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ln this work, we attempted to determine the function of SZRDl,a protein of unknown function using a multidimensional approach. Our selection of SZRDl was based on the presence of four putative binding sites for miR-128 in its 3'UTR.Mi-RNAs are small RNAs that inhibit translation and promote mRNA degradation of target transcripts. They usually have many targets, but effects on individual targets are often modest. Small changes in target transcripts are therefore thought to collaborate to lead to a cellular function. We therefore hypothesized that uncharacterized targets of a mi-RNA might functionally collaborate with other targets of the same mi-RNA. To test this hypothesis we focused on miR-128 and one of its predicted targets of unknown function, SZRDl. MiR-128 targets are implicated in cellular signaling pathways and in a RNA quality control mechanism called Nonsense-Mediated Decay (NMD). we hypothesized that SZRDl could be implicated in the function’s exerted by miR-128.First, we demonstrated that the conserved multiple binding site of miR-128 in the 3'UTR of SZRDl are indeed functional using luciferase reporter genes.The analysis of SZRDl protein function followed a multidimensional approach based on primary sequence analysis, existing high-throughput data and identification of possible partners through tandem affinity purification followed by mass spectrometry:The SZRDl gene gives rise to several transcripts that code for proteins of 133 and 154 amino acids (SZRDl Long and SZRDl Short). We showed that SZRDl Long is excluded from the nuclei, possibly through a predicted nuclear export sequence (NES), and that SZRDl Short is phosphorylated in a cell cycle-dependent manner.We identified several proteins implicated in various aspects of RNA metabolism as interactors of SZRDl: RENTl/UPFl,STRAP, ClQBP, UNR/CSDEl and GEMIN2.We provide preliminary evidence that SZRDl might regulate the function of its interactors RENTl/UPFl and STRAP.Notably, RENTl/UPFl has previously been identified as a target of miR-128. Our data therefore are consistent with the hypothesisthat target genes of miR-128 functionally collaborate. Dans le cadre de ce mémoire, nous avons cherché à établir la fonction de SZRDl en utilisant une approche multidimensionelle. C'est la présence de quatre sites de liaisons potentiels au miR-128 dans la région 3'non traduite (3'UTR) de SZRDl qui a guidé notre choix vers cette protéine.Les mi-ARNs sont de courts ARNs simple brin qui inhibent la traduction et/ou favorisent la dégradation de leurs transcrits cibles. Dans la plupart des cas, chaque mi-ARN possède un grand nombre de cibles mais l'effet sur chacune d'entre elles est souvent modeste. Ilest donc possible que l'association de ces faibles mais nombreux changements ait une importance fonctionnelle. Nous avons donc émis l'hypothèse qu'une cible non caractérisée d'un mi-ARN pourraient collaborer avec d'autres cibles de ce même mi-ARN. Nous avons testé cette hypothèse en étudiant le miR-128 et une de ses cibles prédites mais dont la fonction est inconnue, SZRDl. Les cibles connues du miR-128 jouent un rôle dans la signalisation cellulaire et dans un mécanisme de contrôle de la qualité des ARNs appelé « Nonsense-Mediated Decay » ou NMD. Dès lors, nous avons émis l'hypothèse selon laquelle la protéine SZRDl pourrait être impliquée dans les fonctions connues de miR-128.Nous avons tout d'abord démontré, grâce à un test rapporteur luciférase, que les sites de liaisons conservés pour le miR-128 présents dans la région 3'UTR de SZRDl sont fonctionnels et que SZRDl semble bien être une cible du miR-128. Afin de déterminer la fonction de SZRDl,nous avons utilisé une approche multidimensionnelle basée sur l'analyse de la séquence primaire en résidus aminés de la protéine, sur des données issues d'analyses à haut débit et sur l'identification de partenaires potentiels par purification d'affinité en tandem suivie de spectrométrie de masse :Le gène SZRDl donne lieu à plusieurs transcrits codants pour des protéines de 133 et 154 résidus aminés (SZRDl Courte et SZRDl Longue, respectivement) . Nous avons montré que SZRDl Longue peut être exclue du noyau, probablement dû à la présence d'une séquence d'export nucléaire (NES) et que SZRDl Courte est phosphorylée à certains stades du cycle cellulaire.Nous avons identifié plusieurs protéines interactantes de SZRDl qui sont impliquées à différents niveaux du métabolisme de I'ARN : RENTl/UPFl, STRAP, ClQBP, UNR/CSDl et GEMIN2.Nous avons obtenus des données préliminaires suggérant que SZRDl est capable de réguler,la fonction de certaines de ses protéines partenaires comme RENTl/UPFl et STRAP.Il est important de noter que RENTl/UPFl a également été décrit comme étant une cible du miR- 1Nos données sont donc en accord avec l'hypothèse selon laquelle les gènes cibles du miR- 128 collaborent fonctionnellement.

