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1.
Proc Natl Acad Sci U S A ; 113(10): 2768-73, 2016 Mar 08.
Artículo en Inglés | MEDLINE | ID: mdl-26888284

RESUMEN

The shaping of organs in plants depends on the intercellular flow of the phytohormone auxin, of which the directional signaling is determined by the polar subcellular localization of PIN-FORMED (PIN) auxin transport proteins. Phosphorylation dynamics of PIN proteins are affected by the protein phosphatase 2A (PP2A) and the PINOID kinase, which act antagonistically to mediate their apical-basal polar delivery. Here, we identified the ROTUNDA3 (RON3) protein as a regulator of the PP2A phosphatase activity in Arabidopsis thaliana. The RON3 gene was map-based cloned starting from the ron3-1 leaf mutant and found to be a unique, plant-specific gene coding for a protein with high and dispersed proline content. The ron3-1 and ron3-2 mutant phenotypes [i.e., reduced apical dominance, primary root length, lateral root emergence, and growth; increased ectopic stages II, IV, and V lateral root primordia; decreased auxin maxima in indole-3-acetic acid (IAA)-treated root apical meristems; hypergravitropic root growth and response; increased IAA levels in shoot apices; and reduced auxin accumulation in root meristems] support a role for RON3 in auxin biology. The affinity-purified PP2A complex with RON3 as bait suggested that RON3 might act in PIN transporter trafficking. Indeed, pharmacological interference with vesicle trafficking processes revealed that single ron3-2 and double ron3-2 rcn1 mutants have altered PIN polarity and endocytosis in specific cells. Our data indicate that RON3 contributes to auxin-mediated development by playing a role in PIN recycling and polarity establishment through regulation of the PP2A complex activity.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Proteína Fosfatasa 2/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Regulación de la Expresión Génica de las Plantas , Hibridación in Situ , Proteínas de Transporte de Membrana/genética , Microscopía Confocal , Modelos Biológicos , Mutación , Hojas de la Planta/genética , Hojas de la Planta/crecimiento & desarrollo , Hojas de la Planta/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
2.
Nature ; 485(7396): 119-22, 2012 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-22504182

RESUMEN

The phytohormone auxin acts as a prominent signal, providing, by its local accumulation or depletion in selected cells, a spatial and temporal reference for changes in the developmental program. The distribution of auxin depends on both auxin metabolism (biosynthesis, conjugation and degradation) and cellular auxin transport. We identified in silico a novel putative auxin transport facilitator family, called PIN-LIKES (PILS). Here we illustrate that PILS proteins are required for auxin-dependent regulation of plant growth by determining the cellular sensitivity to auxin. PILS proteins regulate intracellular auxin accumulation at the endoplasmic reticulum and thus auxin availability for nuclear auxin signalling. PILS activity affects the level of endogenous auxin indole-3-acetic acid (IAA), presumably via intracellular accumulation and metabolism. Our findings reveal that the transport machinery to compartmentalize auxin within the cell is of an unexpected molecular complexity and demonstrate this compartmentalization to be functionally important for a number of developmental processes.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas Portadoras/metabolismo , Homeostasis , Ácidos Indolacéticos/metabolismo , Espacio Intracelular/metabolismo , Familia de Multigenes , Arabidopsis/citología , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Proteínas Portadoras/genética , Retículo Endoplásmico/metabolismo , Genes de Plantas/genética , Germinación , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Plantas Modificadas Genéticamente , Regiones Promotoras Genéticas/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo
3.
Planta ; 245(1): 15-30, 2017 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-27541497

RESUMEN

MAIN CONCLUSION: This study showed that Bacillus amyloliquefaciens UCMB5113 colonizing Arabidopsis roots changed root structure and promoted growth implying the usability of this strain as a novel tool to support sustainable crop production. Root architecture plays a crucial role for plants to ensure uptake of water, minerals and nutrients and to provide anchorage in the soil. The root is a dynamic structure with plastic growth and branching depending on the continuous integration of internal and environmental factors. The rhizosphere contains a complex microbiota, where some microbes can colonize plant roots and support growth and stress tolerance. Here, we report that the rhizobacterium Bacillus amyloliquefaciens subsp. plantarum UCMB5113 stimulated the growth of Arabidopsis thaliana Col-0 by increased lateral root outgrowth and elongation and root-hair formation, although primary root elongation was inhibited. In addition, the growth of the above ground tissues was stimulated by UCMB5113. Specific hormone reporter gene lines were tested which suggested a role for at least auxin and cytokinin signaling during rhizobacterial modulation of Arabidopsis root architecture. UCMB5113 produced cytokinins and indole-3-acetic acid, and the formation of the latter was stimulated by root exudates and tryptophan. The plant growth promotion effect by UCMB5113 did not appear to depend on jasmonic acid in contrast to the disease suppression effect in plants. UCMB5113 exudates inhibited primary root growth, while a semi-purified lipopeptide fraction did not and resulted in the overall growth promotion indicating an interplay of many different bacterial compounds that affect the root growth of the host plant. This study illustrates that beneficial microbes interact with plants in root development via classic and novel signals.


