Caracterización de la familia génica de las nucleosidasas en Phaseolus vulgaris

Los nucleótidos son moléculas de vital importancia en todos los seres vivos, ya que, además de ser componentes esenciales de los ácidos nucleicos, participan en procesos bioenergéticos, forman parte de cofactores de enzimas y de segundos mensajeros, y son precursores para la síntesis de metabolitos primarios y secundarios. En plantas, los nucleótidos de purina y de pirimidina pueden proceder de la síntesis de novo, el reciclado de nucleósidos y bases nitrogenadas o pueden absorberse directamente del suelo. Como en la mayoría de animales y microorganismos, el catabolismo de pirimidinas en plantas conlleva la reducción de uracilo y timina hasta CO2, NH3 y β-alanina o β - aminoisobutirato, respectivamente. En cuanto al catabolismo de purinas, las plantas pueden oxidarlas completamente y recuperar el nitrógeno presente en ellas, mientras que la mayoría de los animales no. Otra particularidad del metabolismo de purinas en plantas es su relevancia en las leguminosas ureídicas, ya que la nucleobase xantina es el precursor de los ureidos alantoína y alantoato, moléculas claves en el transporte y almacenamiento de nitrógeno en este grupo de leguminosas. Además, en los últimos años se ha sugerido una conexión importante entre el catabolismo de los ácidos nucleicos y nucleótidos y el de ureidos en distintos procesos en los que se produce una elevada movilización de nitrógeno. Tanto las rutas de síntesis de novo como las de reciclaje de nucleótidos confluyen en la formación de nucleótidos monofosfato, que son los sustratos de las 5’-nucleotidasas, fosfatasas que catalizan el primer paso de la ruta de degradación. Un elemento clave en la ruta de degradación de purinas es la formación del nucleósido xantosina, ya que marca un punto de no retorno en la degradación de purinas, pues no puede ser reciclada. El siguiente paso en la ruta de degradación de purinas y pirimidinas lo catalizan las nucleosidasas (nucleósido hidrolasas, NSH), enzimas que hidrolizan los nucleósidos hasta la base nitrogenada y ribosa, y que se han postulado como enzimas clave en el control del equilibrio entre degradación y reciclaje de nucleótidos. Las nucleosidasas también pueden jugar un papel importante en procesos de señalización, ya que para que el eATP, una molécula señalizadora extracelular, actúe de manera eficiente, debe de poder ser retirado del apoplasto una vez ejerza su función, lo que ocurre por la acción concertada de apirasas y nucleotidasas y nucleosidasas extracelulares. Estas nucleosidasas también podrían participar en la recuperación de nucleósidos del suelo o los que son liberados cuando una célula se daña. El objetivo principal de este trabajo ha sido la caracterización de la familia génica de las nucleosidasas en la leguminosa ureídica Phaseolus vulgaris. Para ello, se han clonado los genes que codifican dichas enzimas en judía, lo que ha permitido determinar que dicha familia está formada por, al menos, cuatro genes, dos de localización citosólica (PvNSH1 y PvNSH2) y dos de localización apoplástica (PvNSH3 y PvNSH4). Además, los ORFs de dichos genes se han sobreexpresado en sistemas heterólogos, lo que ha posibilitado la purificación de las enzimas correspondientes y su posterior caracterización. PvNSH1 presenta alta afinidad por nucleósidos de purina y pirimidina, mientras que PvNSH2 parece más específica de purinas y, en particular, de xantosina. En cuanto a las nucleosidasas apoplásticas, PvNSH4 es activa a pH ácido y utiliza, principalmente, inosina como sustrato, mientras que no se ha podido caracterizar la actividad de PvNSH3. Cabe destacar, que el análisis filogenético de las nucleosidasas apoplásticas sugiere que estas enzimas tienen un origen evolutivo relativamente reciente, ya que sólo se han identificado en algunos grupos de plantas, pero no en organismos pertenecientes a otros reinos. Por otro lado, se ha investigado el papel fisiológico de las nucleosidasas en judía. Los resultados obtenidos muestran que PvNSH1 y PvNSH2 juegan un papel relevante durante la germinación y el desarrollo postgerminativo de judía, lo que resalta la importancia del metabolismo de nucleótidos en esta fase de desarrollo. Además, se ha puesto de manifiesto la importancia de estas enzimas en los nódulos de judía, en los que su expresión y actividad es muy elevada, aunque no se ha podido demostrar una relación entre sus niveles y los de ureidos ni en raíces ni en nódulos. En cuanto a las nucleosidasas con posible localización apoplástica, se ha demostrado esta ubicación celular y que PvNSH4 podría estar implicada en situaciones de respuesta a estrés y, en concreto, en aquellas que están mediadas por la hormona metil jasmonato. Por otro lado, no se ha podido obtener información