Funktion und Biogenese kleiner RNAs in Dictyostelium discoideum

RNA mediated gene silencing pathways are highly conserved among eukaryotes and they have been well investigated in animals and in plants. Longer dsRNA molecules trigger the silencing pathways: RNase III proteins and their dsRNA binding protein (dsRBP) partners recognize those molecules as a substrate and process 21 nucleotide long microRNAs (miRNAs) or small interfering RNAs (siRNAs). Some organisms encode RNA dependent RNA polymerases (RdRPs), which are able to expand the pool of existing siRNAs. Argonaute proteins are able to bind small regulatory RNAs and are subsequently recruited to target mRNAs by base complementary. This leads in turn to transcriptional or posttranscriptional silencing of respective genes. The Dictyostelium discoideum genome encodes two Dicer homologues (DrnA and DrnB), five Argonaute proteins (AgnA to AgnE) and three RdRPs (RrpA to RrpC). In addition, the amoeba is known to express miRNAs and siRNAs, while the latter derive mainly from the DIRS-1 retrotransposon. One part of this work focused on the miRNA biogenesis pathway of D. discoideum. It was shown that the dsRNA binding protein RbdB is a necessary component for miRNA processing in the amoeba. There were no mature miRNAs detectable by Northern blot analysis in rbdB- strains, which is also true for drnB mutants. Moreover, primary miRNA-transcripts (pri-miRNAs) accumulated in rbdB- and drnB- strains. Fluorescence microscopy studies showed a nuclear localization of RbdB. RbdB accumulated in distinct perinucleolar foci. These were reminiscent of plant dicing bodies that contain essential protein components for miRNA processing. It is well known that RNase III enzymes and dsRBPs work together during miRNA processing in higher eukaryotes. This work demonstrated that the same is true for members of the amoebozoa supergroup. In Arabidopsis the nuclear zinc finger protein Serrate (SE) is also necessary for miRNA processing. The D. discoideum homologue SrtA, however, is not relevant which has been shown by the analysis of the respective knockdown strain. MiRNAs are known to be differentially expressed in several RNAi knockout strains. The accumulation of miRNAs in agnA- strains and a strong decrease in rbdB- strains were criteria that could thus be successfully used (among others) to identify and validate new miRNAs candidates by Illumina®-RNA sequencing. In another part of this study, the silencing and amplification of the DIRS-1 retrotransposons was analyzed in more detail. It was already known that DIRS-1 transcripts and extrachromosomal DIRS-1 DNA molecules accumulated in agnA- strains. This phenotype was correlated with the loss of endogenous DIRS-1 siRNAs in the knockout strain. By deep sequencing analysis of small RNAs from the AX2 wild type and the agnA- strain, the strong decrease of endogenous DIRS-1 siRNAs in the mutant strain (accounting for 70 %) could be confirmed. Further analysis of the data revealed an unequal distribution of DIRS-1 derived siRNAs along the retroelement in the wild type strain, since only very few of them matched the inverted terminal repeats (ITRs) and the 5’- half of the first open reading frame (ORF). Besides, sense and antisense siRNAs were asymmetrically distributed, as well. By using different reporter constructs it was shown indirectly that AgnA is necessary for the RrpC mediated production of secondary DIRS-1 siRNAs. These analyses also demonstrated an amplification of siRNAs in 5’- and in 3’-direction. Further analysis of the agnA- strain revealed that not only DIRS-1 sense transcripts but also ORF2 and ORF3 encoded proteins were enriched. In contrast, the ORF1 encoded protein GAG was equally expressed in the mutant and the wild type. This might reflect the unequal distribution of endogenous DIRS-1 siRNAs along the retrotransposon. Southern Blot and PCR-analyses showed that extrachromosomal DIRS-1 DNA molecules are present in the cytoplasm of angA- strains and that they are complementary to sense transcripts of intact DIRS-1 elements. Thus, the extrachromosomal DIRS-1 intermediates are likely incomplete cDNA molecules generated by the DIRS-1 encoded reverse transcriptase. One could hypothesize that virus like particles (VLPs) are the places of DIRS-1 cDNA synthesis. At least, DIRS-1 GAG proteins interact and fluorescence microscopy studies showed that they localize in distinct cytoplasmic foci which accumulate in close proximity to the nuclei.