Asunto(s)
Arabidopsis/crecimiento & desarrollo , Arabidopsis/microbiología , Bacillus amyloliquefaciens/fisiología , Interacciones Huésped-Patógeno , Arabidopsis/efectos de los fármacos , Bacillus amyloliquefaciens/efectos de los fármacos , Brasinoesteroides/farmacología , Citocininas/farmacología , Giberelinas/farmacología , Interacciones Huésped-Patógeno/efectos de los fármacos , Ácidos Indolacéticos/farmacología , Lipopéptidos/farmacología , Reguladores del Crecimiento de las Plantas/farmacología , Raíces de Plantas/anatomía & histología , Raíces de Plantas/efectos de los fármacos , Plantones/efectos de los fármacos , Plantones/crecimiento & desarrollo
4.
New Phytol ; 211(1): 65-74, 2016 07.
Artículo en Inglés | MEDLINE | ID: mdl-27240710

RESUMEN

Plant development mediated by the phytohormone auxin depends on tightly controlled cellular auxin levels at its target tissue that are largely established by intercellular and intracellular auxin transport mediated by PIN auxin transporters. Among the eight members of the Arabidopsis PIN family, PIN6 is the least characterized candidate. In this study we generated functional, fluorescent protein-tagged PIN6 proteins and performed comprehensive analysis of their subcellular localization and also performed a detailed functional characterization of PIN6 and its developmental roles. The localization study of PIN6 revealed a dual localization at the plasma membrane (PM) and endoplasmic reticulum (ER). Transport and metabolic profiling assays in cultured cells and Arabidopsis strongly suggest that PIN6 mediates both auxin transport across the PM and intracellular auxin homeostasis, including the regulation of free auxin and auxin conjugates levels. As evidenced by the loss- and gain-of-function analysis, the complex function of PIN6 in auxin transport and homeostasis is required for auxin distribution during lateral and adventitious root organogenesis and for progression of these developmental processes. These results illustrate a unique position of PIN6 within the family of PIN auxin transporters and further add complexity to the developmentally crucial process of auxin transport.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Membrana Celular/metabolismo , Retículo Endoplásmico/metabolismo , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Evolución Molecular , Homeostasis , Proteínas de Transporte de Membrana/genética , Filogenia , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/metabolismo , Plantas Modificadas Genéticamente
5.
Plant Physiol ; 167(1): 102-17, 2015 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-25475669

RESUMEN

Vitamin B(6) (pyridoxal 5'-phosphate) is an essential cofactor of many metabolic enzymes. Plants biosynthesize the vitamin de novo employing two enzymes, pyridoxine synthase1 (PDX1) and PDX2. In Arabidopsis (Arabidopsis thaliana), there are two catalytically active paralogs of PDX1 (PDX1.1 and PDX1.3) producing the vitamin at comparable rates. Since single mutants are viable but the pdx1.1 pdx1.3 double mutant is lethal, the corresponding enzymes seem redundant. However, the single mutants exhibit substantial phenotypic differences, particularly at the level of root development, with pdx1.3 being more impaired than pdx1.1. Here, we investigate the differential regulation of PDX1.1 and PDX1.3 by identifying factors involved in their disparate phenotypes. Swapped-promoter experiments clarify the presence of distinct regulatory elements in the upstream regions of both genes. Exogenous sucrose (Suc) triggers impaired ethylene production in both mutants but is more severe in pdx1.3 than in pdx1.1. Interestingly, Suc specifically represses PDX1.1 expression, accounting for the stronger vitamin B6 deficit in pdx1.3 compared with pdx1.1. Surprisingly, Suc enhances auxin levels in pdx1.1, whereas the levels are diminished in pdx1.3. In the case of pdx1.3, the previously reported reduced meristem activity combined with the impaired ethylene and auxin levels manifest the specific root developmental defects. Moreover, it is the deficit in ethylene production and/or signaling that triggers this outcome. On the other hand, we hypothesize that it is the increased auxin content of pdx1.1 that is responsible for the root developmental defects observed therein. We conclude that PDX1.1 and PDX1.3 play partially nonredundant roles and are differentially regulated as manifested in disparate root growth impairment morphologies.