acerca de la función de PvNSH3, ya que se han detectado niveles de expresión muy reducidos y sin cambios significativos en ninguna de las situaciones estudiadas.Nucleotides are molecules of vital importance in all living organisms, as they are not only essential components of nucleic acids, but also participate in bioenergetic processes, are core element of cofactors and second messengers and are precursors for the synthesis of primary and secondary metabolites. In plants, purine and pyrimidine nucleotides can be originate from de novo synthesis, salvage of nucleosides and nucleobases or can be absorbed directly from the soil. As in most animals and microorganisms, pyrimidine catabolism in plants involves the reduction of uracil and thymine to CO2, NH3 and β-alanine or β-aminoisobutyrate, respectively. Regarding purine catabolism, plants can completely oxidise purines and recover the nitrogen present in them, while most animals cannot. Another peculiarity of purine metabolism in plants is its relevance in ureidic legumes, since the nucleobase xanthine is the precursor of the ureides allantoin and allantoate, key molecules in the transport and storage of nitrogen in this group of legumes. Furthermore, in recent years, an important connection between the catabolism of nucleic acids and nucleotides and that of ureides has been suggested in different processes involving high nitrogen mobilisation. Both the de novo synthesis and nucleotide salvage pathways converge in the formation of monophosphate nucleotides, which are the substrates of 5'-nucleotidases, phosphatases that catalyse the first step of the degradation pathway. A key element in the purine degradation pathway is the formation of the nucleoside xanthosine, as it marks a point of no return in purine degradation, because it cannot be recycled. The next step in purine and pyrimidine degradation pathways is catalysed by nucleosidases (nucleoside hydrolases, NSH), enzymes that hydrolyse nucleosides to the nucleobase and ribose, and which have been postulated as key enzymes in controlling the balance between nucleotide salvage and degradation. Nucleosidases may also play an important role in signalling processes, since for eATP, an extracellular signalling molecule, to act efficiently, it must be removed from the apoplast after exerting its function, which is achieved by the concerted action of apyrases and nucleotidases and extracellular nucleosidases. These nucleosidases may also be involved in the recicling of nucleosides from the soil or those that are released from damaged cells. The main objective of this work has been the characterisation of the gene family of nucleosidases in the ureidic legume Phaseolus vulgaris. For this purpose, the genes encoding these enzymes in common bean have been cloned, which has allowed us to determine that this family is formed by at least four genes, two located in the cytosol (PvNSH1 and PvNSH2) and two located in the apoplast (PvNSH3 and PvNSH4). In addition, the ORFs of these genes have been overexpressed in heterologous systems, which has allowed the purification of the corresponding enzymes and their subsequent characterisation. PvNSH1 has a high affinity for purine and pyrimidine nucleosides, whereas PvNSH2 appears to be more specific for purines and, in particular, for xanthosine. Regarding the apoplastic nucleosidases, PvNSH4 is active at acidic pH and uses mainly inosine as substrate, while the activity of PvNSH3 could not be characterised. It should be noted that the phylogenetic analysis of apoplastic nucleosidases suggests that these enzymes have a relatively recent evolutionary origin, as they have only been identified in some groups of plants, but not in organisms belonging to other kingdoms. On the other hand, the physiological role of nucleosidases in common bean has been investigated. The results obtained show that PvNSH1 and PvNSH2 play a relevant role during germination and postgerminative development, which highlights the importance of nucleotide metabolism at this stage of plant development. Furthermore, the importance of these enzymes in common bean nodules has been demonstrated, in which their expression and activity are very high, although no relationship between their levels and those of ureides in either roots or nodules has been demonstrated. Regarding the nucleosidases with predicted apoplastic localisation, this cellular location has been demonstrated, and that PvNSH4 could be involved in the response to stress and, specifically, in those stresses mediated by the hormone methyl jasmonate. On the other hand, it has not been possible to gain information about the function of PvNSH3, as the levels of expression detected were very low and without significant changes in any of the situations studied

