RNA Pull-down Procedure to Identify RNA Targets of a Long Non-coding RNA

International audienceLong non-coding RNA (lncRNA), which are sequences of more than 200 nucleotides without a defined reading frame, belong to the regulatory non-coding RNA's family. Although their biological functions remain largely unknown, the number of these lncRNAs has steadily increased and it is now estimated that humans may have more than 10,000 such transcripts. Some of these are known to be involved in important regulatory pathways of gene expression which take place at the transcriptional level, but also at different steps of RNA co-and post-transcriptional maturation. In the latter cases, RNAs that are targeted by the lncRNA have to be identified. That's the reason why it is useful to develop a method enabling the identification of RNAs associated directly or indirectly with a lncRNA of interest. This protocol, which was inspired by previously published protocols allowing the isolation of a lncRNA together with its associated chromatin sequences, was adapted to permit the isolation of associated RNAs. We determined that two steps are critical for the efficiency of this protocol. The first is the design of specific anti-sense DNA oligonucleotide probes able to hybridize to the lncRNA of interest. To this end, the lncRNA secondary structure was predicted by bioinformatics and anti-sense oligonucleotide probes were designed with a strong affinity for regions that display a low probability of internal base pairing. The second crucial step of the procedure relies on the fixative conditions of the tissue or cultured cells that have to preserve the network between all molecular partners. Coupled with high throughput RNA sequencing, this RNA pull-down protocol can provide the whole RNA interactome of a lncRNA of interest

Discutindo a educação ambiental no cotidiano escolar: desenvolvimento de projetos na escola formação inicial e continuada de professores

A presente pesquisa buscou discutir como a Educação Ambiental (EA) vem sendo trabalhada, no Ensino Fundamental e como os docentes desta escola compreendem e vem inserindo a EA no cotidiano escolar., em uma escola estadual do município de Tangará da Serra/MT, Brasil. Para tanto, realizou-se entrevistas com os professores que fazem parte de um projeto interdisciplinar de EA na escola pesquisada. Verificou-se que o projeto da escola não vem conseguindo alcançar os objetivos propostos por: desconhecimento do mesmo, pelos professores; formação deficiente dos professores, não entendimento da EA como processo de ensino-aprendizagem, falta de recursos didáticos, planejamento inadequado das atividades. A partir dessa constatação, procurou-se debater a impossibilidade de tratar do tema fora do trabalho interdisciplinar, bem como, e principalmente, a importância de um estudo mais aprofundado de EA, vinculando teoria e prática, tanto na formação docente, como em projetos escolares, a fim de fugir do tradicional vínculo “EA e ecologia, lixo e horta”.Facultad de Humanidades y Ciencias de la Educació

Evidence for the presence of noradrenergic neurons and their inhibitory action on luteinizing hormone-releasing hormone release in cultured fetal rat hypothalamic cells

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Convergent serotonin and GABA innervation of VIP neurons in the suprachiasmatic nucleus demonstrated by triple labeling in the rat

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Evidence for GABA control of serotonin metabolism in the rat suprachiasmatic area

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Regulation by estradiol of GABAA and GABAB binding sites in the diencephalon of the rat: an autoradiographic study

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Light‐Induced Variations in AP‐1 Binding Activity and Composition in the Rat Suprachiasmatic Nucleus

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8-OH-DPAT regulates the amplitude and the phase of LH surge in ovariectomized steroid-primed rats

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Long‐term variations of AP‐1 composition after CRH stimulation: consequence on POMC gene regulation

International audienceIt has been shown previously that the CRH‐induced POMC gene transcription in the corticotroph cell line AtT‐20 involves an increase in AP‐1 DNA binding activity that remained elevated for at least 24 h, while induction of c‐fos was transient. We showed here that there were dramatic changes in protein components of AP‐1 including an initial recruitment of the transcriptional activators c‐Fos and Jun‐B then of Fra‐2 and Jun‐D. Changes in AP‐1 composition were concomitant with a decrease in POMC mRNA. Moreover, the presence of Fra‐2/Jun‐D dimers suppressed the CRH‐induction of c‐fos mRNA expression as well as c‐Fos/Jun‐B recruitment in AP‐1 complexes, suggesting the existence of autoregulatory loops of AP‐1 composition that involve complex interactions between the different members of the Jun and Fos families. It is concluded that CRH stimulation of corticotroph cells involves successive recruitment of activators and repressors, possibly contributing to prevent over expression of POMC