Asunto(s)
Arabidopsis/crecimiento & desarrollo , Reguladores del Crecimiento de las Plantas/fisiología , Raíces de Plantas/crecimiento & desarrollo , Vitamina B 6/fisiología , Arabidopsis/metabolismo , Arabidopsis/fisiología , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/fisiología , Liasas de Carbono-Nitrógeno , Homeostasis/fisiología , Ácidos Indolacéticos/metabolismo , Transferasas de Grupos Nitrogenados/metabolismo , Transferasas de Grupos Nitrogenados/fisiología , Fenotipo , Reguladores del Crecimiento de las Plantas/metabolismo , Vitamina B 6/biosíntesis
6.
Ann Bot ; 117(6): 1037-44, 2016 05.
Artículo en Inglés | MEDLINE | ID: mdl-27098087

RESUMEN

BACKGROUND AND AIMS: The typical rootless linear shoots of aquatic carnivorous plants exhibit clear, steep polarity associated with very rapid apical shoot growth. The aim of this study was to determine how auxin and cytokinin contents are related to polarity and shoot growth in such plants. METHODS: The main auxin and cytokinin metabolites in separated shoot segments and turions of two carnivorous plants, Aldrovanda vesiculosa and Utricularia australis, were analysed using ultra-high-performance liquid chromatography coupled with triple quad mass spectrometry. KEY RESULTS: In both species, only isoprenoid cytokinins were identified. Zeatin cytokinins predominated in the apical parts, with their concentrations decreasing basipetally, and the trans isomer predominated in A. vesiculosa whereas the cis form was more abundant in U australis. Isopentenyladenine-type cytokinins, in contrast, increased basipetally. Conjugated cytokinin metabolites, the O-glucosides, were present at high concentrations in A. vesiculosa but only in minute amounts in U. australis. N(9)-glucoside forms were detected only in U. australis, with isopentenyladenine-9-glucoside (iP9G) being most abundant. In addition to free indole-3-acetic acid (IAA), indole-3-acetamide (IAM), IAA-aspartate (IAAsp), IAA-glutamate (IAGlu) and IAA-glycine (IAGly) conjugates were identified. CONCLUSIONS: Both species show common trends in auxin and cytokinin levels, the apical localization of the cytokinin biosynthesis and basipetal change in the ratio of active cytokinins to auxin, in favour of auxin. However, our detailed study of cytokinin metabolic profiles also revealed that both species developed different regulatory mechanisms of active cytokinin content; on the level of their degradation, in U. australis, or in the biosynthesis itself, in the case of A. vesiculosa Results indicate that the rapid turnover of these signalling molecules along the shoots is essential for maintaining the dynamic balance between the rapid polar growth and development of the apical parts and senescence of the older, basal parts of the shoots.


Asunto(s)
Citocininas/metabolismo , Ácidos Indolacéticos/metabolismo , Magnoliopsida/fisiología , Organismos Acuáticos , Carnivoría , Droseraceae/fisiología , Brotes de la Planta/metabolismo
7.
Plant Cell Rep ; 35(1): 77-89, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26403461

RESUMEN

KEY MESSAGE: Habituated embryogenic line of pumpkin contained more CKs and IAA, but less ABA than the non-habituated line. Pronounced hypomethylation correlated with the absence of 2,4-D, addition of 5-azaC, and the process of habituation. A comparative analysis between habituated and non-habituated embryogenic cultures of pumpkin (Cucurbita pepo L.) in relation to endogenous phytohormones, global DNA methylation, and developmental and regeneration capacities of the cultures was conducted. The analysis revealed more cytokinins (CKs) and indole-3-acetic acid (IAA), but less abscisic acid (ABA) in the habituated HEC line than in the non-habituated DEC line. Ribosides and ribotides were the most abundant CK forms in both HEC and DEC lines (75.9 and 57.6 %, respectively). HEC contained more free-base CKs (5.8 vs. 3.2 %), whereas DEC contained considerably more O-glycosides (39.1 vs. 18.3 %). Although prevalence of IAA was common for both lines, relative ratio of CKs and ABA differed between DEC and HEC lines. ABA was prevailing over CKs in DEC, while CKs prevailed over ABA in HEC line. Taking into account the importance of ABA for embryo maturation, the reduced endogenous ABA content in HEC line might be the reason for a 5-fold reduction in regeneration capacity compared to DEC. Both habituated and non-habituated embryogenic lines were highly methylated in the presence of 2,4-dichlorophenoxyacetic acid (2,4-D). Pronounced hypomethylation correlated with the absence of 2,4-D, addition of 5-azacytidine (5-azaC), but also with the process of habituation. The habituated line was resistant to the effect of hypomethylation drug 5-azaC and remained highly methylated even after the addition of 5-azaC. Also, 5-azaC did not change the developmental pattern in the habituated line, indicating the existence of separate mechanisms by which 2,4-D influences global DNA methylation in comparison to habituation-related global DNA methylation.