Automatic marbling prediction of sliced dry-cured ham using image segmentation, texture analysis and regression

Dry-cured ham is a traditional Mediterranean meat product consumed throughout the world. This product is very variable in terms of composition and quality. Consumer’s acceptability of this product is influenced by different factors, in particular, visual intramuscular fat and its distribution across the slice, also known as marbling. On-line marbling assessment is of great interest for the industry for classification purposes. However, until now this assessment has been traditionally carried out by panels of experts and this methodology cannot be implement in industry. We propose a complete automatic system to predict marbling degree of dry-cured ham slices, which combines: (1) the color texture features of regions of interest (ROIs) extracted automatically for each muscle; and (2) machine learning models to predict the marbling. For the ROIs extraction algorithm more than the 90% of pixels of the ROI fall into the true muscle. The proposed system achieves a correlation of 0.92 using the support vector regression and a set of color texture features including statistics of each channel of RGB color image and Haralick’s coefficients of its gray-level version. The mean absolute error was 0.46, which is lower than the standard desviation (0.5) of the marbling scores evaluated by experts. This high accuracy in the marbling prediction for sliced dry-cured ham would allow to deploy its application in the dry-cured ham industryThis work has received financial support from the Xunta de Galicia (Centro singular de investigación de Galicia, accreditation 2020– 2023) and the European Union (European Regional Development Fund–ERDF), Project ED431G-2019/04. IRTA’s contribution was also funded by the CCLabel project (RTI-2018- 096883-R-C41) and the CERCA programme from Generalitat de CatalunyaS

A highly sensitive method for the reassessment and quantification of 239Pu in urine samples based on a 1 MV accelerator mass spectrometry system

A new and highly sensitive method for the determination of 239Pu in human urine has been developed permitting the reassessment of planchets initially prepared for alpha spectrometry (AS) analysis in the context of internal dosimetry. A set of urine samples (volume: 500 mL) was spiked with known quantities of 239Pu, ranging from 2 to 120 fg (4.6 mBq–0.3 mBq), employing 14 pg (2.05 mBq) of 242Pu as internal standard. The Pu was purified by ion-chromatography using BioRad AG1X2 anion-exchange resins (Bio-Rad Laboratories Inc.). The chemical yield was determined by alphaspectrometry, being about 80%. Afterwards, the planchets so obtained were leached with diluted HNO3 and the dissolved plutonium was determined by Accelerator Mass Spectrometry (AMS) at the Centro Nacional de Aceleradores (CNA) in Seville, Spain. The minimum detectable activity (MDA) for the AMS measurements was determined through the study of a set of procedural blanks, giving figures of about 0.44 fg (1 mBq) per sample. This contrasts with the MDA obtained by AS for the same set of samples, of about 50 fg (0.1 mBq). The results now presented helps to demonstrate that the routine measurement of 239Pu at ultra trace levels in human urine samples is possible with the new-generation of compact AMS systems, offering a highly sensitive method for the reassessing of planchets prepared for bioassay purposes

Reassessment of 239Pu on planchets from human urine samples at ultra-trace levels using Aridus-ICP-SFMS and AMS

New analytical methods developed at the facilities here, based on two ultra-sensitive mass spectrometry (MS) techniques, inductively coupled plasma sector field mass spectrometer with a desolvator system (Aridus-ICP-SFMS) and accelerator MS (AMS), have been applied in this work for the reassessment of 239Pu in alpha spectrometry (AS) planchets corresponding to spiked human urine samples. The obtained 239Pu minimum detectable activities (MDAs) values by Aridus-ICP-SFMS and AMS were 3 fg (∼6.92 mBq) and 0.4 fg (∼0.92 mBq), respectively, per sample, which are much better than those attainable by AS [50 fg (∼115.3 mBq) of 239Pu per sample, approximately]. Therefore, it is demonstrated that the MS techniques employed in this work are very powerful tools for internal dosimetry studies in human urine samples, giving excellent results when the reassessment of AS planchets is needed (samples with a Pu concentration below or at the MDA levels measurable by AS)

Family CEO affect and R&D investments of family firms: The moderation effect of family ownership structure

