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1.
Plant Physiol ; 192(1): 34-55, 2023 05 02.
Artículo en Inglés | MEDLINE | ID: mdl-36789623

RESUMEN

While various labs had shown cell division-inducing activity in a variety of plant extracts for over a decade, the identification of zeatin (Z) in 1964, the first known naturally occurring cytokinin, belongs to Letham and co-workers. Using extracts from maize (Zea mays), they were the first to obtain crystals of pure Z and in sufficient quantity for structural determination by MS, NMR, chromatography, and mixed melting-point analysis. This group also crystallized Z-9-riboside (ZR) from coconut (Cocos nucifera) milk. However, their chemical contributions go well beyond the identification of Z and ZR and include two unambiguous syntheses of trans-Z (to establish stereochemistry), the synthesis of 3H-cytokinins that facilitated metabolic studies, and the synthesis of deuterated internal standards for accurate mass spectral quantification. Letham and associates also unequivocally identified Z nucleotide, the 7-and 9-glucoside conjugates of Z, and the O-glucosides of Z, ZR, dihydro Z (DHZ) and DHZR as endogenous compounds and as metabolites of exogenous Z. Their contributions to the role of cytokinins in plant physiology and development were also substantial, especially the role of cytokinins moving in the xylem. These biological advances are described and briefly related to the genetic/molecular biological contributions of others that established that plants have an absolute requirement for cytokinin.


Asunto(s)
Aniversarios y Eventos Especiales , Zeatina , Humanos , Zeatina/química , Zeatina/metabolismo , Zeatina/farmacología , Citocininas/metabolismo
2.
Int J Mol Sci ; 24(6)2023 Mar 09.
Artículo en Inglés | MEDLINE | ID: mdl-36982301

RESUMEN

Psb28 is a soluble protein in the photosystem II (PSII) complex, but its role in the drought stress response of wheat remains unclear. Here, we functionally characterized the TaPsb28 gene, which positively regulates drought tolerance in wheat. When the full-length 546-bp TaPsb28 cDNA was transferred into Arabidopsis thaliana, it was located in the guard cell chloroplast around the stroma. Overexpression of TaPsb28 conferred drought tolerance, as exhibited by the increases in the survival rate. Transgenic plants maintained lower MDA content and higher chlorophyll content by inducing chlorophyll synthase (ChlG) gene transcription. The content of abscisic acid (ABA) and zeatin increased significantly in wild-type (WT) plants under drought stress, and the transcriptional expression levels of RD22, dihydroflavonol 4-reductase (DFR) and anthocyanin reductase (ANR) genes were induced, thus enhancing the contents of endogenous cyanidin, delphinidin, and proanthocyanidins. However, in transgenic plants, although anthocyanins were further aggregated, the ABA increase was inhibited, zeatin was restored to the control level under drought stress, and stomatal closure was promoted. These findings indicate ABA and zeatin have opposite synergistic effects in the process of drought tolerance caused by TaPsb28 because only after the effect of zeatin is alleviated can ABA better play its role in promoting anthocyanin accumulation and stomatal closure, thus enhancing the drought tolerance of transgenic plants. The results suggest that overexpression of TaPsb28 exerts a positive role in the drought response by influencing the functional metabolism of endogenous hormones. The understanding acquired through the research laid a foundation for further in-depth investigation of the function of TaPsb28 in drought resistance in wheat, especially its relationship with anthocyanidin accumulation.


Asunto(s)
Arabidopsis , Arabidopsis/fisiología , Antocianinas/farmacología , Resistencia a la Sequía , Triticum/fisiología , Zeatina/farmacología , Estrés Fisiológico , Plantas Modificadas Genéticamente/metabolismo , Ácido Abscísico/metabolismo , Sequías , Oxidorreductasas/metabolismo , Regulación de la Expresión Génica de las Plantas , Proteínas de Plantas/metabolismo
3.
Appl Environ Microbiol ; 88(6): e0216021, 2022 03 22.
Artículo en Inglés | MEDLINE | ID: mdl-35108066