Asunto(s)
Cucurbita/genética , Epigénesis Genética/efectos de los fármacos , Ácidos Indolacéticos/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Ácido 2,4-Diclorofenoxiacético/farmacología , Ácido Abscísico/metabolismo , Azacitidina/farmacología , Cucurbita/efectos de los fármacos , Cucurbita/embriología , Citocininas/metabolismo , Metilación de ADN/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Herbicidas/farmacología , Técnicas de Embriogénesis Somática de Plantas
8.
Plant J ; 77(1): 97-107, 2014 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-24180465

RESUMEN

The unfolded protein response (UPR) is a signaling network triggered by overload of protein-folding demand in the endoplasmic reticulum (ER), a condition termed ER stress. The UPR is critical for growth and development; nonetheless, connections between the UPR and other cellular regulatory processes remain largely unknown. Here, we identify a link between the UPR and the phytohormone auxin, a master regulator of plant physiology. We show that ER stress triggers down-regulation of auxin receptors and transporters in Arabidopsis thaliana. We also demonstrate that an Arabidopsis mutant of a conserved ER stress sensor IRE1 exhibits defects in the auxin response and levels. These data not only support that the plant IRE1 is required for auxin homeostasis, they also reveal a species-specific feature of IRE1 in multicellular eukaryotes. Furthermore, by establishing that UPR activation is reduced in mutants of ER-localized auxin transporters, including PIN5, we define a long-neglected biological significance of ER-based auxin regulation. We further examine the functional relationship of IRE1 and PIN5 by showing that an ire1 pin5 triple mutant enhances defects of UPR activation and auxin homeostasis in ire1 or pin5. Our results imply that the plant UPR has evolved a hormone-dependent strategy for coordinating ER function with physiological processes.


Asunto(s)
Arabidopsis/fisiología , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/genética , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Respuesta de Proteína Desplegada , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Regulación hacia Abajo , Retículo Endoplásmico/metabolismo , Estrés del Retículo Endoplásmico , Regulación de la Expresión Génica de las Plantas , Homeostasis , Ácidos Indolacéticos/análisis , Proteínas de Transporte de Membrana/metabolismo , Modelos Biológicos , Mutación , Fenotipo , Reguladores del Crecimiento de las Plantas/análisis , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/fisiología , Proteínas Quinasas/genética , Proteínas Quinasas/metabolismo , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Plantones/genética , Plantones/fisiología , Especificidad de la Especie
9.
J Exp Bot ; 66(21): 6927-43, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26276866

RESUMEN

The shape of the maternal pericarp affects cereal grain mass and yield. Pericarp growth was analysed by magnetic resonance imaging (MRI), revealing topological maps of mobile water in developing pericarp of barley (Hordeum vulgare) and displaying tissue regions actively elongating in specific temporal-spatial patterns. Correlation analysis of MRI signals and growth rates reveals that growth in length is mediated by dorsal and also lateral rather than ventral regions. Growth in thickness is related to ventral regions. Switching from dorsal to ventral growth is associated with differential expression of axial regulators of the HD-ZipIII and Kanadi/Ettin types, and NPH3 photoreceptors, suggesting light-mediated auxin re-distribution. Auxin increases with the highest levels in the basal pericarp at 6 days after fertilization (DAF), together with transcriptionally up-regulated auxin transport and signalling. Gibberellin biosynthesis is transcriptionally up-regulated only later, and levels of bioactive gibberellins increase from 7 to 13 DAF, with higher levels in ventral than dorsal regions. Differential gene expression related to cell expansion indicates genes related to apoplast acidification, wall relaxation, sugar cleavage, water transport, and cell wall biosynthesis. Candidate genes potentially involved in pericarp extension are distinguished by their temporal expression, representing potential isoforms responsible for dorsal-mediated early growth in length or ventral-mediated late growth in thickness.


Asunto(s)
Regulación de la Expresión Génica de las Plantas , Hordeum/crecimiento & desarrollo , Reguladores del Crecimiento de las Plantas/metabolismo , Proteínas de Plantas/genética , Semillas/crecimiento & desarrollo , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Giberelinas/metabolismo , Hordeum/genética , Hordeum/metabolismo , Ácidos Indolacéticos/metabolismo , Imagen por Resonancia Magnética , Proteínas de Plantas/metabolismo , Semillas/genética , Semillas/metabolismo
10.
J Exp Bot ; 66(9): 2569-82, 2015 May.
Artículo en Inglés | MEDLINE | ID: mdl-25873679