The last decades have seen increasing interest in the determinants of heterogeneity in family firm innovation. In this study, we respond to recent calls to address the micro-level mechanisms behind innovation in family firms. Specifically, we analyze the effect of family CEO affect, namely positive and negative affective traits, on the R&D investment decisions of family firms. We also analyze the moderating effect of family ownership structure on the influence of CEO affect on these strategic decisions. Consistent with affect maintenance arguments, our findings of a sample of 142 Spanish family firms show that positive family CEO affect negatively influences the R&D investments of family firms, while negative affect positively influences these investments. The results also show that family CEO ownership and family CEO branch ownership strengthen the effects of family CEO affect on R&D investments, whereas ownership concentration in other family branches weakens these effects.This work was supported by grant from the Spanish Ministry of Science and Innovation (Ministerio de Ciencia e Innovacion) ´ PID2020-120288 GB-I00

Las exposiciones interactivas para escolares como herramienta de motivación de los estudiantes de Ingeniería Forestal y del Medio Natural

El perfil de alumnado de nuevo ingreso en el Grado en Ingeniería Forestal y del Medio Natural (IFMN) de la UEX es, con frecuencia, el de un titulado de ciclo formativo de grado superior con importantes carencias en las materias de tipo técnico, lo cual reduce su motivación para el estudio de éstas. Por otra parte, en el Grado en IFMN del Centro Universitario de Plasencia existe una preocupación por la innovación docente (llevada a cabo a través de varios proyectos competitivos) para lograr una mejor formación del alumnado tanto en competencias técnicas como en competencias transversales (entre las que se incluye la inteligencia emocional) y, paralelamente, mejorar las tasas de éxito y rendimiento de las materias implicadas. En este sentido, se ha diseñado una estrategia innovadora, bajo el lema general de 'FOREXPO: enseñando se aprende más', consistente en una exposición interactiva sobre Espacios Naturales Protegidos, Física y Medio Ambiente, que es ofrecida por los estudiantes universitarios de Ingeniería Forestal a los escolares de la ciudad de Plasencia, que visitan dicha exposición cada primavera, desde 2013, para aprender de manera directa y amena a través de maquetas y exposiciones de tipo interactivo donde pueden experimentar, observar, etc.La novedad de esta estrategia denominada FOREXPO es que combina diversas metodologías innovadoras como el ABP (Aprendizaje Basado en Problemas, ya que los estudiantes universitarios deben confeccionar las maquetas o exposiciones con un objetivo final), el ApS (puesto que en su aprendizaje realizan un servicio a la sociedad, como es mostrar ciencia cercana y divertida a los escolares de primaria de la ciudad de Plasencia), el “learning by teaching” o enseñando se aprende más (pues les empodera y motiva enormemente poder hablar a un público no especializado sobre cuestiones científicas de una cierta dificultad como la Física ambiental) y, por último, supone una colaboración entre dos materias del Grado, de forma que no se funcione de manera estanca.Esta experiencia innovadora es muy bien valorada por los estudiantes universitarios, tanto por la autoestima científica que les proporciona en materias difíciles como Física, como por las competencias transversales adquiridas al realizar las tareas organizativas que el ABP plantea como objetivo secundario. Asimismo, la acogida por parte de los centros educativos de primaria es muy buena en la ciudad, tanto por el contenido científico expuesto como por el buen hacer de los “monitores” universitarios y por la posibilidad de conocer la titulación en una especie de jornada de puertas abiertas que contribuye a difundir también la labor del Ingeniero Forestal en la sociedad, especialmente mediante lo expuesto en la materia de Gestión de Espacios Naturales Protegidos

S-Like Ribonuclease T2 Genes Are Induced during Mobilisation of Nutrients in Cotyledons from Common Bean

