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1.
Plant Cell ; 2024 Mar 27.
Artículo en Inglés | MEDLINE | ID: mdl-38781425

RESUMEN

PROTON GRADIENT REGULATION5 (PGR5) is thought to promote cyclic electron flow, and its deficiency impairs photosynthetic control and increases photosensitivity of photosystem (PS) I, leading to seedling lethality under fluctuating light (FL). By screening for Arabidopsis (Arabidopsis thaliana) suppressor mutations that rescue the seedling lethality of pgr5 plants under FL, we identified a portfolio of mutations in 12 different genes. These mutations affect either PSII function, cytochrome b6f (cyt b6f) assembly, plastocyanin (PC) accumulation, the CHLOROPLAST FRUCTOSE-1,6-BISPHOSPHATASE1 (cFBP1), or its negative regulator ATYPICAL CYS HIS-RICH THIOREDOXIN2 (ACHT2). The characterization of the mutants indicates that the recovery of viability can in most cases be explained by the restoration of PSI donor side limitation, which is caused by reduced electron flow to PSI due to defects in PSII, cyt b6f, or PC. Inactivation of cFBP1 or its negative regulator ACHT2 results in increased levels of the NADH dehydrogenase-like complex. This increased activity may be responsible for suppressing the pgr5 phenotype under FL conditions. Plants that lack both PGR5 and DE-ETIOLATION-INDUCED PROTEIN1 (DEIP1)/NEW TINY ALBINO1 (NTA1), previously thought to be essential for cyt b6f assembly, are viable and accumulate cyt b6f. We suggest that PGR5 can have a negative effect on the cyt b6f complex and that DEIP1/NTA1 can ameliorate this negative effect.

2.
Plant Physiol ; 190(3): 1866-1882, 2022 10 27.
Artículo en Inglés | MEDLINE | ID: mdl-35946785

RESUMEN

The PROTON GRADIENT REGULATION5 (PGR5) protein is required for trans-thylakoid proton gradient formation and acclimation to fluctuating light (FL). PGR5 functionally interacts with two other thylakoid proteins, PGR5-like 1 (PGRL1) and 2 (PGRL2); however, the molecular details of these interactions are largely unknown. In the Arabidopsis (Arabidopsis thaliana) pgr5-1 mutant, the PGR5G130S protein accumulates in only small amounts. In this work, we generated a knockout allele of PGR5 (pgr5-Cas) using CRISPR-Cas9 technology. Like pgr5-1, pgr5-Cas is seedling-lethal under FL, but photosynthesis and particularly cyclic electron flow, as well as chlorophyll content, are less severely affected in both pgr5-Cas and pgrl1ab (which lacks PGRL1 and PGR5) than in pgr5-1. These differences are associated with changes in the levels of 260 proteins, including components of the Calvin-Benson cycle, photosystems II and I, and the NDH complex, in pgr5-1 relative to the wild type (WT), pgr5-Cas, and pgrl1ab. Some of the differences between pgr5-1 and the other mutant lines could be tentatively assigned to second-site mutations in the pgr5-1 line, identified by whole-genome sequencing. However, others, particularly the more pronounced photosynthetic defects and PGRL1 depletion (compared to pgr5-Cas), are clearly due to specific negative effects of the amino-acid substitution in PGR5G130S, as demonstrated by complementation analysis. Moreover, pgr5-1 and pgr5-Cas plants are less tolerant to long-term exposure to high light than pgrl1ab plants. These results imply that, in addition to the previously reported necessity of PGRL1 for optimal PGR5 function, PGR5 is required alongside PGRL1 to avoid harmful effects on plant performance.


Asunto(s)
Proteínas de Arabidopsis , Arabidopsis , Proteínas del Complejo del Centro de Reacción Fotosintética , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/metabolismo , Protones , Complejo de Proteína del Fotosistema I/genética , Complejo de Proteína del Fotosistema I/metabolismo , Transporte de Electrón , Fotosíntesis/genética , Luz , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Proteínas de la Membrana/metabolismo
3.
Plant J ; 106(1): 23-40, 2021 04.
Artículo en Inglés | MEDLINE | ID: mdl-33368770