RESUMEN

Phloroglucinol improves shoot formation and somatic embryogenesis in several horticultural and grain crops, but its function in microalgae remains unclear. Here, we found that sufficiently high concentrations of phloroglucinol significantly increased fucoxanthin synthesis, growth, and photosynthetic efficiency in the microalga Thalassiosira pseudonana. These results suggested that the role of phloroglucinol is conserved across higher plants and microalgae. Further analysis showed that, after phloroglucinol treatment, the contents of cis-zeatin and brassinolide in T. pseudonana increased significantly, while the contents of trans-zeatin, N6-isopentenyladenine (iP), auxin, and gibberellin were unaffected. Indeed, functional studies showed that the effects of cis-zeatin and brassinolide in T. pseudonana were similar to those of phloroglucinol. Knockout of key enzyme genes in the cis-zeatin synthesis pathway of T. pseudonana or treatment of T. pseudonana with a brassinolide synthesis inhibitor (brassinazole) significantly reduced growth and fucoxanthin content in T. pseudonana, and phloroglucinol treatment partially alleviated these inhibitory effects. However, phloroglucinol treatment was ineffective when the cis-zeatin and brassinolide pathways were simultaneously inhibited. These results suggested that the cis-zeatin and brassinolide signaling pathways are independent regulators of fucoxanthin synthesis in T. pseudonana and that phloroglucinol affects both pathways. Thus, this study not only characterizes the mechanism by which phloroglucinol promotes fucoxanthin synthesis but also demonstrates the roles of cis-zeatin and brassinolide in T. pseudonana. IMPORTANCE Here, we demonstrate that phloroglucinol, a growth promoter in higher plants, also increases growth and fucoxanthin synthesis in the microalga Thalassiosira pseudonana and therefore may have substantial practical application for industrial fucoxanthin production. Phloroglucinol treatment also induced the synthesis of cis-zeatin and brassinolide in T. pseudonana, and the cis-zeatin and brassinolide signaling pathways were implicated in the phloroglucinol-driven increases in T. pseudonana growth and fucoxanthin synthesis. Thus, our work clarified the molecular mechanism of phloroglucinol promoting the growth and fucoxanthin synthesis of Thalassiosira pseudonana and suggested that cis-zeatin and brassinolide, in addition to phloroglucinol, have potential utility as inducers of increased microalgal fucoxanthin production.


Asunto(s)
Diatomeas , Zeatina , Brasinoesteroides , Floroglucinol/metabolismo , Esteroides Heterocíclicos , Xantófilas , Zeatina/metabolismo , Zeatina/farmacología
4.
New Phytol ; 224(1): 242-257, 2019 10.
Artículo en Inglés | MEDLINE | ID: mdl-31230346

RESUMEN

Phosphate (Pi) is an essential nutrient for all organisms. Roots are underground organs, but the majority of the root biology studies have been done on root systems growing in the presence of light. Root illumination alters the Pi starvation response (PSR) at different intensities. Thus, we have analyzed morphological, transcriptional and physiological responses to Pi starvation in dark-grown roots. We have identified new genes and pathways regulated by Pi starvation that were not described previously. We also show that Pi-starved plants increase the cis-zeatin (cZ) : trans-zeatin (tZ) ratio. Transcriptomic analyses show that tZ preferentially represses cell cycle and PSR genes, whereas cZ induces genes involved in cell and root hair elongation and differentiation. In fact, cZ-treated seedlings show longer root system as well as longer root hairs compared with tZ-treated seedlings, increasing the total absorbing surface. Mutants with low cZ concentrations do not allocate free Pi in roots during Pi starvation. We propose that Pi-starved plants increase the cZ : tZ ratio to maintain basal cytokinin responses and allocate Pi in the root system to sustain its growth. Therefore, cZ acts as a PSR hormone that stimulates root and root hair elongation to enlarge the root absorbing surface and to increase Pi concentrations in roots.


Asunto(s)
Fosfatos/deficiencia , Raíces de Plantas/metabolismo , Zeatina/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/metabolismo , Arabidopsis/efectos de la radiación , Proliferación Celular/efectos de los fármacos , Proliferación Celular/efectos de la radiación , Regulación de la Expresión Génica de las Plantas/efectos de los fármacos , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Luz , Reguladores del Crecimiento de las Plantas/farmacología , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/efectos de la radiación , Brotes de la Planta/efectos de los fármacos , Brotes de la Planta/metabolismo , Brotes de la Planta/efectos de la radiación , Zeatina/farmacología
5.
J Theor Biol ; 404: 182-205, 2016 09 07.
Artículo en Inglés | MEDLINE | ID: mdl-27157127

RESUMEN

The growth of the root of Arabidopsis thaliana is sustained by the meristem, a region of cell proliferation and differentiation which is located in the root apex and generates cells which move shootwards, expanding rapidly to cause root growth. The balance between cell division and differentiation is maintained via a signalling network, primarily coordinated by the hormones auxin, cytokinin and gibberellin. Since these hormones interact at different levels of spatial organisation, we develop a multi-scale computational model which enables us to study the interplay between these signalling networks and cell-cell communication during the specification of the root meristem. We investigate the responses of our model to hormonal perturbations, validating the results of our simulations against experimental data. Our simulations suggest that one or more additional components are needed to explain the observed expression patterns of a regulator of cytokinin signalling, ARR1, in roots not producing gibberellin. By searching for novel network components, we identify two mutant lines that affect significantly both root length and meristem size, one of which also differentially expresses a central component of the interaction network (SHY2). More generally, our study demonstrates how a multi-scale investigation can provide valuable insight into the spatio-temporal dynamics of signalling networks in biological tissues.


