The CRISPR/Cas technology is enabling targeted genome editing in multiple organisms with unprecedented accuracy and specificity by using RNA-guided nucleases. A critical point when planning a CRISPR/Cas experiment is the design of the guide RNA (gRNA), which directs the nuclease and associated machinery to the desired genomic location. This gRNA has to fulfil the requirements of the nuclease and lack homology with other genome sites that could lead to off-target effects. Here we introduce the Breaking-Cas system for the design of gRNAs for CRISPR/Cas experiments, including those based in the Cas9 nuclease as well as others recently introduced. The server has unique features not available in other tools, including the possibility of using all eukaryotic genomes available in ENSEMBL (currently around 700), placing variable PAM sequences at 5′ or 3′ and setting the guide RNA length and the scores per nucleotides. It can be freely accessed at: http://bioinfogp.cnb.csic.es/tools/breakingcas, and the code is available upon request.

No abstract

Drug repositioning, using known drugs for treating conditions different from those the drug was originally designed to treat, is an important drug discovery tool that allows for a faster and cheaper development process by using drugs that are already approved or in an advanced trial stage for another purpose. This is especially relevant for orphan diseases because they affect too few people to make drug research de novo economically viable. In this paper we present NFFinder, a bioinformatics tool for identifying potential useful drugs in the context of orphan diseases. NFFinder uses transcriptomic data to find relationships between drugs, diseases and a phenotype of interest, as well as identifying experts having published on that domain. The application shows in a dashboard a series of graphics and tables designed to help researchers formulate repositioning hypotheses and identify potential biological relationships between drugs and diseases. NFFinder is freely available at http://nffinder.cnb.csic.es.

Our data identify novel protective mechanisms involved in RSV tolerance operating at system level in a gut microbe. These insights could influence the way RSV is used for a better management of gut microbial ecosystems to obtain associated health benefits.

During ontogeny, fetal liver (FL) acts as a major site for hematopoietic stem cell (HSC) maturation and expansion, whereas HSCs in the adult bone marrow (ABM) are largely quiescent. HSCs in the FL possess faster repopulation capacity as compared with ABM HSCs. However, the molecular mechanism regulating the greater self-renewal potential of FL HSCs has not yet extensively been assessed. Recently, we published RNA sequencing-based gene expression analysis on FL HSCs from 14.5-day mouse embryo (E14.5) in comparison to the ABM HSCs. We reanalyzed these data to identify key transcriptional regulators that play important roles in the expansion of HSCs during development. The comparison of FL E14.5 with ABM HSCs identified more than 1,400 differentially expressed genes. More than 200 genes were shortlisted based on the gene ontology (GO) annotation term "transcription." By morpholino-based knockdown studies in zebrafish, we assessed the function of 18 of these regulators, previously not associated with HSC proliferation. Our studies identified a previously unknown role for tdg, uhrf1, uchl5, and ncoa1 in the emergence of definitive hematopoiesis in zebrafish.In conclusion, we demonstrate that identification of genes involved in transcriptional regulation differentially expressed between expanding FL HSCs and quiescent ABM HSCs, uncovers novel regulators of HSC function.

We have used cryo Electron Tomography, proteomics and immunolabeling to study centrosomes isolated from the young lamb thymus, an efficient source of quiescent differentiated cells. We compared the proteome of thymocyte centrosomes to data published for KE37 cells, focusing on proteins associated with centriole disengagement and centrosome separation. The data obtained enhances our understanding of the protein system joining the centrioles, a system comprised of a branched network of fibers linked to an apparently amorphous density that was partially characterized here. A number of proteins were localized to the amorphous density by immunolabeling (C-NAP1, cohesin SMC1, condensin SMC4 and NCAPD2), yet not DNA. In conjuction, these data not only extend our understanding of centrosomes but they will help refine the model that focus on the protein system associated with the centriolar junction.

No abstract

Considering that time is an important dimension in anthropological discourse and a key element of the "hidden curriculum" of school, this paper presents the results of an ethnography on time in a high school in João Pessoa, Paraíba, Brazil. We discuss how teaching/learning processes are organized and experienced in time, paying close attention to the limitations and possibilities revealed by interactions between teachers and students. We first identify the pedagogical temporal units that compose the dimension of structural time at the school, and then identify aspects of time in practice (experiences and strategies). We argue that school time is both a constraint and an opportunity, unfolding in a series of metaphors ("save time," "waste time," "kill time", "fill time", "pass the time"), which enable ways of thinking about the conflicts present in daily school life. ResumoConsiderando que o tempo é uma importante dimensão para a reflexão antropológica, e um elemento fundamental no "currículo oculto" da ins- Mónica Franch and Josilene P. de Souza aprendizagem são organizados e vivenciados temporalmente, dando atenção aos limites e possibilidades que se revelam nas interações entre professores e alunos. Para isso, identificamos, em primeiro lugar, as unidades temporais pedagógicas, que compõem a dimensão do tempo estrutural da escola pesquisada para em seguida identificar aspectos do tempo na prática (experiên-cias e estratégias). Defendemos que o tempo escolar emerge simultaneamente como constrangimento e como oportunidade, se desdobrando em uma série de metáforas ("ganhar tempo", "perder tempo", "matar o tempo", "preencher o tempo", "passar o tempo"), que permitem pensar alguns impasses presentes no cotidiano escolar.