RESUMEN

Sugar has only recently been identified as a key player in triggering bud outgrowth, while hormonal control of bud outgrowth is already well established. To get a better understanding of sugar control, the present study investigated how sugar availability modulates the hormonal network during bud outgrowth in Rosa hybrida. Other plant models, for which mutants are available, were used when necessary. Buds were grown in vitro to manipulate available sugars. The temporal patterns of the hormonal regulatory network were assessed in parallel with bud outgrowth dynamics. Sucrose determined bud entrance into sustained growth in a concentration-dependent manner. Sustained growth was accompanied by sustained auxin production in buds, and sustained auxin export in a DR5::GUS-expressing pea line. Several events occurred ahead of sucrose-stimulated bud outgrowth. Sucrose upregulated early auxin synthesis genes (RhTAR1, RhYUC1) and the auxin efflux carrier gene RhPIN1, and promoted PIN1 abundance at the plasma membrane in a pPIN1::PIN1-GFP-expressing tomato line. Sucrose downregulated both RwMAX2, involved in the strigolactone-transduction pathway, and RhBRC1, a repressor of branching, at an early stage. The presence of sucrose also increased stem cytokinin content, but sucrose-promoted bud outgrowth was not related to that pathway. In these processes, several non-metabolizable sucrose analogues induced sustained bud outgrowth in R. hybrida, Pisum sativum, and Arabidopsis thaliana, suggesting that sucrose was involved in a signalling pathway. In conclusion, we identified potential hormonal candidates for bud outgrowth control by sugar. They are central to future investigations aimed at disentangling the processes that underlie regulation of bud outgrowth by sugar.


Asunto(s)
Reguladores del Crecimiento de las Plantas/metabolismo , Rosa/crecimiento & desarrollo , Sacarosa/metabolismo , Transporte Biológico , Citocininas/metabolismo , Flores/genética , Flores/crecimiento & desarrollo , Flores/metabolismo , Regulación de la Expresión Génica de las Plantas , Ácidos Indolacéticos/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Rosa/genética , Rosa/metabolismo , Transducción de Señal
11.
Nature ; 459(7250): 1136-40, 2009 Jun 25.
Artículo en Inglés | MEDLINE | ID: mdl-19506555

RESUMEN

The plant signalling molecule auxin provides positional information in a variety of developmental processes by means of its differential distribution (gradients) within plant tissues. Thus, cellular auxin levels often determine the developmental output of auxin signalling. Conceptually, transmembrane transport and metabolic processes regulate the steady-state levels of auxin in any given cell. In particular, PIN auxin-efflux-carrier-mediated, directional transport between cells is crucial for generating auxin gradients. Here we show that Arabidopsis thaliana PIN5, an atypical member of the PIN gene family, encodes a functional auxin transporter that is required for auxin-mediated development. PIN5 does not have a direct role in cell-to-cell transport but regulates intracellular auxin homeostasis and metabolism. PIN5 localizes, unlike other characterized plasma membrane PIN proteins, to endoplasmic reticulum (ER), presumably mediating auxin flow from the cytosol to the lumen of the ER. The ER localization of other PIN5-like transporters (including the moss PIN) indicates that the diversification of PIN protein functions in mediating auxin homeostasis at the ER, and cell-to-cell auxin transport at the plasma membrane, represent an ancient event during the evolution of land plants.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/fisiología , Retículo Endoplásmico/metabolismo , Homeostasis/fisiología , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Arabidopsis/clasificación , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Células Cultivadas , Técnicas de Inactivación de Genes , Proteínas de Transporte de Membrana/genética , Mutación , Fenotipo , Filogenia , Reguladores del Crecimiento de las Plantas/metabolismo
12.
Planta ; 240(1): 55-76, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24677098

RESUMEN

Plant hormones act as chemical messengers in the regulation of myriads of physiological processes that occur in plants. To date, nine groups of plant hormones have been identified and more will probably be discovered. Furthermore, members of each group may participate in the regulation of physiological responses in planta both alone and in concert with members of either the same group or other groups. The ideal way to study biochemical processes involving these signalling molecules is 'hormone profiling', i.e. quantification of not only the hormones themselves, but also their biosynthetic precursors and metabolites in plant tissues. However, this is highly challenging since trace amounts of all of these substances are present in highly complex plant matrices. Here, we review advances, current trends and future perspectives in the analysis of all currently known plant hormones and the associated problems of extracting them from plant tissues and separating them from the numerous potentially interfering compounds.