Asunto(s)
Arabidopsis/metabolismo , Meristema/citología , Meristema/metabolismo , Modelos Biológicos , Reguladores del Crecimiento de las Plantas/metabolismo , Transducción de Señal , Arabidopsis/efectos de los fármacos , Proteínas de Arabidopsis/metabolismo , Transporte Biológico/efectos de los fármacos , Giberelinas/metabolismo , Ácidos Indolacéticos/metabolismo , Meristema/efectos de los fármacos , Tamaño de los Órganos/efectos de los fármacos , Reproducibilidad de los Resultados , Transducción de Señal/efectos de los fármacos , Triazoles/farmacología , Zeatina/farmacología
6.
Proc Natl Acad Sci U S A ; 110(4): 1548-53, 2013 Jan 22.
Artículo en Inglés | MEDLINE | ID: mdl-23319631

RESUMEN

Maintaining nitric oxide (NO) homeostasis is essential for normal plant physiological processes. However, very little is known about the mechanisms of NO modulation in plants. Here, we report a unique mechanism for the catabolism of NO based on the reaction with the plant hormone cytokinin. We screened for NO-insensitive mutants in Arabidopsis and isolated two allelic lines, cnu1-1 and 1-2 (continuous NO-unstressed 1), that were identified as the previously reported altered meristem program 1 (amp1) and as having elevated levels of cytokinins. A double mutant of cnu1-2 and nitric oxide overexpression 1 (nox1) reduced the severity of the phenotypes ascribed to excess NO levels as did treating the nox1 line with trans-zeatin, the predominant form of cytokinin in Arabidopsis. We further showed that peroxinitrite, an active NO derivative, can react with zeatin in vitro, which together with the results in vivo suggests that cytokinins suppress the action of NO most likely through direct interaction between them, leading to the reduction of endogenous NO levels. These results provide insights into NO signaling and regulation of its bioactivity in plants.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Citocininas/metabolismo , Óxido Nítrico/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Arabidopsis/crecimiento & desarrollo , Proteínas de Arabidopsis/genética , Carboxipeptidasas/genética , Carboxipeptidasas/metabolismo , Citocininas/química , Citocininas/genética , Flores/crecimiento & desarrollo , Flores/metabolismo , Genes de Plantas , Mutación , Donantes de Óxido Nítrico/farmacología , Nitroprusiato/farmacología , Ácido Peroxinitroso/metabolismo , Fenotipo , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN de Planta/genética , ARN de Planta/metabolismo , Zeatina/metabolismo , Zeatina/farmacología
7.
Proc Natl Acad Sci U S A ; 110(24): 10028-33, 2013 Jun 11.
Artículo en Inglés | MEDLINE | ID: mdl-23720308

RESUMEN

Cytokinins are plant hormones that play critical roles in growth and development. In Arabidopsis, the transcriptional response to cytokinin is regulated by action of type-B Arabidopsis response regulators (ARRs). Although central elements in the cytokinin signal transduction pathway have been identified, mechanisms controlling output remain to be elucidated. Here we demonstrate that a family of F-box proteins, called the kiss me deadly (KMD) family, targets type-B ARR proteins for degradation. KMD proteins form an S-phase kinase-associated PROTEIN1 (SKP1)/Cullin/F-box protein (SCF) E3 ubiquitin ligase complex and directly interact with type-B ARR proteins. Loss-of-function KMD mutants stabilize type-B ARRs and exhibit an enhanced cytokinin response. In contrast, plants with elevated KMD expression destabilize type-B ARR proteins leading to cytokinin insensitivity. Our results support a model in which an SCF(KMD) complex negatively regulates cytokinin responses by controlling levels of a key family of transcription factors.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Citocininas/farmacología , Proteínas de Unión al ADN/metabolismo , Proteínas F-Box/metabolismo , Factores de Transcripción/metabolismo , Secuencia de Aminoácidos , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Unión al ADN/genética , Proteínas F-Box/clasificación , Proteínas F-Box/genética , Immunoblotting , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Microscopía Fluorescente , Datos de Secuencia Molecular , Familia de Multigenes , Mutación , Filogenia , Reguladores del Crecimiento de las Plantas/farmacología , Plantas Modificadas Genéticamente , Unión Proteica , Protoplastos/metabolismo , Homología de Secuencia de Aminoácido , Transducción de Señal , Factores de Transcripción/genética , Zeatina/farmacología
8.
Curr Genet ; 61(4): 653-63, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-25986972

RESUMEN

An efficient system for shoot regeneration and Agrobacterium tumefaciens-mediated transformation of Brassica oleracea cv. Green Marvel cultivar is described. This study focuses on developing shoot regeneration from hypocotyl explants of broccoli cv. Green Marvel using thidiazuron (TDZ), zeatin, and kinetin, the optimization of factors affecting Agrobacterium-mediated transformation of the hypocotyl explants with heat-resistant cDNA, followed by the confirmation of transgenicity of the regenerants. High shoot regeneration was observed in 0.05-0.1 mg dm(-3) TDZ. TDZ at 0.1 mg dm(-3) produced among the highest percentage of shoot regeneration (96.67 %) and mean number of shoot formation (6.17). The highest percentage (13.33 %) and mean number (0.17) of putative transformant production were on hypocotyl explants subjected to preculture on shoot regeneration medium (SRM) with 200 µM acetosyringone. On optimization of bacterial density and inoculation time, the highest percentage and mean number of putative transformant production were on hypocotyl explants inoculated with a bacterial dilution of 1:5 for 30 min. Polymerase chain reaction (PCR) assay indicated a transformation efficiency of 8.33 %. The luciferase assay showed stable integration of the Arabidopsis thaliana HSP101 (AtHSP101) cDNA in the transgenic broccoli regenerants. Three out of five transgenic lines confirmed through PCR showed positive hybridization bands of the AtHSP101 cDNA through Southern blot analysis. The presence of AtHSP101 transcripts in the three transgenic broccoli lines indicated by reverse transcription-PCR (RT-PCR) confirmed the expression of the gene. In conclusion, an improved regeneration system has been established from hypocotyl explants of broccoli followed by successful transformation with AtHSP101 for resistance to high temperature.