RESUMEN

Acclimation is the capacity to adapt to environmental changes within the lifetime of an individual. This ability allows plants to cope with the continuous variation in ambient conditions to which they are exposed as sessile organisms. Because environmental changes and extremes are becoming even more pronounced due to the current period of climate change, enhancing the efficacy of plant acclimation is a promising strategy for mitigating the consequences of global warming on crop yields. At the cellular level, the chloroplast plays a central role in many acclimation responses, acting both as a sensor of environmental change and as a target of cellular acclimation responses. In this Perspective article, we outline the activities of the Green Hub consortium funded by the German Science Foundation. The main aim of this research collaboration is to understand and strategically modify the cellular networks that mediate plant acclimation to adverse environments, employing Arabidopsis, tobacco (Nicotiana tabacum) and Chlamydomonas as model organisms. These efforts will contribute to 'smart breeding' methods designed to create crop plants with improved acclimation properties. To this end, the model oilseed crop Camelina sativa is being used to test modulators of acclimation for their potential to enhance crop yield under adverse environmental conditions. Here we highlight the current state of research on the role of gene expression, metabolism and signalling in acclimation, with a focus on chloroplast-related processes. In addition, further approaches to uncovering acclimation mechanisms derived from systems and computational biology, as well as adaptive laboratory evolution with photosynthetic microbes, are highlighted.


Asunto(s)
Hojas de la Planta/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Arabidopsis/fisiología , Camellia/genética , Camellia/metabolismo , Camellia/fisiología , Chlamydomonas/genética , Chlamydomonas/metabolismo , Chlamydomonas/fisiología , Hojas de la Planta/genética , Biología de Sistemas/métodos , Nicotiana/genética , Nicotiana/metabolismo , Nicotiana/fisiología
4.
Plant J ; 99(3): 521-535, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-31002470

RESUMEN

GUN1 integrates retrograde signals in chloroplasts but the underlying mechanism is elusive. FUG1, a chloroplast translation initiation factor, and GUN1 are co-expressed at the transcriptional level, and FUG1 co-immunoprecipitates with GUN1. We used mutants of GUN1 (gun1-103) and FUG1 (fug1-3) to analyse their functional relationship at the physiological and system-wide level, the latter including transcriptome and proteome analyses. Absence of GUN1 aggravates the effects of decreased FUG1 levels on chloroplast protein translation, resulting in transiently more pronounced phenotypes regarding photosynthesis, leaf colouration, growth and cold acclimation. The gun1-103 mutation also enhances variegation in the var2 mutant, increasing the fraction of white sectors, while fug1-3 suppresses the var2 phenotype. The transcriptomes of fug1-3 and gun1-103 plants are very similar, but absence of GUN1 alone has almost no effect on protein levels, whereas steady-state levels of chloroplast proteins are markedly decreased in fug1-3. In fug1 gun1 double mutants, effects on transcriptomes and particularly on proteomes are enhanced. Our results show that GUN1 function becomes critical when chloroplast proteostasis is perturbed by decreased rates of synthesis (fug1) or degradation (var2) of chloroplast proteins, or by low temperatures. The functions of FUG1 and GUN1 appear to be related, corroborating the view that GUN1 helps to maintain chloroplast protein homeostasis (proteostasis).


Asunto(s)
Proteínas de Arabidopsis/genética , Arabidopsis/genética , Proteínas de Cloroplastos/genética , Cloroplastos/genética , Proteínas de Unión al ADN/genética , Factor 2 Eucariótico de Iniciación/genética , Proteostasis/genética , Aclimatación/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/clasificación , Proteínas de Arabidopsis/metabolismo , Proteínas de Cloroplastos/metabolismo , Cloroplastos/metabolismo , Frío , Proteínas de Unión al ADN/metabolismo , Factor 2 Eucariótico de Iniciación/metabolismo , Perfilación de la Expresión Génica , Regulación de la Expresión Génica de las Plantas , Mutación , Fenotipo , Filogenia , Hojas de la Planta/genética , Hojas de la Planta/metabolismo , Plantas Modificadas Genéticamente
5.
Proc Natl Acad Sci U S A ; 114(45): 12069-12074, 2017 11 07.
Artículo en Inglés | MEDLINE | ID: mdl-29078290