Asunto(s)
Reguladores del Crecimiento de las Plantas , Plantas/química , Reguladores del Crecimiento de las Plantas/química , Reguladores del Crecimiento de las Plantas/aislamiento & purificación , Transducción de Señal
13.
Physiol Plant ; 148(2): 214-31, 2013 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-23043692

RESUMEN

In Pinus radiata D. Don, the transition from the juvenile to the mature phase is characterized by a reduction in the tree's organogenic potential, which is usually reverted in breeding programs by reinvigoration procedures to enable vegetative propagation. In this work, we have determined the best culture conditions for in vitro reinvigoration of radiata pine buds, tested different cytokinin (CK) types [N6-benzyladenine (BA), meta-topolin (mT) and trans-zeatin] and concentrations (25 and 50 µM), and studied the effect of culture conditions on endogenous CK and indole-3-acetic acid (IAA) levels at different stages of the organogenic process. To this end, the levels of 43 CKs and IAA were determined in P. radiata buds before and during the reinvigoration process. When BA or mT was applied to the induction medium, we did not observe any significant increase or decrease in endogenous isoprenoid CK content. We also report for the first time the presence of O-glucosides in non-treated P. radiata explants from the field and remark the importance of O-glucosides as storage forms.


Asunto(s)
Citocininas/metabolismo , Ácidos Indolacéticos/metabolismo , Pinus/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Compuestos de Bencilo , Cromatografía Liquida , Citocininas/análisis , Citocininas/farmacología , Glucósidos/análisis , Glucósidos/metabolismo , Ácidos Indolacéticos/análisis , Cinetina/farmacología , Espectrometría de Masas , Pinus/efectos de los fármacos , Pinus/crecimiento & desarrollo , Reguladores del Crecimiento de las Plantas/análisis , Reguladores del Crecimiento de las Plantas/farmacología , Brotes de la Planta/efectos de los fármacos , Brotes de la Planta/crecimiento & desarrollo , Brotes de la Planta/metabolismo , Purinas , Terpenos/análisis , Terpenos/metabolismo , Árboles , Zeatina/farmacología
14.
Proc Natl Acad Sci U S A ; 107(2): 918-22, 2010 Jan 12.
Artículo en Inglés | MEDLINE | ID: mdl-20080776

RESUMEN

The plant hormone auxin plays a crucial role in regulating plant development and plant architecture. The directional auxin distribution within tissues depends on PIN transporters that are polarly localized on the plasma membrane. The PIN polarity and the resulting auxin flow directionality are mediated by the antagonistic actions of PINOID kinase and protein phosphatase 2A. However, the contribution of the PIN phosphorylation to the polar PIN sorting is still unclear. Here, we identified an evolutionarily conserved phosphorylation site within the central hydrophilic loop of PIN proteins that is important for the apical and basal polar PIN localizations. Inactivation of the phosphorylation site in PIN1(Ala) resulted in a predominantly basal targeting and increased the auxin flow to the root tip. In contrast, the outcome of the phosphomimic PIN1(Asp) manipulation was a constitutive, PINOID-independent apical targeting of PIN1 and an increased auxin flow in the opposite direction. Furthermore, the PIN1(Asp) functionally replaced PIN2 in its endogenous expression domain, revealing that the phosphorylation-dependent polarity regulation contributes to functional diversification within the PIN family. Our data suggest that PINOID-independent PIN phosphorylation at one single site is adequate to change the PIN polarity and, consequently, to redirect auxin fluxes between cells and provide the conceptual possibility and means to manipulate auxin-dependent plant development and architecture.


Asunto(s)
Arabidopsis/genética , Sustitución de Aminoácidos , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Retículo Endoplásmico/metabolismo , Flores/genética , Flores/metabolismo , Genes Reporteros , Ácidos Indolacéticos/metabolismo , Proteínas de Transporte de Membrana/genética , Proteínas de Transporte de Membrana/metabolismo , Mutagénesis Sitio-Dirigida , Mutación , Fosforilación , Raíces de Plantas/genética , Raíces de Plantas/fisiología , Brotes de la Planta/genética , Brotes de la Planta/metabolismo
15.
Bioinformatics ; 27(10): 1404-12, 2011 May 15.
Artículo en Inglés | MEDLINE | ID: mdl-21450717

RESUMEN

MOTIVATION: Hormone pathway interactions are crucial in shaping plant development, such as synergism between the auxin and brassinosteroid pathways in cell elongation. Both hormone pathways have been characterized in detail, revealing several feedback loops. The complexity of this network, combined with a shortage of kinetic data, renders its quantitative analysis virtually impossible at present. RESULTS: As a first step towards overcoming these obstacles, we analyzed the network using a Boolean logic approach to build models of auxin and brassinosteroid signaling, and their interaction. To compare these discrete dynamic models across conditions, we transformed them into qualitative continuous systems, which predict network component states more accurately and can accommodate kinetic data as they become available. To this end, we developed an extension for the SQUAD software, allowing semi-quantitative analysis of network states. Contrasting the developmental output depending on cell type-specific modulators enabled us to identify a most parsimonious model, which explains initially paradoxical mutant phenotypes and revealed a novel physiological feature. AVAILABILITY: The package SQUADD is freely available via the Bioconductor repository at http://www.bioconductor.org/help/bioc-views/release/bioc/html/SQUADD.html.