Asunto(s)
Agrobacterium tumefaciens/genética , Brassica/genética , ADN Complementario/genética , Hipocótilo/genética , Proteínas de Plantas/genética , Factores de Transcripción/genética , Transformación Genética , Acetofenonas/farmacología , Adaptación Fisiológica/genética , Agrobacterium tumefaciens/metabolismo , Arabidopsis/genética , Brassica/efectos de los fármacos , Brassica/metabolismo , Brassica/microbiología , ADN Complementario/metabolismo , Expresión Génica , Vectores Genéticos , Calor , Hipocótilo/efectos de los fármacos , Hipocótilo/metabolismo , Hipocótilo/microbiología , Cinetina/farmacología , Compuestos de Fenilurea/farmacología , Reguladores del Crecimiento de las Plantas/farmacología , Proteínas de Plantas/metabolismo , Plantas Modificadas Genéticamente , ARN Mensajero/genética , ARN Mensajero/metabolismo , Regeneración/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tiadiazoles/farmacología , Factores de Transcripción/metabolismo , Transgenes , Zeatina/farmacología
9.
Cryo Letters ; 35(2): 119-28, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-24869644

RESUMEN

BACKGROUND: Thymus lotocephalus is a rare endemic species from the Algarve, Portugal, and is legally protected by Portuguese and European legislation. OBJECTIVE: The aim is to develop a cryopreservation protocol for T. lotocephalus shoot tips, as an alternative approach for the long-term conservation of this species. METHODS: Several methods (droplet-vitrification, vitrification and encapsulation-dehydration) were tested. Conditions regarding the subculture period, cold-hardening and preculture were optimized. Cryopreserved shoot tips were also assessed for their genetic stability using RAPD markers. RESULTS: Droplet-vitrification presented the best results. The best regrowth of cryopreserved shoot tips obtained eight weeks after rewarming was 67%. This was accomplished with four weeks subculture period of in vitro-donor plants at 25 degree C, preculture of excised shoot tips for one day on MS medium containing 0.3 M sucrose, treatment in PVS2 for 60 min, and MS supplemented with 0.2 mg per L zeatin as recovery medium. The assessment using RAPD markers observed variation at a low frequency and shoots regenerated from cryopreserved apices showed normal development compared to the regular in vitro-grown shoots. CONCLUSION: Droplet-vitrification is thus a viable method for the cryopreservation of T. lotocephalus shoot tips.


Asunto(s)
Criopreservación , ADN de Plantas/genética , Genoma de Planta , Brotes de la Planta/fisiología , Thymus (Planta)/fisiología , Vitrificación , Adaptación Fisiológica , Conservación de los Recursos Naturales , Crioprotectores/farmacología , Medios de Cultivo , Desecación , Especies en Peligro de Extinción , Inestabilidad Genómica , Reguladores del Crecimiento de las Plantas/farmacología , Brotes de la Planta/efectos de los fármacos , Portugal , Técnica del ADN Polimorfo Amplificado Aleatorio , Sacarosa/farmacología , Thymus (Planta)/efectos de los fármacos , Zeatina/farmacología
10.
Nat Commun ; 15(1): 8489, 2024 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-39353942

RESUMEN

Plants growing at high densities can detect competitors through changes in the composition of light reflected by neighbours. In response to this far-red-enriched light, plants elicit adaptive shade avoidance responses for light capture, but these need to be balanced against other input signals, such as nutrient availability. Here, we investigated how Arabidopsis integrates shade and nitrate signalling. We unveiled that nitrate modulates shade avoidance via a previously unknown shade response pathway that involves root-derived trans-zeatin (tZ) signal and the BEE1 transcription factor as an integrator of light and cytokinin signalling. Under nitrate-sufficient conditions, tZ promotes hypocotyl elongation specifically in the presence of supplemental far-red light. This occurs via PIF transcription factors-dependent inhibition of type-A ARRs cytokinin response inhibitors. Our data thus reveal how plants co-regulate responses to shade cues with root-derived information about nutrient availability, and how they restrict responses to this information to specific light conditions in the shoot.


Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Citocininas , Regulación de la Expresión Génica de las Plantas , Luz , Nitratos , Fitocromo , Raíces de Plantas , Transducción de Señal , Arabidopsis/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Raíces de Plantas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/efectos de los fármacos , Nitratos/metabolismo , Citocininas/metabolismo , Citocininas/farmacología , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Fitocromo/metabolismo , Hipocótilo/crecimiento & desarrollo , Hipocótilo/metabolismo , Zeatina/metabolismo , Zeatina/farmacología , Factores de Transcripción/metabolismo , Factores de Transcripción/genética
11.
Cells ; 13(8)2024 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-38667301

RESUMEN

Phytohormones, particularly cytokinin trans-zeatin (tZ), were studied for their impact on the green alga Desmodesmus armatus under cadmium (Cd) stress, focusing on growth, metal accumulation, and stress response mechanisms. Using atomic absorption spectroscopy for the Cd level and high-performance liquid chromatography for photosynthetic pigments and phytochelatins, along with spectrophotometry for antioxidants and liquid chromatography-mass spectrometry for phytohormones, we found that tZ enhances Cd uptake in D. armatus, potentially improving phycoremediation of aquatic environments. Cytokinin mitigates Cd toxicity by regulating internal phytohormone levels and activating metal tolerance pathways, increasing phytochelatin synthase activity and phytochelatin accumulation essential for Cd sequestration. Treatment with tZ and Cd also resulted in increased cell proliferation, photosynthetic pigment and antioxidant levels, and antioxidant enzyme activities, reducing oxidative stress. This suggests that cytokinin-mediated mechanisms in D. armatus enhance its capacity for Cd uptake and tolerance, offering promising avenues for more effective aquatic phycoremediation techniques.


Asunto(s)
Antioxidantes , Cadmio , Chlorophyta , Zeatina , Cadmio/toxicidad , Zeatina/metabolismo , Zeatina/farmacología , Antioxidantes/metabolismo , Antioxidantes/farmacología , Chlorophyta/efectos de los fármacos , Chlorophyta/metabolismo , Estrés Oxidativo/efectos de los fármacos , Fotosíntesis/efectos de los fármacos , Fitoquelatinas/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Reguladores del Crecimiento de las Plantas/metabolismo
12.
Biomolecules ; 14(5)2024 Apr 25.
Artículo en Inglés | MEDLINE | ID: mdl-38785924

RESUMEN

Cytokinins (CKs) and abscisic acid (ABA) play an important role in the life of both plants and pathogenic fungi. However, the role of CKs and ABA in the regulation of fungal growth, development and virulence has not been sufficiently studied. We compared the ability of two virulent isolates (SnB and Sn9MN-3A) and one avirulent isolate (Sn4VD) of the pathogenic fungus Stagonospora nodorum Berk. to synthesize three groups of hormones (CKs, ABA and auxins) and studied the effect of exogenous ABA and zeatin on the growth, sporulation and gene expression of necrotrophic effectors (NEs) and transcription factors (TFs) in them. Various isolates of S. nodorum synthesized different amounts of CKs, ABA and indoleacetic acid. Using exogenous ABA and zeatin, we proved that the effect of these hormones on the growth and sporulation of S. nodorum isolates can be opposite, depends on both the genotype of the isolate and on the concentration of the hormone and is carried out through the regulation of carbohydrate metabolism. ABA and zeatin regulated the expression of fungal TF and NE genes, but correlation analysis of these parameters showed that this effect depended on the genotype of the isolate. This study will contribute to our understanding of the role of the hormones ABA and CKs in the biology of the fungal pathogen S. nodorum.


Asunto(s)
Ácido Abscísico , Ascomicetos , Citocininas , Ácido Abscísico/metabolismo , Citocininas/metabolismo , Ascomicetos/metabolismo , Ascomicetos/patogenicidad , Ascomicetos/genética , Ascomicetos/efectos de los fármacos , Virulencia , Regulación Fúngica de la Expresión Génica/efectos de los fármacos , Enfermedades de las Plantas/microbiología , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Zeatina/metabolismo , Zeatina/farmacología , Esporas Fúngicas/crecimiento & desarrollo , Esporas Fúngicas/metabolismo , Esporas Fúngicas/efectos de los fármacos , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética
13.
BMC Plant Biol ; 13: 115, 2013 Aug 12.
Artículo en Inglés | MEDLINE | ID: mdl-23937221