Germination and seedling development are crucial phases in a plant’s life cycle with economical and agronomical implications. The RNA quality in seeds is linked to seed viability, being an important agronomic trait since this leads to a loss in germination efficiency. In addition, RNA can be an important phosphorous reservoir in seeds, affecting the efficiency of the mobilisation of nutrients towards the seedlings. However, knowledge about the physiological function of ribonucleases during germination and seedling development is scarce. We analysed the ribonuclease activities of cotyledons during these processes and the expression of S-like ribonucleases T2. Ribonuclease activity was detected in cotyledons at 1 day after imbibition and the specific activity increased during germination and seedling development, reaching a maximal value at 10 days after imbibition. At this stage, the levels of proteins and RNA in cotyledons were very low. Using in-gel assays, three ribonucleases were detected with apparent molecular masses of 16, 17 and 19 kDa along cotyledon ontogeny. The S-like ribonucleases T2 family consists of four genes in common bean (PvRNS1 to PvRNS4). The expression of PvRNS1, PvRNS2 and PvRNS4 increased in the phase of nutrient mobilisation in cotyledons. The expression of PvRNS1 increased 1000 fold in cotyledons, from 1 to 6 days after imbibition. The suppression of the induction of ribonuclease activity and gene expression in decapitated seedlings suggests that the regulatory signal comes from the developing axes. These results clearly state that S-like ribonucleases T2 are involved in RNA turnover in cotyledons during seedling development

Brain Structural Correlates of Reward Sensitivity and Impulsivity in Adolescents with Normal and Excess Weight

Introduction: Neuroscience evidence suggests that adolescent obesity is linked to brain dysfunctions associated with enhanced reward and somatosensory processing and reduced impulse control during food processing. Comparatively less is known about the role of more stable brain structural measures and their link to personality traits and neuropsychological factors on the presentation of adolescent obesity. Here we aimed to investigate regional brain anatomy in adolescents with excess weight vs. lean controls. We also aimed to contrast the associations between brain structure and personality and cognitive measures in both groups. Methods: Fifty-two adolescents (16 with normal weight and 36 with excess weight) were scanned using magnetic resonance imaging and completed the Sensitivity to Punishment and Sensitivity to Reward Questionnaire (SPSRQ), the UPPS-P scale, and the Stroop task. Voxel-based morphometry (VBM) was used to assess possible between-group differences in regional gray matter (GM) and to measure the putative differences in the way reward and punishment sensitivity, impulsivity and inhibitory control relate to regional GM volumes, which were analyzed using both region of interest (ROI) and whole brain analyses. The ROIs included areas involved in reward/somatosensory processing (striatum, somatosensory cortices) and motivation/impulse control (hippocampus, prefrontal cortex). Results: Excess weight adolescents showed increased GM volume in the right hippocampus. Voxel-wise volumes of the second somatosensory cortex (SII) were correlated with reward sensitivity and positive urgency in lean controls, but this association was missed in excess weight adolescents. Moreover, Stroop performance correlated with dorsolateral prefrontal cortex volumes in controls but not in excess weight adolescents. Conclusion: Adolescents with excess weight have structural abnormalities in brain regions associated with somatosensory processing and motivation

The stellar content of giant H II regions in NGC 7714

This is an electronic version of an article published in The Astrophysical Journal. García-Vargas, M.L., González-Delgado, R.M., Pérez, E., Alloin, D., Díaz, A. and E. Terlevich. The stellar content of giant H II regions in NGC 7714. The Astrophysical Journal 478 (1997): 112-12

Protocolo del estudio: Demanda y práctica farmacéutica en afección bucofaríngea en España. Estudio ACTUA

Introducción y justificación: La afección bucofaríngea, y más concretamente el dolor de garganta, por su prevalencia y relación con el uso inadecuado de medicamentos tiene una alta importancia en salud, el farmacéutico contribuye a que el usuario alcance una automedicación adecuada a través de los servicios de atención farmacéutica. Sin embargo, se conoce poco en el ámbito de la farmacia comunitaria sobre los usuarios que demandan esta atención, así como las consultas e intervenciones de los farmacéuticos.Aplicabilidad de los resultados: Conocer las características de la demanda y la práctica farmacéutica en afección bucofaríngea permitirá establecer estrategias sanitarias destinadas a optimizar la asistencia sanitaria.Objetivos: Caracterizar la práctica farmacéutica en afección bucofaríngea realizada en farmacias comunitarias españolas.Material y métodos: Estudio observacional descriptivo transversal. En farmacias comunitarias voluntarias del territorio español. La población de estudio serán los usuarios que acudan a las farmacias por una afección bucofaríngea. La duración del trabajo de campo será de tres meses. Las variables contempladas en el estudio serán aquellas que caracterizan al usuario, a la consulta realizada, y a la intervención del farmacéutico. Se realizará un análisis estadístico descriptivo de los datos (univariante y multivariante por la técnica de correspondencias múltiples). Se garantizará la confidencialidad y el consentimiento informado de los participantes