RESUMEN

Thiol-dependent redox regulation allows the rapid adaptation of chloroplast function to unpredictable changes in light intensity. Traditionally, it has been considered that chloroplast redox regulation relies on photosynthetically reduced ferredoxin (Fd), thioredoxins (Trxs), and an Fd-dependent Trx reductase (FTR), the Fd-FTR-Trxs system, which links redox regulation to light. More recently, a plastid-localized NADPH-dependent Trx reductase (NTR) with a joint Trx domain, termed NTRC, was identified. NTRC efficiently reduces 2-Cys peroxiredoxins (Prxs), thus having antioxidant function, but also participates in redox regulation of metabolic pathways previously established to be regulated by Trxs. Thus, the NTRC, 2-Cys Prxs, and Fd-FTR-Trxs redox systems may act concertedly, but the nature of the relationship between them is unknown. Here we show that decreased levels of 2-Cys Prxs suppress the phenotype of the Arabidopsis thaliana ntrc KO mutant. The excess of oxidized 2-Cys Prxs in NTRC-deficient plants drains reducing power from chloroplast Trxs, which results in low efficiency of light energy utilization and impaired redox regulation of Calvin-Benson cycle enzymes. Moreover, the dramatic phenotype of the ntrc-trxf1f2 triple mutant, lacking NTRC and f-type Trxs, was also suppressed by decreased 2-Cys Prxs contents, as the ntrc-trxf1f2-Δ2cp mutant partially recovered the efficiency of light energy utilization and exhibited WT rate of CO2 fixation and growth phenotype. The suppressor phenotype was not caused by compensatory effects of additional chloroplast antioxidant systems. It is proposed that the Fd-FTR-Trx and NTRC redox systems are linked by the redox balance of 2-Cys Prxs, which is crucial for chloroplast function.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Peroxirredoxinas/metabolismo , Fotosíntesis/fisiología , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Antioxidantes/metabolismo , Arabidopsis/metabolismo , Cloroplastos/metabolismo , Ferredoxinas/metabolismo , Oxidación-Reducción , Plastidios/metabolismo , Tiorredoxinas/metabolismo
6.
Plant Physiol ; 169(3): 1766-86, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26338951

RESUMEN

Two different thiol redox systems exist in plant chloroplasts, the ferredoxin-thioredoxin (Trx) system, which depends on ferredoxin reduced by the photosynthetic electron transport chain and, thus, on light, and the NADPH-dependent Trx reductase C (NTRC) system, which relies on NADPH and thus may be linked to sugar metabolism in the dark. Previous studies suggested, therefore, that the two different systems may have different functions in plants. We now report that there is a previously unrecognized functional redundancy of Trx f1 and NTRC in regulating photosynthetic metabolism and growth. In Arabidopsis (Arabidopsis thaliana) mutants, combined, but not single, deficiencies of Trx f1 and NTRC led to severe growth inhibition and perturbed light acclimation, accompanied by strong impairments of Calvin-Benson cycle activity and starch accumulation. Light activation of key enzymes of these pathways, fructose-1,6-bisphosphatase and ADP-glucose pyrophosphorylase, was almost completely abolished. The subsequent increase in NADPH-NADP(+) and ATP-ADP ratios led to increased nitrogen assimilation, NADP-malate dehydrogenase activation, and light vulnerability of photosystem I core proteins. In an additional approach, reporter studies show that Trx f1 and NTRC proteins are both colocalized in the same chloroplast substructure. Results provide genetic evidence that light- and NADPH-dependent thiol redox systems interact at the level of Trx f1 and NTRC to coordinately participate in the regulation of the Calvin-Benson cycle, starch metabolism, and growth in response to varying light conditions.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Arabidopsis/fisiología , Arabidopsis/efectos de la radiación , Proteínas de Arabidopsis/genética , Cloroplastos/metabolismo , Glucosa-1-Fosfato Adenililtransferasa/genética , Glucosa-1-Fosfato Adenililtransferasa/metabolismo , Malato-Deshidrogenasa (NADP+)/genética , Malato-Deshidrogenasa (NADP+)/metabolismo , Metaboloma , Oxidación-Reducción , Fenotipo , Fotosíntesis/efectos de la radiación , Complejo de Proteína del Fotosistema I/metabolismo , Hojas de la Planta/enzimología , Hojas de la Planta/genética , Hojas de la Planta/fisiología , Hojas de la Planta/efectos de la radiación , Transpiración de Plantas/efectos de la radiación , Almidón/metabolismo , Reductasa de Tiorredoxina-Disulfuro/genética
7.
Plant Cell Environ ; 39(4): 804-22, 2016 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-26476233