RESUMEN

BACKGROUND: Lepidium campestre is an undomesticated oilseed species with a great potential to become a new crop for both food and industrial feedstocks production. Genetic modification is needed for further improving the oil quantity and quality of Lepidium. Studies on in vitro shoot regeneration of Lepidium are very limited and there is no transformation protocol available. RESULTS: We have investigated the effects of different factors, especially the type, concentration and combination of plant growth regulators (PGRs) on in vitro shoot regeneration of Lepidium. The results showed that the 2,4-D treatment was crucial to shoot regeneration from different explants. The duration of 2,4-D exposure between 2-4 days did not show significant difference in shoot regeneration, while the effect of 2,4-D concentration varied greatly depending on the type of explants and cytokinins used, for example, the low concentration of 2,4-D combined with TDZ significantly increased the regeneration frequency of hypocotyls. Cotyledon and hypocotyl explants responded differently to cytokinin, for example, TDZ was more effective than zeatin in promoting shoot regeneration from hypocotyls, but did not affect the regeneration of cotyledons which was more affected by high concentration of zeatin. The results also showed that NAA was not effective for shoot regeneration. Germination in light increased the regeneration frequency compared to that in dark. After optimization of the different conditions, an efficient regeneration protocol was developed with the regeneration efficiency of 92.7%. Using this protocol, the transformation frequency of 6% in average was achieved. The presence of transgenes in the transgenic lines was confirmed by GUS staining, PCR and Southern blot analyses. CONCLUSION: Through systematic investigation of important factors affecting in vitro shoot regeneration, we have developed an efficient regeneration and transformation protocol for the genetic modification of Lepidium campestre. The method may also be applied to the related species.


Asunto(s)
Lepidium/crecimiento & desarrollo , Reguladores del Crecimiento de las Plantas/farmacología , Regeneración , Transformación Genética , Ácido 2,4-Diclorofenoxiacético , Citocininas/farmacología , Ingeniería Genética/métodos , Lepidium/genética , Compuestos de Fenilurea/farmacología , Brotes de la Planta/crecimiento & desarrollo , Plantas Modificadas Genéticamente/crecimiento & desarrollo , Tiadiazoles/farmacología , Zeatina/farmacología
14.
Plant Physiol ; 160(1): 319-31, 2012 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-22811434

RESUMEN

cis-Zeatin (cZ) is generally regarded as a cytokinin with little or no activity, compared with the highly active trans-zeatin (tZ). Although recent studies suggested possible roles for cZ, its physiological significance remains unclear. In our studies with rice (Oryza sativa), cZ inhibited seminal root elongation and up-regulated cytokinin-inducible genes, and its activities were comparable to those of tZ. Tracer experiments showed that exogenously supplied cZ-riboside was mainly converted into cZ derivatives but scarcely into tZ derivatives, indicating that isomerizations of cZ derivatives into tZ derivatives are a minor pathway in rice cytokinin metabolism. We identified three putative cZ-O-glucosyltransferases (cZOGT1, cZOGT2, and cZOGT3) in rice. The cZOGTs preferentially catalyzed O-glucosylation of cZ and cZ-riboside rather than tZ and tZ-riboside in vitro. Transgenic rice lines ectopically overexpressing the cZOGT1 and cZOGT2 genes exhibited short-shoot phenotypes, delay of leaf senescence, and decrease in crown root number, while cZOGT3 overexpressor lines did not show shortened shoots. These results propose that cZ activity has a physiological impact on the growth and development of rice.


Asunto(s)
Citocininas/metabolismo , Glucosiltransferasas/metabolismo , Oryza/enzimología , Proteínas de Plantas/metabolismo , Zeatina/farmacología , Secuencia de Aminoácidos , Clonación Molecular , Activación Enzimática , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Glucosiltransferasas/genética , Isomerismo , Datos de Secuencia Molecular , Oryza/efectos de los fármacos , Oryza/genética , Oryza/crecimiento & desarrollo , Fenotipo , Proteínas de Plantas/genética , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/enzimología , Raíces de Plantas/crecimiento & desarrollo , Brotes de la Planta/efectos de los fármacos , Brotes de la Planta/crecimiento & desarrollo , Plantas Modificadas Genéticamente/efectos de los fármacos , Plantas Modificadas Genéticamente/enzimología , Plantas Modificadas Genéticamente/genética , Alineación de Secuencia , Zeatina/metabolismo
15.
Ann Bot ; 112(7): 1395-407, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-24061489

RESUMEN

BACKGROUND AND AIMS: Adventitious roots (ARs) are part of the root system in numerous plants, and are required for successful micropropagation. In the Arabidopsis thaliana primary root (PR) and lateral roots (LRs), the quiescent centre (QC) in the stem cell niche of the meristem controls apical growth with the involvement of auxin and cytokinin. In arabidopsis, ARs emerge in planta from the hypocotyl pericycle, and from different tissues in in vitro cultured explants, e.g. from the stem endodermis in thin cell layer (TCL) explants. The aim of this study was to investigate the establishment and maintenance of the QC in arabidopsis ARs, in planta and in TCL explants, because information about this process is still lacking, and it has potential use for biotechnological applications. METHODS: Expression of PR/LR QC markers and auxin influx (LAX3)/efflux (PIN1) genes was investigated in the presence/absence of exogenous auxin and cytokinin. Auxin was monitored by the DR5::GUS system and cytokinin by immunolocalization. The expression of the auxin-biosynthetic YUCCA6 gene was also investigated by in situ hybridization in planta and in AR-forming TCLs from the indole acetic acid (IAA)-overproducing superroot2-1 mutant and its wild type. KEY RESULTS: The accumulation of auxin and the expression of the QC marker WOX5 characterized the early derivatives of the AR founder cells, in planta and in in vitro cultured TCLs. By determination of PIN1 auxin efflux carrier and LAX3 auxin influx carrier activities, an auxin maximum was determined to occur at the AR tip, to which WOX5 expression was restricted, establishing the positioning of the QC. Cytokinin caused a restriction of LAX3 and PIN1 expression domains, and concomitantly the auxin biosynthesis YUCCA6 gene was expressed in the apex. CONCLUSIONS: In ARs formed in planta and TCLs, the QC is established in a similar way, and auxin transport and biosynthesis are involved through cytokinin tuning.