Asunto(s)
Arabidopsis/metabolismo , Ácidos Indolacéticos/metabolismo , Modelos Biológicos , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Procesos de Crecimiento Celular
16.
J Med Chem ; 49(22): 6500-9, 2006 Nov 02.
Artículo en Inglés | MEDLINE | ID: mdl-17064068

RESUMEN

In a routine screening of our small-molecule compound collection we recently identified 4-arylazo-3,5-diamino-1H-pyrazoles as a novel group of ATP antagonists with moderate potency against CDK2-cyclin E. A preliminary SAR study based on 35 analogues suggests ways in which the pharmacophore could be further optimized, for example, via substitutions in the 4-aryl ring. Enzyme kinetics studies with the lead compound and X-ray crystallography of an inhibitor-CDK2 complex demonstrated that its mode of inhibition is competitive. Functional kinase assays confirmed the selectivity toward CDKs, with a preference for CDK9-cyclin T1. The most potent inhibitor, 4-[(3,5-diamino-1H-pyrazol-4-yl)diazenyl]phenol 31b (CAN508), reduced the frequency of S-phase cells of the cancer cell line HT-29 in antiproliferation assays. Further observed cellular effects included decreased phosphorylation of the retinoblastoma protein and the C-terminal domain of RNA polymerase II, inhibition of mRNA synthesis, and induction of the tumor suppressor protein p53, all of which are consistent with inhibition of CDK9.


Asunto(s)
Compuestos Azo/síntesis química , Compuestos Azo/farmacología , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/farmacología , Pirazoles/síntesis química , Pirazoles/farmacología , Antimetabolitos , Bromodesoxiuridina , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Cristalografía por Rayos X , Quinasa 2 Dependiente de la Ciclina/antagonistas & inhibidores , Humanos , Immunoblotting , Modelos Moleculares , ARN/biosíntesis , ARN/aislamiento & purificación , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transcripción Reversa/efectos de los fármacos , Relación Estructura-Actividad , Especificidad por Sustrato
17.
N Biotechnol ; 33(5 Pt B): 728-734, 2016 Sep 25.
Artículo en Inglés | MEDLINE | ID: mdl-26921706

RESUMEN

Flax (Linum usitatissimum L.) is an important crop for the production of oil and fiber. In vitro manipulations of flax are used for genetic improvement and breeding while improvements in adventitious root formation are important for biotechnological programs focused on regeneration and vegetative propagation of genetically valuable plant material. Additionally, flax hypocotyl segments possess outstanding morphogenetic capacity, thus providing a useful model for the investigation of flax developmental processes. Here, we investigated the crosstalk between hydrogen peroxide and auxin with respect to reprogramming flax hypocotyl cells for root morphogenetic development. Exogenous auxin induced the robust formation of adventitious roots from flax hypocotyl segments while the addition of hydrogen peroxide further enhanced this process. The levels of endogenous auxin (indole-3-acetic acid; IAA) were positively correlated with increased root formation in response to exogenous auxin (1-Naphthaleneacetic acid; NAA). Histochemical staining of the hypocotyl segments revealed that hydrogen peroxide and peroxidase, but not superoxide, were positively correlated with root formation. Measurements of antioxidant enzyme activities showed that endogenous levels of hydrogen peroxide were controlled by peroxidases during root formation from hypocotyl segments. In conclusion, hydrogen peroxide positively affected flax adventitious root formation by regulating the endogenous auxin levels. Consequently, this agent can be applied to increase flax regeneration capacity for biotechnological purposes such as improved plant rooting.


Asunto(s)
Lino/efectos de los fármacos , Lino/crecimiento & desarrollo , Peróxido de Hidrógeno/farmacología , Antioxidantes/metabolismo , Biotecnología , Reprogramación Celular/efectos de los fármacos , Lino/metabolismo , Peróxido de Hidrógeno/metabolismo , Hipocótilo/efectos de los fármacos , Hipocótilo/crecimiento & desarrollo , Hipocótilo/metabolismo , Ácidos Indolacéticos/metabolismo , Ácidos Naftalenoacéticos/farmacología , Reguladores del Crecimiento de las Plantas/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/metabolismo
18.
J Inorg Biochem ; 99(3): 776-86, 2005 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-15708799

