angiosperms have evolved the phloem for the long-distance transport of metabolites. The complex process of phloem development involves genes that only occur in vascular plant lineages. For example, in Arabidopsis thaliana, the BREVIS RADIX (BRX) gene is required for continuous root protophloem differentiation, together with PROTEIN KINASE ASSOCIATED WITH BRX (PAX). BRX and its BRX-LIKE (BRXL) homologs are composed of four highly conserved domains including the signature tandem BRX domains that are separated by variable spacers. Nevertheless, BRX family proteins have functionally diverged. For instance, BRXL2 can only partially replace BRX in the root protophloem. This divergence is reflected in physiologically relevant differences in protein behavior, such as auxin-induced plasma membrane dissociation of BRX, which is not observed for BRXL2. Here we dissected the differential functions of BRX family proteins using a set of amino acid substitutions and domain swaps. Our data suggest that the plasma membrane-associated tandem BRX domains are both necessary and sufficient to convey the biological outputs of BRX function and therefore constitute an important regulatory entity. Moreover, PAX target phosphosites in the linker between the two BRX domains mediate the auxin-induced plasma membrane dissociation. Engineering these sites into BRXL2 renders this modified protein auxin-responsive and thereby increases its biological activity in the root protophloem context.
Asunto(s)
Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Animales , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Membrana Celular/metabolismo , Femenino , Regulación de la Expresión Génica de las Plantas , Familia de Multigenes , Oocitos/metabolismo , Plantas Modificadas Genéticamente , Dominios Proteicos , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Selaginellaceae/química , Xenopus laevisRESUMEN
N-1-naphthylphthalamic acid (NPA) is a key inhibitor of directional (polar) transport of the hormone auxin in plants. For decades, it has been a pivotal tool in elucidating the unique polar auxin transport-based processes underlying plant growth and development. Its exact mode of action has long been sought after and is still being debated, with prevailing mechanistic schemes describing only indirect connections between NPA and the main transporters responsible for directional transport, namely PIN auxin exporters. Here we present data supporting a model in which NPA associates with PINs in a more direct manner than hitherto postulated. We show that NPA inhibits PIN activity in a heterologous oocyte system and that expression of NPA-sensitive PINs in plant, yeast, and oocyte membranes leads to specific saturable NPA binding. We thus propose that PINs are a bona fide NPA target. This offers a straightforward molecular basis for NPA inhibition of PIN-dependent auxin transport and a logical parsimonious explanation for the known physiological effects of NPA on plant growth, as well as an alternative hypothesis to interpret past and future results. We also introduce PIN dimerization and describe an effect of NPA on this, suggesting that NPA binding could be exploited to gain insights into structural aspects of PINs related to their transport mechanism.
Asunto(s)
Transporte Biológico Activo/efectos de los fármacos , Ácidos Indolacéticos/metabolismo , Ftalimidas/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Proteínas de Plantas/metabolismo , Animales , Arabidopsis/efectos de los fármacos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Transporte Biológico Activo/genética , Dimerización , Espectrometría de Masas , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Oocitos/efectos de los fármacos , Fosforilación , Ftalimidas/farmacología , Reguladores del Crecimiento de las Plantas/antagonistas & inhibidores , Reguladores del Crecimiento de las Plantas/genética , Proteínas de Plantas/genética , Saccharomyces cerevisiae/metabolismo , Nicotiana/efectos de los fármacos , Nicotiana/metabolismo , XenopusRESUMEN
Plant growth and development are regulated by a tightly controlled interplay between cell division, cell expansion and cell differentiation during the entire plant life cycle from seed germination to maturity and seed propagation. To explore some of the underlying molecular mechanisms in more detail, we selected different aerial tissue types of the model plant Arabidopsis thaliana, namely rosette leaf, flower and silique/seed and performed proteomic, phosphoproteomic and transcriptomic analyses of sequential growth stages using tandem mass tag-based mass spectrometry and RNA sequencing. With this exploratory multi-omics dataset, development dynamics of photosynthetic tissues can be investigated from different angles. As expected, we found progressive global expression changes between growth stages for all three omics types and often but not always corresponding expression patterns for individual genes on transcript, protein and phosphorylation site level. The biggest difference between proteomic- and transcriptomic-based expression information could be observed for seed samples. Proteomic and transcriptomic data is available via ProteomeXchange and ArrayExpress with the respective identifiers PXD018814 and E-MTAB-7978.