RESUMEN

High irradiances may lead to photooxidative stress in plants, and non-photochemical quenching (NPQ) contributes to protection against excess excitation. One of the NPQ mechanisms, qE, involves thermal dissipation of the light energy captured. Importantly, plants need to tune down qE under light-limiting conditions for efficient utilization of the available quanta. Considering the possible redox control of responses to excess light implying enzymes, such as thioredoxins, we have studied the role of the NADPH thioredoxin reductase C (NTRC). Whereas Arabidopsis thaliana plants lacking NTRC tolerate high light intensities, these plants display drastically elevated qE, have larger trans-thylakoid ΔpH and have 10-fold higher zeaxanthin levels under low and medium light intensities, leading to extremely low linear electron transport rates. To test the impact of the high qE on plant growth, we generated an ntrc-psbs double-knockout mutant, which is devoid of qE. This double mutant grows faster than the ntrc mutant and has a higher chlorophyll content. The photosystem II activity is partially restored in the ntrc-psbs mutant, and linear electron transport rates under low and medium light intensities are twice as high as compared with plants lacking ntrc alone. These data uncover a new role for NTRC in the control of photosynthetic yield.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/enzimología , Arabidopsis/efectos de la radiación , Cloroplastos/enzimología , Luz , Fotosíntesis/efectos de la radiación , Reductasa de Tiorredoxina-Disulfuro/metabolismo , Arabidopsis/efectos de los fármacos , Arabidopsis/fisiología , Proteínas de Arabidopsis/genética , Clorofila/metabolismo , Cloroplastos/efectos de los fármacos , Cloroplastos/efectos de la radiación , Ditiotreitol/farmacología , Transporte de Electrón/efectos de los fármacos , Transporte de Electrón/efectos de la radiación , Fluorescencia , Técnicas de Inactivación de Genes , Mutación/genética , Nigericina/farmacología , Peroxirredoxinas/metabolismo , Fotosíntesis/efectos de los fármacos , Complejo de Proteína del Fotosistema I/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Reductasa de Tiorredoxina-Disulfuro/genética , Xantófilas/metabolismo
8.
J Exp Bot ; 67(6): 1951-64, 2016 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-26842981

RESUMEN

Redox regulation plays a central role in the adaptation of chloroplast metabolism to light. Extensive biochemical analyses in vitro have identified f-type thioredoxins (Trxs) as the most important catalysts for light-dependent reduction and activation of the enzymes of the Calvin-Benson cycle. However, the precise function of type f Trxs in vivo and their impact on plant growth are still poorly known. To address this issue we have generated an Arabidopsis thaliana double knock-out mutant, termed trxf1f2, devoid of both f1 and f2 Trxs. Despite the essential function previously proposed for f-type Trxs, the visible phenotype of the trxf1f2 double mutant was virtually indistinguishable from the wild type when grown under a long-day photoperiod. However, the Trx f-deficient plants showed growth inhibition under a short-day photoperiod which was not rescued at high light intensity. The absence of f-type Trxs led to significantly lower photosynthetic electron transport rates and higher levels of non-photochemical energy quenching. Notably, the Trx f null mutant suffered from a shortage of photosystem I electron acceptors and delayed activation of carbon dioxide fixation following a dark-light transition. Two redox-regulated Calvin-Benson cycle enzymes, fructose 1,6-bisphosphatase (FBPase) and Rubisco activase, showed retarded and incomplete reduction in the double mutant upon illumination, compared with wild-type plants. These results show that the function of f-type Trxs in the rapid activation of carbon metabolism in response to light is not entirely compensated for by additional plastid redox systems, and suggest that these Trxs have an important role in the light adjustment of photosynthetic metabolism.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/crecimiento & desarrollo , Arabidopsis/metabolismo , Carbono/metabolismo , Tiorredoxinas en Cloroplasto/metabolismo , Fotoperiodo , Arabidopsis/enzimología , Arabidopsis/genética , Dióxido de Carbono/metabolismo , Transporte de Electrón/efectos de la radiación , Regulación de la Expresión Génica de las Plantas/efectos de la radiación , Cinética , Luz , Mutación/genética , Oxidación-Reducción/efectos de la radiación , Fotosíntesis/efectos de la radiación , Complejo de Proteína del Fotosistema I/metabolismo , Complejo de Proteína del Fotosistema II/metabolismo , Desarrollo de la Planta/efectos de la radiación , Plantas Modificadas Genéticamente , ARN Mensajero/genética , ARN Mensajero/metabolismo
10.
Trends Plant Sci ; 27(6): 577-587, 2022 06.
Artículo en Inglés | MEDLINE | ID: mdl-35012879