Asunto(s)
Arabidopsis/citología , Arabidopsis/metabolismo , Citocininas/metabolismo , Ácidos Indolacéticos/metabolismo , Raíces de Plantas/citología , Raíces de Plantas/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico/efectos de los fármacos , Transporte Biológico/genética , Biomarcadores/metabolismo , División Celular/efectos de los fármacos , División Celular/genética , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Glucuronidasa/metabolismo , Proteínas de Homeodominio/metabolismo , Hipocótilo/efectos de los fármacos , Hipocótilo/crecimiento & desarrollo , Indoles/farmacología , Isopenteniladenosina/análogos & derivados , Isopenteniladenosina/metabolismo , Cinetina/farmacología , Proteínas de Transporte de Membrana/metabolismo , Oxigenasas de Función Mixta/genética , Oxigenasas de Función Mixta/metabolismo , Modelos Biológicos , Raíces de Plantas/efectos de los fármacos , Plantones/efectos de los fármacos , Plantones/crecimiento & desarrollo , Zeatina/farmacología
16.
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
17.
Phytopathology ; 103(9): 888-99, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23593941

RESUMEN

The soil phytopathogen Agrobacterium tumefaciens causes crown gall disease in a wide range of plant species. The neoplastic growth at the infection sites is caused by transferring, integrating, and expressing transfer DNA (T-DNA) from A. tumefaciens into plant cells. A trans-zeatin synthesizing (tzs) gene is located in the nopaline-type tumor-inducing plasmid and causes trans-zeatin production in A. tumefaciens. Similar to known virulence (Vir) proteins that are induced by the vir gene inducer acetosyringone (AS) at acidic pH 5.5, Tzs protein is highly induced by AS under this growth condition but also constitutively expressed and moderately upregulated by AS at neutral pH 7.0. We found that the promoter activities and protein levels of several AS-induced vir genes increased in the tzs deletion mutant, a mutant with decreased tumorigenesis and transient transformation efficiencies, in Arabidopsis roots. During AS induction and infection of Arabidopsis roots, the tzs deletion mutant conferred impaired growth, which could be rescued by genetic complementation and supplementing exogenous cytokinin. Exogenous cytokinin also repressed vir promoter activities and Vir protein accumulation in both the wild-type and tzs mutant bacteria with AS induction. Thus, the tzs gene or its product, cytokinin, may be involved in regulating AS-induced vir gene expression and, therefore, affect bacterial growth and virulence during A. tumefaciens infection.


Asunto(s)
Agrobacterium tumefaciens/fisiología , Arabidopsis/microbiología , Proteínas Bacterianas/genética , Regulación Bacteriana de la Expresión Génica , Reguladores del Crecimiento de las Plantas/farmacología , Tumores de Planta/microbiología , Acetofenonas/farmacología , Agrobacterium tumefaciens/genética , Agrobacterium tumefaciens/crecimiento & desarrollo , Agrobacterium tumefaciens/patogenicidad , Proteínas Bacterianas/metabolismo , Citocininas/metabolismo , Citocininas/farmacología , Expresión Génica , Genes Reporteros , Prueba de Complementación Genética , Concentración de Iones de Hidrógeno , Reguladores del Crecimiento de las Plantas/metabolismo , Raíces de Plantas/microbiología , Plásmidos/genética , Eliminación de Secuencia , Regulación hacia Arriba , Virulencia , Factores de Virulencia/genética , Factores de Virulencia/metabolismo , Zeatina/metabolismo , Zeatina/farmacología
18.
Plant Cell Physiol ; 53(7): 1334-43, 2012 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-22642989

RESUMEN

Cytokinins are involved in key developmental processes in rice (Oryza sativa), including the regulation of cell proliferation and grain yield. However, the in vivo action of histidine kinases (OsHks), putative cytokinin receptors, in rice cytokinin signaling remains elusive. This study examined the function and characteristics of OsHk3, 4 and 6 in rice. OsHk6 was highly sensitive to isopentenyladenine (iP) and was capable of restoring cytokinin-dependent ARR6 reporter expression in the ahk2 ahk3 Arabidopsis mutant upon treatment with 1 nM iP. OsHk4 recognized trans-zeatin (tZ) and iP, while OsHk3 scarcely induced cytokinin signaling activity. OsHk4 and OsHk6 mediated the canonical two-component signaling cascade of Arabidopsis to induce phosphorylation of ARR2. OsHk4 and OsHk6 were highly expressed in spikelets, suggesting that tZ and iP might play key roles in grain development. OsHk6 formed a self-interacting homomer in rice protoplasts, although the trans-phosphorylation activity between subunits was much lower than the intra-molecular trans-phosphorylation activity. This indicates that the action mechanism of OsHks is evolutionarily diverged from bacterial histidine kinases. Ectopic expression of OsHk6 in rice calli promoted green pigmentation and subsequent shoot induction, further supporting an OsHk6 in planta function as a cytokinin receptor. From the results of this study, OsHks are homomeric cytokinin receptors with distinctive cytokinin preferences in rice.