Introduction to Knowledge Representation

Focussing on Open Data and the need for cleaning data

Material didàctic per a l'àrea d'anglès a un centre trilingüe de secundària : com treballar lectures a classe. 'Material didáctico para el área de inglés en un centro trilingüe de secundaria : cómo trabajar lecturas en clase'.

Resumen tomado de la publicación

The narrative of arguing the personalised news

Bay 9 are hoping to pioneer a way to encourage postgrads and staff in the lab to get over the fear of presenting their work to the group. The members of the bay will each give a 6m40s Pecha Kucha explaining their current research work through pictures. The topics of the pecha kuchas are: - Citizen Participation in News: An analysis of the landscape of online journalism (Jonny) - Argumentation on the Social Web (Tom) - From Narrative Systems to Ubiquitous Computing for Psychology - and everything in between (Charlie) - Is it worth sharing user model data? (Rikki)

Com raspallar-se les dents : el fil dental. 'Como cepillarse los dientes : el hilo dental'.

Resumen del vídeo en catalán

Los hábitos saludables.

Se trata de una unidad didáctica sobre la importancia de los valores alimentarios, y de la importancia de los mismos en la sociedad de hoy, a través de la historia de Charlie y la fabrica de chocolate se pretende concienciar a los niños de la importancia de una alimentación saludable y potenciar otros valores como la generosidad, la amistad y la familia.

Tocar, oír, probar ... ­Juguemos con los sentidos!

El proyecto propone descubrir las posibilidades de todos los sentidos a través de un taller de experiencias en el que participan, en principio, niños de 0 a 2 años que se amplia a los niños de 2 a 3 y de 3 a 4. El taller sirve para trabajar los distintos objetivos y contenidos relacionados con el proyecto educativo y el proyecto curricular: el cuerpo y la propia imagen, juego y movimiento, los objetos, y lenguaje oral. Al inicio del curso se prepararon materiales, y se revisaron y adaptaron los contenidos teniendo en cuenta los resultados de la evaluación inicial realizada. A partir de enero se iniciaron las sesiones de trabajo de media hora con cada grupo y nivel durante una semana. En cada una de ellas se seguían los siguientes pasos: presentación de los objetos o materiales que se iban a utilizar; experimentación y expresión de sensaciones e ideas (manipulación de materiales, observación); puesta en común; y recogida de material. Los materiales utilizados han sido muy variados y en algunos casos los niños también han participado en su elaboración: molinillos de viento, tablillas de distintas texturas (lija, raso, arpillera), soportes de madera con tiras de papel seda, cajas decoradas para guardar materiales, frasquitos llenos de agua tintada, etc. La evaluación continua prevista en el proyecto ha permitido modificar puntualmente algunos aspectos.

Qué bonito es mi huerto.

Experiencia que consiste en la creación de un huerto escolar para el alumnado de tres y cuatro años. Los objetivos son, entre otros: desarrollar la coordinación visomanual y las habilidades manipulativas necesarias para manejar y explorar objetos con mayor precisión; aceptar pequeñas frustraciones y manifestar una actitud positiva ante las dificultades. Se desarrolla en tres fases: recopilación de materiales y útiles, preparación de la tierra y las semillas y mantenimiento y recolección de los frutos. En la valoración se destaca el logro de experiencias gratificantes para los niños y la creación de un espacio de juego-trabajo..

Superstars.

El volumen tiene dos historias que tratan situaciones reales que pueden tener los niños. En la primera, Andy está una semana con mamá y otra semana con papá, ya que están divorciados. En 'El proyecto Lottie' Charlie encuentra su doble, cuya vida es mucho más dura que la suya. Para niños que estan pasando por una experiencia similar. Los libros están redactados en primera persona para hacer las historias más reales. Contiene : the suitcase kid ; the lottie project. .

The town mouse and the country mouse.

Es una nueva versión de la fábula de Esopo en la que se narra la historia de dos ratones, Charlie, que vive en el campo y Tyler, que vive en la ciudad. Cuando se visitan mutuamente, cada uno se sorprende de la forma de vivir que lleva el otro, pero, finalmente, a cada uno le gusta el lugar en el que vive.

Edexcel A2 physics revision guide : for SHAP and concept-led approaches.