RESUMEN

Under natural environmental conditions, changes in light intensity and temperature are closely interwoven, and of all organelles, only chloroplasts react strongly upon alterations of these two parameters. We review increasing evidence indicating that changes in chloroplast metabolism are critical for the comprehensive cellular answer in a challenging environment. This cellular answer starts with rapid modifications of thylakoid-located processes, followed by modifications in the stroma and transport activities across the chloroplast envelope. We propose that the 'modulators' involved contribute to plant stress tolerance and that deciphering of their characteristics is essential to understand 'acclimation'. Especially in times of climatic changes, we must gain knowledge on physiological reactions that might become instrumental for directed breeding strategies aiming to develop stress-tolerant crop plants.


Asunto(s)
Cloroplastos , Fitomejoramiento , Cloroplastos/metabolismo , Luz , Fotosíntesis , Plantas/metabolismo , Estrés Fisiológico/fisiología , Temperatura , Tilacoides/metabolismo
11.
Antioxidants (Basel) ; 10(6)2021 Jun 03.
Artículo en Inglés | MEDLINE | ID: mdl-34204867

RESUMEN

Non-photochemical quenching (NPQ) protects plants from the detrimental effects of excess light. NPQ is rapidly induced by the trans-thylakoid proton gradient during photosynthesis, which in turn requires PGR5/PGRL1-dependent cyclic electron flow (CEF). Thus, Arabidopsis thaliana plants lacking either protein cannot induce transient NPQ and die under fluctuating light conditions. Conversely, the NADPH-dependent thioredoxin reductase C (NTRC) is required for efficient energy utilization and plant growth, and in its absence, transient and steady-state NPQ is drastically increased. How NTRC influences NPQ and functionally interacts with CEF is unclear. Therefore, we generated the A. thaliana line pgr5 ntrc, and found that the inactivation of PGR5 suppresses the high transient and steady-state NPQ and impaired growth phenotypes observed in the ntrc mutant under short-day conditions. This implies that NTRC negatively influences PGR5 activity and, accordingly, the lack of NTRC is associated with decreased levels of PGR5, possibly pointing to a mechanism to restrict upregulation of PGR5 activity in the absence of NTRC. When exposed to high light intensities, pgr5 ntrc plants display extremely impaired photosynthesis and growth, indicating additive effects of lack of both proteins. Taken together, these findings suggest that the interplay between NTRC and PGR5 is relevant for photoprotection and that NTRC might regulate PGR5 activity.

12.
Nat Commun ; 12(1): 3941, 2021 06 24.
Artículo en Inglés | MEDLINE | ID: mdl-34168134

RESUMEN

In plants, inactivation of either of the thylakoid proteins PGR5 and PGRL1 impairs cyclic electron flow (CEF) around photosystem I. Because PGR5 is unstable in the absence of the redox-active PGRL1, but not vice versa, PGRL1 is thought to be essential for CEF. However, we show here that inactivation of PGRL2, a distant homolog of PGRL1, relieves the need for PGRL1 itself. Conversely, high levels of PGRL2 destabilize PGR5 even when PGRL1 is present. In the absence of both PGRL1 and PGRL2, PGR5 alters thylakoid electron flow and impairs plant growth. Consequently, PGR5 can operate in CEF on its own, and is the target of the CEF inhibitor antimycin A, but its activity must be modulated by PGRL1. We conclude that PGRL1 channels PGR5 activity, and that PGRL2 triggers the degradation of PGR5 when the latter cannot productively interact with PGRL1.