MIRN128 microRNA, human --- RNA Processing, Post-Transcriptional

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Crithidia --- DNA, Mitochondrial --- RNA --- RNA Processing, Post-Transcriptional --- Gene Expression Regulation --- Transcription, Genetic --- RNA --- genetics --- chemistry --- chemistry --- genetics

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The presence of modified nucleotides in cellular RNAs has been known for decades and over 100 distinct RNA modifications have been characterized to date. While the exact role of many of these modifications is still unclear, many are highly conserved across evolution and most contribute to the overall fitness of the organism. In recent years, new methods and bioinformatics approaches have been developed for the dissection of modification pathways and functions. These methods intersect a number of related fields, ranging from RNA processing to comparative genomics and systems biology. In add

RNA editing. --- ARN --- Edition --- RNA Editing --- RNA Processing, Post-Transcriptional --- Metabolism --- Biochemical Processes --- Gene Expression Regulation --- Biochemical Phenomena --- Metabolic Phenomena --- Genetic Processes --- Chemical Processes --- Chemical Phenomena --- Genetic Phenomena --- Phenomena and Processes --- Editing, RNA --- Messenger RNA editing --- mRNA editing --- Genetic regulation

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RNA Modification provides a useful examination of the science and its role in biological regulation, the current frontier of life science research, and includes various RNA modications and their role in gene expression. It represents the most up-to-date knowledge and protocols available today.

• Dynamic RNA modifications and their roles in biological regulation are the current frontier of life science research
• This volume of Methods in Enzymology represents up to date knowledge and protocols

RNA Processing, Post-Transcriptional --- Pseudouridine --- RNA, Transfer --- Gene Expression Regulation --- Uridine --- Biochemical Processes --- RNA --- Metabolism --- Genetic Processes --- Ribonucleosides --- Pyrimidine Nucleosides --- Biochemical Phenomena --- Chemical Processes --- Nucleic Acids --- Metabolic Phenomena --- Nucleosides --- Genetic Phenomena --- Nucleic Acids, Nucleotides, and Nucleosides --- Phenomena and Processes --- Chemical Phenomena --- Pyrimidines --- Chemicals and Drugs --- Heterocyclic Compounds, 1-Ring --- Heterocyclic Compounds --- Animal Biochemistry --- Human Anatomy & Physiology --- Health & Biological Sciences --- RNA editing. --- ARN --- pharmacokinetics --- Edition --- Editing, RNA --- Messenger RNA editing --- mRNA editing --- RNA Processing, Post-Transcriptional. --- pharmacokinetics. --- Genetic regulation

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In Post-Transcriptional Gene Regulation, renowned authors present current technical approaches to most aspects of post-transcriptional control and provide a useful and versatile laboratory bench resource. With chapters split into sections covering bioinformatics, fundamental aspects of the study of RNA biology, and techniques for specific aspects of RNA biology, the expert authors have filled the book with invaluable tricks of the trade, perfected in their state-of-the-art laboratories. This new volume from the Methods in Molecular Biology series is convinently divided into three sections. The first section presents a series of bioinformatic approaches to address the use of RNA databases and algorithms to the study of post-transcriptional regulation involving untranslated regions of transcripts. In the second section, a series of methods applicable to fundamental issues in mRNA biology are presented. These include RNA structure/function, mRNP analysis and novel methods for mRNA labeling and isolation. The third section of this volume presents methodologies to study particular aspects of post-transcriptional control. This section includes methods for the study of alternative splicing and 3’ end processing, mRNA localization, mRNA translation, mRNA stability and si/miRNA regulation. Collectively, Post-Transcriptional Gene Regulation provides the reader with a useful and versatile laboratory bench resource that will become an essential reference in the field.

Genetic regulation --- Messenger RNA --- Cellular control mechanisms --- Bioinformatics --- Gene Expression Regulation --- RNA Processing, Post-Transcriptional --- Computational Biology --- Bioinformatics. --- Régulation génétique --- ARN messager --- Régulation cellulaire --- Bio-informatique --- Laboratory manuals --- Manuels de laboratoire --- Informational RNA --- Messenger ribonucleic acid --- mRNA --- Protein transcript --- Protein transcripts --- Template RNA --- Bio-informatics --- Biological informatics --- Biology --- Information science --- Computational biology --- Systems biology --- Data processing --- Cell regulation --- Biological control systems --- Cell metabolism --- RNA --- Gene expression --- Gene expression regulation --- Gene regulation --- Biosynthesis --- Molecular genetics --- Regulation --- Biochemistry. --- Cytology. --- Human genetics. --- Biochemistry, general. --- Cell Biology. --- Human Genetics. --- Cell biology --- Cellular biology --- Cells --- Cytologists --- Genetics --- Heredity, Human --- Human biology --- Physical anthropology --- Biological chemistry --- Chemical composition of organisms --- Organisms --- Physiological chemistry --- Chemistry --- Medical sciences --- Composition --- Genetic regulation - Laboratory manuals --- Messenger RNA - Laboratory manuals --- Cellular control mechanisms - Laboratory manuals --- Gene Expression Regulation - laboratory manuals --- RNA Processing, Post-Transcriptional - Laboratory Manuals