Asunto(s)
Arabidopsis , Proteoma , Arabidopsis/genética , Perfilación de la Expresión Génica , Proteoma/genética , Proteómica , TranscriptomaRESUMEN
Seed dormancy is an adaptive trait defining where and when plants are established. Diverse signals from the environment are used to decide when to initiate seed germination, a process driven by the expansion of cells within the embryo. How these signals are integrated and transduced into the biomechanical changes that drive embryo growth remains poorly understood. Using Arabidopsis seeds, we demonstrate that cell-wall-loosening EXPANSIN (EXPA) genes promote gibberellic acid (GA)-mediated germination, identifying EXPAs as downstream molecular targets of this developmental phase transition. Molecular interaction screening identified transcription factors (TFs) that bind to both EXPA promoter fragments and DELLA GA-response regulators. A subset of these TFs is targeted each by nitric oxide (NO) and the phytochrome-interacting TF PIL5. This molecular interaction network therefore directly links the perception of an external environmental signal (light) and internal hormonal signals (GA and NO) with downstream germination-driving EXPA gene expression. Experimental validation of this network established that many of these TFs mediate GA-regulated germination, including TCP14/15, RAP2.2/2.3/2.12, and ZML1. The reduced germination phenotype of the tcp14 tcp15 mutant seed was partially rescued through ectopic expression of their direct target EXPA9. The GA-mediated control of germination by TCP14/15 is regulated through EXPA-mediated control of cell wall loosening, providing a mechanistic explanation for this phenotype and a previously undescribed role for TCPs in the control of cell expansion. This network reveals the paths of signal integration that culminate in seed germination and provides a resource to uncover links between the genetic and biomechanical bases of plant growth.
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Arabidopsis/metabolismo , Germinación/fisiología , Semillas/metabolismo , Ácido Abscísico/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Regulación de la Expresión Génica de las Plantas/genética , Germinación/genética , Giberelinas/metabolismo , Fitocromo/metabolismo , Latencia en las Plantas/genética , Latencia en las Plantas/fisiología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Regiones Promotoras Genéticas/genética , Semillas/genética , Factores de Transcripción/metabolismoRESUMEN
Plants are essential for life and are extremely diverse organisms with unique molecular capabilities1. Here we present a quantitative atlas of the transcriptomes, proteomes and phosphoproteomes of 30 tissues of the model plant Arabidopsis thaliana. Our analysis provides initial answers to how many genes exist as proteins (more than 18,000), where they are expressed, in which approximate quantities (a dynamic range of more than six orders of magnitude) and to what extent they are phosphorylated (over 43,000 sites). We present examples of how the data may be used, such as to discover proteins that are translated from short open-reading frames, to uncover sequence motifs that are involved in the regulation of protein production, and to identify tissue-specific protein complexes or phosphorylation-mediated signalling events. Interactive access to this resource for the plant community is provided by the ProteomicsDB and ATHENA databases, which include powerful bioinformatics tools to explore and characterize Arabidopsis proteins, their modifications and interactions.
Asunto(s)
Proteínas de Arabidopsis/análisis , Proteínas de Arabidopsis/química , Arabidopsis/química , Espectrometría de Masas , Proteoma/análisis , Proteoma/química , Proteómica , Secuencias de Aminoácidos , Arabidopsis/anatomía & histología , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/biosíntesis , Proteínas de Arabidopsis/genética , Bases de Datos de Proteínas , Conjuntos de Datos como Asunto , Regulación de la Expresión Génica de las Plantas , Anotación de Secuencia Molecular , Sistemas de Lectura Abierta , Especificidad de Órganos , Fosfoproteínas/análisis , Fosfoproteínas/química , Fosfoproteínas/genética , Fosforilación , Proteoma/biosíntesis , Proteoma/genética , ARN Mensajero/análisis , ARN Mensajero/biosíntesis , ARN Mensajero/genética , TranscriptomaRESUMEN
DELLA proteins are repressors of the gibberellin (GA) hormone signaling pathway that act mainly by regulating transcription factor activities in plants. GAs induce DELLA repressor protein degradation and thereby control a number of critical developmental processes as well as responses to stresses such as cold. The strong effect of cold temperatures on many physiological processes has rendered it difficult to assess, based on phenotypic criteria, the role of GA and DELLAs in plant growth during cold stress. Here, we uncover substantial differences in the GA transcriptomes between plants grown at ambient temperature (21°C) and plants exposed to cold stress (4°C) in Arabidopsis (Arabidopsis thaliana). We further identify over 250, to the largest extent previously unknown, DELLA-transcription factor interactions using the yeast two-hybrid system. By integrating both data sets, we reveal that most members of the nine-member GRF (GROWTH REGULATORY FACTOR) transcription factor family are DELLA interactors and, at the same time, that several GRF genes are targets of DELLA-modulated transcription after exposure to cold stress. We find that plants with altered GRF dosage are differentially sensitive to the manipulation of GA and hence DELLA levels, also after cold stress, and identify a subset of cold stress-responsive genes that qualify as targets of this DELLA-GRF regulatory module.