Asunto(s)
Regulación de la Expresión Génica de las Plantas , Isopenteniladenosina/farmacología , Oryza/enzimología , Proteínas Quinasas/química , Receptores de Superficie Celular/química , Arabidopsis/química , Arabidopsis/genética , Proteínas de Arabidopsis , Proliferación Celular , Citocininas/química , Citocininas/farmacología , Activación Enzimática , Genes Reporteros , Histidina Quinasa , Inmunoprecipitación , Mutagénesis Sitio-Dirigida , Oryza/efectos de los fármacos , Oryza/genética , Fosforilación , Proteínas de Plantas/química , Proteínas de Plantas/genética , Brotes de la Planta/química , Brotes de la Planta/genética , Plantas Modificadas Genéticamente/enzimología , Plantas Modificadas Genéticamente/genética , Proteínas Quinasas/genética , Protoplastos/química , Receptores de Superficie Celular/genética , Transducción de Señal , Transfección , Zeatina/farmacología
19.
Sci Rep ; 11(1): 6494, 2021 03 22.
Artículo en Inglés | MEDLINE | ID: mdl-33753792

RESUMEN

Cross-talk between phytohormones and sugars is intensely involved in plant metabolism, growth and regeneration. We documented alterations in cytokinin (CK) homeostasis in four developmental stages during de novo shoot organogenesis (DNSO) of kohlrabi (Brassica oleracea var. gongylodes cv. Vienna Purple) seedlings induced by exogenous CKs, trans-zeatin (transZ) and thidiazuron (TDZ), added together with elevated sucrose concentration (6% and 9%). Significant impact of CK and sucrose treatment and their interaction was recorded in all investigated stages, including plantlet development before calli formation (T1 and T2), calli formation (T3) and shoot regeneration (T4). Results showed remarkable increase in total CK levels for transZ treatment, particularly with 9% sucrose. This trend was observed for all physiological and structural groups of CKs. Application of TDZ contributed to little or no increase in CK levels regardless of sucrose concentration. Analysis of expression profiles of organogenesis-related genes involved in auxin transport, CK response, shoot apical meristem formation and cell division revealed that higher sugar concentration significantly downregulated the analysed genes, particularly in T3. This continued on TDZ, but transZ induced an opposite effect with 9% sucrose in T4, increasing gene activity. Our results demonstrated that phytohormone metabolism might be triggered by sucrose signalling in kohlrabi DNSO.


Asunto(s)
Brassica/metabolismo , Genes de Plantas , Compuestos de Fenilurea/farmacología , Reguladores del Crecimiento de las Plantas/farmacología , Sacarosa/metabolismo , Tiadiazoles/farmacología , Zeatina/farmacología , Brassica/genética , Brassica/crecimiento & desarrollo , Regulación de la Expresión Génica de las Plantas , Homeostasis , Meristema/efectos de los fármacos , Meristema/genética , Meristema/crecimiento & desarrollo
20.
Tsitologiia ; 52(12): 1024-30, 2010.
Artículo en Ruso | MEDLINE | ID: mdl-21427982

RESUMEN

Here we have shown that 24 hours after addition of zeatin to the nutrient solution the cytokinin content in xylem sap of wheat plants appears to be about 10 times lower that in the nutrient solution. Cytokinins accumulated mostly in roots and not in shoots of treated plants. These data demonstrate the existence of some barrier on cytokinin pathway from the nutrient solution to the plant shoot. With the help of Sudan III an increase in lignin and suberin deposition in the endodermis could be detected, being stronger with the increase in the distance from the root tip. The increase in deposition of suberin and lignin coincided with the decrease in cytokinin immunolabeling in root cells revealed with the help of monoclonal cytokinin antibodies and the second gold-labelled antibodies. Simultaneously exogenous cytokinins accumulated in root stele cells showing that the Casparian band was not only barrier on cytokinin pathway to plant shoot. It is concluded that high cytokinin immunolabe ling in the stele parenchyma cells around the stele vessels demonstrated accumulation of cytokinins by these cells, which could be important in regulation of cytokinin loading to the xylem vessels during there transport to the shoot. The role of cytokinin transporters is discussed.


Asunto(s)
Citocininas/metabolismo , Reguladores del Crecimiento de las Plantas/farmacología , Raíces de Plantas/metabolismo , Triticum/metabolismo , Xilema/metabolismo , Zeatina/farmacología , Transporte Biológico/efectos de los fármacos , Transporte Biológico/fisiología , Citocininas/farmacología , Raíces de Plantas/citología , Triticum/citología , Xilema/citología
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