Asunto(s)
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de la Membrana/metabolismo , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Antimicina A/farmacología , Arabidopsis/efectos de los fármacos , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas Fluorescentes Verdes/genética , Luz , Proteínas de la Membrana/genética , Mutación , Fotosíntesis , Proteínas del Complejo del Centro de Reacción Fotosintética/genética , Plantas Modificadas Genéticamente , Estabilidad Proteica
13.
J Bacteriol ; 192(13): 3345-51, 2010 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-20400539

RESUMEN

It is well established that respiratory organisms use proton motive force to produce ATP via F-type ATP synthase aerobically and that this process may reverse during anaerobiosis to produce proton motive force. Here, we show that Shewanella oneidensis strain MR-1, a nonfermentative, facultative anaerobe known to respire exogenous electron acceptors, generates ATP primarily from substrate-level phosphorylation under anaerobic conditions. Mutant strains lacking ackA (SO2915) and pta (SO2916), genes required for acetate production and a significant portion of substrate-level ATP produced anaerobically, were tested for growth. These mutant strains were unable to grow anaerobically with lactate and fumarate as the electron acceptor, consistent with substrate-level phosphorylation yielding a significant amount of ATP. Mutant strains lacking ackA and pta were also shown to grow slowly using N-acetylglucosamine as the carbon source and fumarate as the electron acceptor, consistent with some ATP generation deriving from the Entner-Doudoroff pathway with this substrate. A deletion strain lacking the sole F-type ATP synthase (SO4746 to SO4754) demonstrated enhanced growth on N-acetylglucosamine and a minor defect with lactate under anaerobic conditions. ATP synthase mutants grown anaerobically on lactate while expressing proteorhodopsin, a light-dependent proton pump, exhibited restored growth when exposed to light, consistent with a proton-pumping role for ATP synthase under anaerobic conditions. Although S. oneidensis requires external electron acceptors to balance redox reactions and is not fermentative, we find that substrate-level phosphorylation is its primary anaerobic energy conservation strategy. Phenotypic characterization of an ackA deletion in Shewanella sp. strain MR-4 and genomic analysis of other sequenced strains suggest that this strategy is a common feature of Shewanella.


Asunto(s)
Proteínas Bacterianas/metabolismo , Shewanella/crecimiento & desarrollo , Shewanella/metabolismo , Anaerobiosis , Proteínas Bacterianas/genética , Cromatografía Líquida de Alta Presión , Prueba de Complementación Genética , Modelos Biológicos , Mutación , Fosforilación , Shewanella/genética
14.
Appl Environ Microbiol ; 76(13): 4123-9, 2010 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-20453141

RESUMEN

Microorganisms can use complex photosystems or light-dependent proton pumps to generate membrane potential and/or reduce electron carriers to support growth. The discovery that proteorhodopsin is a light-dependent proton pump that can be expressed readily in recombinant bacteria enables development of new strategies to probe microbial physiology and to engineer microbes with new light-driven properties. Here, we describe functional expression of proteorhodopsin and light-induced changes in membrane potential in the bacterium Shewanella oneidensis strain MR-1. We report that there were significant increases in electrical current generation during illumination of electrochemical chambers containing S. oneidensis expressing proteorhodopsin. We present evidence that an engineered strain is able to consume lactate at an increased rate when it is illuminated, which is consistent with the hypothesis that proteorhodopsin activity enhances lactate uptake by increasing the proton motive force. Our results demonstrate that there is coupling of a light-driven process to electricity generation in a nonphotosynthetic engineered bacterium. Expression of proteorhodopsin also preserved the viability of the bacterium under nutrient-limited conditions, providing evidence that fulfillment of basic energy needs of organisms may explain the widespread distribution of proteorhodopsin in marine environments.


Asunto(s)
Electricidad , Ingeniería Genética/métodos , Luz , Rodopsina/metabolismo , Shewanella/crecimiento & desarrollo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Biotecnología/métodos , Membrana Celular/metabolismo , Conjugación Genética , Medios de Cultivo , Escherichia coli/genética , Escherichia coli/metabolismo , Potenciales de la Membrana/fisiología , Bombas de Protones/fisiología , Rodopsina/genética , Rodopsina/fisiología , Rodopsinas Microbianas , Shewanella/genética , Shewanella/metabolismo , Shewanella/fisiología
15.
Dent J (Basel) ; 7(3)2019 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-31266163