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Respuesta al Choque por Frío , Factores de Transcripción/metabolismo , Arabidopsis/citología , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Recuento de Células , Tamaño de la Célula , Respuesta al Choque por Frío/efectos de los fármacos , Respuesta al Choque por Frío/genética , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Giberelinas/metabolismo , Giberelinas/farmacología , MicroARNs/genética , MicroARNs/metabolismo , Hojas de la Planta/efectos de los fármacos , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Plantas Modificadas Genéticamente , Unión Proteica/efectos de los fármacos , Transducción de Señal , Transcriptoma/efectos de los fármacos , Transcriptoma/genética , Triazoles/farmacologíaRESUMEN
Exocyst is a heterooctameric protein complex crucial for the tethering of secretory vesicles to the plasma membrane during exocytosis. Compared to other eukaryotes, exocyst subunit EXO70 is represented by many isoforms in land plants whose cell biological and biological roles, as well as modes of regulation remain largely unknown. Here, we present data on the phospho-regulation of exocyst isoform EXO70C2, which we previously identified as a putative negative regulator of exocyst function in pollen tube growth. A comprehensive phosphoproteomic analysis revealed phosphorylation of EXO70C2 at multiple sites. We have now performed localization and functional studies of phospho-dead and phospho-mimetic variants of Arabidopsis EXO70C2 in transiently transformed tobacco pollen tubes and stably transformed Arabidopsis wild type and exo70C2 mutant plants. Our data reveal a dose-dependent effect of AtEXO70C2 overexpression on pollen tube growth rate and cellular architecture. We show that changes of the AtEXO70C2 phosphorylation status lead to distinct outcomes in wild type and exo70c2 mutant cells, suggesting a complex regulatory pattern. On the other side, phosphorylation does not affect the cytoplasmic localization of AtEXO70C2 or its interaction with putative secretion inhibitor ROH1 in the yeast two-hybrid system.
RESUMEN
Certain regions on the surfaces of developing pollen grains exhibit very limited deposition of pollen wall exine. These regions give rise to pollen apertures, which are highly diverse in their patterns and specific for individual species. Arabidopsis thaliana pollen develops three equidistant longitudinal apertures. The precision of aperture formation suggests that, to create them, pollen employs robust mechanisms that generate distinct cellular domains. To identify players involved in this mechanism, we screened natural Arabidopsis accessions and discovered one accession, Martuba, whose apertures form abnormally due to the disruption of the protein kinase D6PKL3. During pollen development, D6PKL3 accumulates at the three plasma membrane domains underlying future aperture sites. Both D6PKL3 localization and aperture formation require kinase activity. Proper D6PKL3 localization is also dependent on a polybasic motif for phosphoinositide interactions, and we identified two phosphoinositides that are specifically enriched at the future aperture sites. The other known aperture factor, INAPERTURATE POLLEN1, fails to aggregate at the aperture sites in d6pkl3 mutants, changes its localization when D6PKL3 is mislocalized, and, in turn, affects D6PKL3 localization. The discovery of aperture factors provides important insights into the mechanisms cells utilize to generate distinct membrane domains, develop cell polarity, and pattern their surfaces.
Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Membrana Celular/metabolismo , Polen/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Membrana Celular/genética , Mutación , Polen/genéticaRESUMEN
The ubiquitin-related protein NEDD8 is conjugated and deconjugated to and from proteins in processes related to ubiquitin conjugation and deconjugation. Neddylation is a well-studied posttranslational modification of Cullin-RING E3 ligases (CRLs). Biochemical and structural studies aiming at understanding the role of NEDD8 in CRL function have now resulted in a convincing model of how neddylation and deneddylation antagonistically regulate CRL stability, conformation, activity as well as degradation substrate receptor exchange. Studies of the Arabidopsis thaliana deneddylation-deficient den1 mutant led to the identification of many low abundant, non-Cullin NEDD8 conjugates. Examination of neddylated AUXIN RESISTANT1 (AXR1), a prominent neddylated protein in den1, suggests, however, that AXR1 neddylation may be an auto-catalytic side-reaction of Cullin-targeted neddylation and that DEN1 may serve to antagonize non-productive, auto-neddylation from substrates to provide free NEDD8 for CRL regulation.