RESUMEN

Cobalt(II) complexes with 6-(2-hydroxybenzylamino)purine (HL1), 6-(2-methoxybenzylamino)purine (HL2), 6-(3-methoxybenzylamino)purine (HL3) and 6-(4-methoxybenzylamino)purine (HL4) of the composition [Co(L1)Cl(H2O)2].H2O (1), [Co(L2)Cl(H2O)2] (2), [Co(L3)2(H2O)2].2H2O (3), [Co(L4)2(H2O)2].2H2O (4) have been synthesized. The compounds have been characterized by elemental analysis, FT-IR, ES+ MS (electrospray mass spectra in the positive ion mode) and electronic spectroscopies, magnetic and conductivity data as tetrahedral high-spin cobalt(II) complexes. The thermal stability of the complexes has also been studied. The cytotoxicity of the complexes (1-4) was determined by a Calcein acetoxymethyl (AM) assay. Human malignant melanoma (G361), human chronic myelogenous erythroleukemia (K562), human osteogenic sarcoma (HOS) and human breast adenocarcinoma (MCF7) cell lines were used for the testing. The molecular structure of 6-(3-methoxybenzylamino)purinium chloride monohydrate, H2L3+.Cl.H2O, i.e. a protonated form of the free HL(3) ligand, has been determined by a single crystal X-ray analysis. The geometry optimisation and infrared frequencies calculations of HL1, HL2, and H2L3+ and H2L4+ were performed using density-functional theory (DFT) calculations at the B3LYP/6-31G* level of the theory. The geometry of complex (1) was optimised at the same level of the theory.


Asunto(s)
Antineoplásicos/síntesis química , Cobalto/química , Citocininas/química , Compuestos Organometálicos/síntesis química , Purinas/química , Antineoplásicos/farmacología , Cobalto/farmacología , Cristalografía por Rayos X/métodos , Espectroscopía de Resonancia por Spin del Electrón/métodos , Fluoresceínas , Humanos , Espectroscopía de Resonancia Magnética/métodos , Modelos Moleculares , Compuestos Organometálicos/farmacología , Purinas/farmacología , Espectrometría de Masa por Ionización de Electrospray/métodos , Temperatura , Células Tumorales Cultivadas
19.
PLoS One ; 10(2): e0117793, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-25695830

RESUMEN

Dark-induced growth (skotomorphogenesis) is primarily characterized by rapid elongation of the hypocotyl. We have studied the role of abscisic acid (ABA) during the development of young tomato (Solanum lycopersicum L.) seedlings. We observed that ABA deficiency caused a reduction in hypocotyl growth at the level of cell elongation and that the growth in ABA-deficient plants could be improved by treatment with exogenous ABA, through which the plants show a concentration dependent response. In addition, ABA accumulated in dark-grown tomato seedlings that grew rapidly, whereas seedlings grown under blue light exhibited low growth rates and accumulated less ABA. We demonstrated that ABA promotes DNA endoreduplication by enhancing the expression of the genes encoding inhibitors of cyclin-dependent kinases SlKRP1 and SlKRP3 and by reducing cytokinin levels. These data were supported by the expression analysis of the genes which encode enzymes involved in ABA and CK metabolism. Our results show that ABA is essential for the process of hypocotyl elongation and that appropriate control of the endogenous level of ABA is required in order to drive the growth of etiolated seedlings.


Asunto(s)
Ácido Abscísico/metabolismo , Oscuridad , Endorreduplicación/efectos de la radiación , Hipocótilo/crecimiento & desarrollo , Hipocótilo/efectos de la radiación , Solanum lycopersicum/crecimiento & desarrollo , Solanum lycopersicum/efectos de la radiación , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Citocininas/biosíntesis , Citocininas/metabolismo , Germinación/efectos de los fármacos , Germinación/efectos de la radiación , Homeostasis/efectos de la radiación , Hipocótilo/citología , Hipocótilo/metabolismo , Solanum lycopersicum/citología , Solanum lycopersicum/metabolismo , Desarrollo de la Planta/efectos de los fármacos , Desarrollo de la Planta/efectos de la radiación , Inhibidores de Proteínas Quinasas/farmacología
20.
Artículo en Inglés | MEDLINE | ID: mdl-12101055

RESUMEN

A single-step, highly specific and easy-to-use method was developed for isolation and purification of melatonin from complex biological matrices. Polyclonal antibodies highly specific against melatonin (with cross-reactivities with related compounds below 0.02%, except for 6-hydroxymelatonin) were raised, characterised by enzyme-linked immunosorbent assay (ELISA) and used for preparation of immunoaffinity gel. Melatonin recovery by the immunoaffinity method was approximately 95%, allowing single-step processing of samples prior to electrospray HPLC-MS analysis (with detection limit 10 fmol). The method was successfully used for determining melatonin in human serum and turned out to be better than the non-specific solid-phase extraction published earlier.


Asunto(s)
Cromatografía de Afinidad/métodos , Cromatografía Líquida de Alta Presión/métodos , Melatonina/aislamiento & purificación , Reacciones Cruzadas , Ensayo de Inmunoadsorción Enzimática , Sensibilidad y Especificidad , Espectrometría de Masa por Ionización de Electrospray
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