RESUMEN

New technologies are changing the therapeutical options to do indirect restorations and new adhesive systems are continuously introduced to be used by clinicians. Different interactions between restorations, adhesive systems components, enamel and dentin require having criteria based on the selection of the adhesive system, ensuring the longevity of the restorations and the preservation of the biological remnant. The adhesion force to the dental tissue is one of the indicatives of the behavior of the adhesive systems and influences the behavior of the treatments with direct and indirect restorations. The objective of this search was to find the adhesive systems with the best results in terms of the adhesion strength of indirect restorations on the dental tissues. The search was conducted in two MEDLINE digital databases (PubMed), and the Cochrane Library with a search strategy based on the combination of MeSH (Medical Subject Headings) keywords. This systematic review used the PRISMA guide (Preferred Reporting Items for Systematic Reviews and Meta-Analysis). According to this review, the 3-step adhesive systems were the best performing and still are the gold standard for the cementing of indirect restorations. In addition, it can be concluded that self-etched adhesive systems reduce the time spent in clinical practice, however at the interface level they behave as permeable membranes more susceptible to degradation.

16.
Sci Rep ; 9(1): 8095, 2019 05 30.
Artículo en Inglés | MEDLINE | ID: mdl-31147586

RESUMEN

Currently, uterus transplantation (UTx) is a clinical option for infertile women. Over the past three decades, treating benign or malignant gynecological diseases with minimally invasive gynecological surgery has improved, providing significant advantages over conventional open surgery. This study addresses the method used for laparoscopic live-donor ovariohysterectomy and graft harvest from a sheep model. Using a microsurgical practice, ten grafts were autotransplanted after uterine perfusion. End-to-end anastomosis techniques were used to approximate veins and arteries. Follow-ups were carried out 2-months after surgery and postoperative studies included ultrasound scan, diagnostic hysteroscopy, vascular angiography, and exploratory laparoscopy. All transplants were completed without complications. After vascular anastomosis, total reperfusion of the tissue was accomplished in all animals without confirmation of arterial or venous thrombosis. Angiographic explorations did not show any statistically significant dissimilarity in the arterial diameters between the different examination times. 3-months after uterine transplantation all animals underwent assisted reproduction techniques. Patent uterine arteries were observed 4, 8 and 12 months after the transplant. 6-months after transplantation, six sheep (60%) became pregnant with assisted reproduction practices. We noticed an increase in the degree of fibrosis of the cervix samples in non-pregnant animals of the transplant group. Laparoscopic surgery can be an advantageous approach for the uterus retrieval procedure during uterine transplantation. However, larger sample sized reports are needed in order to accomplish validation, standardization and wider use of this route.


Asunto(s)
Histerectomía/métodos , Infertilidad Femenina/terapia , Laparoscopía/métodos , Recolección de Tejidos y Órganos/métodos , Útero/trasplante , Animales , Estudios de Factibilidad , Femenino , Fibrosis , Humanos , Laparoscopía/efectos adversos , Donadores Vivos , Modelos Animales , Preservación de Órganos/efectos adversos , Preservación de Órganos/métodos , Perfusión/efectos adversos , Perfusión/métodos , Embarazo , Técnicas Reproductivas Asistidas , Ovinos , Recolección de Tejidos y Órganos/efectos adversos , Trasplante Autólogo/efectos adversos , Trasplante Autólogo/métodos , Útero/patología
17.
Arch Esp Urol ; 71(1): 73-84, 2018 Jan.
Artículo en Español | MEDLINE | ID: mdl-29336335

RESUMEN

Urology needs models of competencies assessment, although there is a wide range of tools not yet integrated into the official training programs. CONTEXT: At present, there is no universal framework for measuring surgeons' level of competence. Urology training programs should provide and consider knowledge, pyschomotor/cognitive skills, and simulator, cadaver or animal models-based training. Validity is a complex concept that refers to the capacity of the evaluation tool, so it is necessary to demonstrate several types of validation to assure the capacity of a method, reinforced with different reliability tests and calculation of internal consistency between evaluators. OBJECTIVE: Based on a structured dossier of surgical skills, classified by groups, the ESSCOLAP® Basic system was proposed with 5 simulator tasks to evaluate basic laparoscopic skills. Once validated in the JUMISC (Spain), the tool was proposed to extend its scope and implementation in other locations. RESULTS: Our system has not yet demonstrated a full validity in the real clinical setting because a predictive validity needs to be demonstrated on the basis of clinical data. It also suffers from a certain range of subjectivity, thus implying clear and defined criteria for any situation. Factors like the number of evaluators and tasks to assess will influence the reliability tests that measure the degree of agreement between evaluators, so that a higher number of evaluated cases would imply a greater reliability of our system. Finally, we assume that the incorporation of this type of tools implies an added cost, charged to the public and private responsible institutions, which will only be considered cost-effective when it is demonstrated its real and positive traceability in health outcomes. CONCLUSIONS: ESSCOLAP® Basic, of quick and simple implementation capacity, has been validated and calibrated for the evaluation of basic technical skills in laparoscopy.