Guía para la revisión de los recursos Edexcel para la especificación General Certificate of Education (GCE ) Física. Salters-Horners Advanced Physics (SHAP) es el material de apoyo oficial. Es un curso completo que cubre la física a través de los contextos de la vida real, que combina los conceptos clave que sustenta la física, con la oportunidad de adquirir las habilidades que los físicos necesitan hoy. Favorece el aprendizaje autónomo con una amplia gama de conocimientos, incluidos el análisis de datos, evaluación crítica de información, comunicación y trabajo cooperativo. Se basa en datos de exámenes reales para ayudar a los estudiantes a evitar errores habituales y construir mejores respuestas. Los ejemplos de preguntas de examen incluyen las de opción múltiple. Tiene las soluciones a las preguntas del texto.

Mutational activation of niche-specific genes provides insight into regulatory networks and bacterial function in a complex environment

The genome of the plant-colonizing bacterium Pseudomonas fluorescens SBW25 harbors a subset of genes that are expressed specifically on plant surfaces. The function of these genes is central to the ecological success of SBW25, but their study poses significant challenges because no phenotype is discernable in vitro. Here, we describe a genetic strategy with general utility that combines suppressor analysis with IVET (SPyVET) and provides a means of identifying regulators of niche-specific genes. Central to this strategy are strains carrying operon fusions between plant environment-induced loci (EIL) and promoterless 'dapB. These strains are prototrophic in the plant environment but auxotrophic on laboratory minimal medium. Regulatory elements were identified by transposon mutagenesis and selection for prototrophs on minimal medium. Approximately 106 mutants were screened for each of 27 strains carrying 'dapB fusions to plant EIL and the insertion point for the transposon determined in approximately 2,000 putative regulator mutants. Regulators were functionally characterized and used to provide insight into EIL phenotypes. For one strain carrying a fusion to the cellulose-encoding wss operon, five different regulators were identified including a diguanylate cyclase, the flagella activator, FleQ, and alginate activator, AmrZ (AlgZ). Further rounds of suppressor analysis, possible by virtue of the SPyVET strategy, revealed an additional two regulators including the activator AlgR, and allowed the regulatory connections to be determined.

Thioquinolobactin, a pseudomonas siderophore with antifungal and anti-pythium activity

Under conditions of iron limitation Pseudomonas fluorescens ATCC 17400 produces two siderophores, pyoverdine, and a second siderophore quinolobactin, which itself results from the hydrolysis of the unstable molecule 8-hydroxy-4-methoxy-2-quinoline thiocarboxylic acid (thioquinolobactin). Pseudomonas fluorescens ATCC 17400 also displays a strong in vitro antagonism against the Oomycete Pythium, which is repressed by iron, suggesting the involvement of a siderophore(s). While a pyoverdine-negative mutant retains most of its antagonism, a thioquinolobactin-negative mutant only slowed-down Pythium growth, and a double pyoverdine-, thioquinolobactin-negative mutant, which does not produce any siderophore, totally lost its antagonism against Pythium. The siderophore thioquinolobactin could be purified and identified from spent medium and showed anti-Pythium activity, but it was quickly hydrolysed to quinolobactin, which we showed has no antimicrobial activity. Analysis of antagonism-affected transposon mutants revealed that genes involved in haem biosynthesis and sulfur assimilation are important for the production of thioquinolobactin and the expression of antagonism.

The evolution of mobile DNAs: when will transposons create phylogenies that look as if there is a master gene?

Some families of mammalian interspersed repetitive DNA, such as the Alu SINE sequence, appear to have evolved by the serial replacement of one active sequence with another, consistent with there being a single source of transposition: the "master gene." Alternative models, in which multiple source sequences are simultaneously active, have been called "transposon models." Transposon models differ in the proportion of elements that are active and in whether inactivation occurs at the moment of transposition or later. Here we examine the predictions of various types of transposon model regarding the patterns of sequence variation expected at an equilibrium between transposition, inactivation, and deletion. Under the master gene model, all bifurcations in the true tree of elements occur in a single lineage. We show that this property will also hold approximately for transposon models in which most elements are inactive and where at least some of the inactivation events occur after transposition. Such tree shapes are therefore not conclusive evidence for a single source of transposition.

Evidence-based psychological therapies: from bench to bedside

Patients with mental health difficulties do not always receive appropriate and recommended psychological treatment for their difficulties, and clinicians are not always appropriately trained to deliver them. This paper considers why this might be the case and provides an overview of the Charlie Waller Institute, a not-for-profit organisation funded by the NHS, University of Reading, and the Charlie Waller Memorial Trust. The Institute seeks to address this problem by training clinicians in wide variety of evidence-based therapies and assessing the impact of this training on clinician knowledge and skill.