Asunto(s)
Competencia Clínica , Urología/educación , Entrenamiento Simulado
18.
Toxins (Basel) ; 9(9)2017 09 19.
Artículo en Inglés | MEDLINE | ID: mdl-28925975

RESUMEN

Viral M-dsRNAs encoding yeast killer toxins share similar genomic organization, but no overall sequence identity. The dsRNA full-length sequences of several known M-viruses either have yet to be completed, or they were shorter than estimated by agarose gel electrophoresis. High-throughput sequencing was used to analyze some M-dsRNAs previously sequenced by traditional techniques, and new dsRNAs from atypical killer strains of Saccharomyces cerevisiae and Torulaspora delbrueckii. All dsRNAs expected to be present in a given yeast strain were reliably detected and sequenced, and the previously-known sequences were confirmed. The few discrepancies between viral variants were mostly located around the central poly(A) region. A continuous sequence of the ScV-M2 genome was obtained for the first time. M1 virus was found for the first time in wine yeasts, coexisting with Mbarr-1 virus in T. delbrueckii. Extra 5'- and 3'-sequences were found in all M-genomes. The presence of repeated short sequences in the non-coding 3'-region of most M-genomes indicates that they have a common phylogenetic origin. High identity between amino acid sequences of killer toxins and some unclassified proteins of yeast, bacteria, and wine grapes suggests that killer viruses recruited some sequences from the genome of these organisms, or vice versa, during evolution.


Asunto(s)
Genoma Viral , ARN Viral/genética , Saccharomyces cerevisiae/virología , Torulaspora/virología , Virus/genética , Genotipo , Secuenciación de Nucleótidos de Alto Rendimiento , Factores Asesinos de Levadura/genética , Fenotipo , Saccharomyces cerevisiae/genética , Torulaspora/genética
19.
Front Plant Sci ; 7: 1050, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-27536301

RESUMEN

Iron limitation is the major factor controlling phytoplankton growth in vast regions of the contemporary oceans. In this study, a combination of thermoluminescence (TL), chlorophyll fluorescence, and P700 absorbance measurements have been used to elucidate the effects of iron deficiency in the photosynthetic electron transport of the marine diatom P. tricornutum. TL was used to determine the effects of iron deficiency on photosystem II (PSII) activity. Excitation of iron-replete P. tricornutum cells with single turn-over flashes induced the appearance of TL glow curves with two components with different peaks of temperature and contributions to the total signal intensity: the B band (23°C, 63%), and the AG band (40°C, 37%). Iron limitation did not significantly alter these bands, but induced a decrease of the total TL signal. Far red excitation did not increase the amount of the AG band in iron-limited cells, as observed for iron-replete cells. The effect of iron deficiency on the photosystem I (PSI) activity was also examined by measuring the changes in P700 redox state during illumination. The electron donation to PSI was substantially reduced in iron-deficient cells. This could be related with the important decline on cytochrome c 6 content observed in these cells. Iron deficiency also induced a marked increase in light sensitivity in P. tricornutum cells. A drastic increase in the level of peroxidation of chloroplast lipids was detected in iron-deficient cells even when grown under standard conditions at low light intensity. Illumination with a light intensity of 300 µE m(-2) s(-1) during different time periods caused a dramatic disappearance in TL signal in cells grown under low iron concentration, this treatment not affecting to the signal in iron-replete cells. The results of this work suggest that iron deficiency induces partial blocking of the electron transfer between PSII and PSI, due to a lower concentration of the electron donor cytochrome c 6. This decreased electron transfer may induce the over-reduction of the plastoquinone pool and consequently the appearance of acceptor side photoinhibition in PSII even at low light intensities. The functionality of chlororespiratory electron transfer pathway under iron restricted conditions is also discussed.

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