Redirecting splicing with bifunctional oligonucleotides

Abstract: Ectopic modulators of alternative splicing are important tools to study the function of splice variants and for correcting mis-splicing events that cause human diseases. Such modulators can be bifunctional oligonucleotides made of an antisense portion that determines target specificity, and a nonhybridizing tail that recruits proteins or RNA/protein complexes that affect splice site selection (TOSS and TOES, respectively, for targeted oligonucleotide silencer of splicing and targeted oligonucleotide enhancer of splicing). The use of TOSS and TOES has been restricted to a handful of targets. To generalize the applicability and demonstrate the robustness of TOSS, we have tested this approach on more than 50 alternative splicing events. Moreover, we have developed an algorithm that can design active TOSS with a success rate of 80%. To produce bifunctional oligonucleotides capable of stimulating splicing, we built on the observation that binding sites for TDP-43 can stimulate splicing and improve U1 snRNP binding when inserted downstream from 50 splice sites. A TOES designed to recruit TDP-43 improved exon 7 inclusion in SMN2. Overall, our study shows that bifunctional oligonucleotides can redirect splicing on a variety of genes, justifying their inclusion in the molecular arsenal that aims to alter the production of splice variants

Efficacité à long terme de la pression positive continue sur la symptomatologie et les troubles respiratoires dans le syndrome d'apnées obstructives du sommeil

Des auteurs ont émis l'hypothèse d'une aggravation du syndrome d'apnées du sommeil (SAOS) au fil du temps. Il a été montré qu'il existe une variation intra-individuelle des niveaux de pression positive continue (PPC) efficace sur les symptômes du SAOS et sur les apnées. L'objectif de ce travail est l'étude de l'efficacité à long terme de la PPC sur les symptômes du SAOS et les apnées d'une part, la fonction respiratoire et la gazométrie artérielle d'autre part. Cette étude rétrospective a rassemblé 132 dossiers consécutifs de la cohorte ayant eu uni diagnostic de SAOS entre 1993 et 2000 et avec un suivi de plus de 4 ans. Un questionnaire, une gazométrie, une spirométrie et un examen polygraphique Ont été réalisés systématiquement lors du diagnostic (tO) et au moins 4 ans après (tl). Entre tO et tl, les symptômes s'améliorent (p<0,0001). La persistance de la somnolence est associée à un âge plus jeune (46 vs 54 ans) (p=0,0008). Il existe une amélioration de l'hématose. Chez les oatients hypoxémiques et normocapniques: augmentation de la PaO2 de 64 à 74 mmHg (p<O,OOI). Chez les patients hypoxémiques et hypercapniques : augmentation de la PaO2 de 64 à 74 mmHg (p<0,001) et diminution de la PaCO2 de 50 à 42 mmHg (p<O,OOI). Les paramètres spirométriques diminuent avec l'âge. Il existe une augmentation de la PPC 1 efficace de 0,4 cmH20 (p=0,002). Cette étude a démontré que les symptômes et les apnées du SAOS sont corrigés chez une majorité de patients. Le niveau de PPC efficace a augmenté. En cas d'insuffisance respiratoire associée, la PPC permet une amélioration des troubles gazométriques. La décroissance des paramètres fonctionnels respiratoires n'est pas modifiée.NANCY1-SCD Medecine (545472101) / SudocPARIS-BIUM (751062103) / SudocSudocFranceF

Boisements sur terres agricoles. Premier bilan en Languedoc-Roussillon

National audienc

A proteomic approach to the identification of heterogeneous nuclear ribonucleoproteins as a new family of poly(ADP-ribose)-binding proteins.

A new class of poly(ADP-ribose) (pADPr)-binding proteins, heterogeneous nuclear ribonucleoproteins (hnRNPs), has been identified by a proteomic approach using matrix-assisted laser-desorption-ionization time-of-flight ('MALDI-TOF') MS. Liquid-phase isoelectric focusing with a Rotofor cell (Bio-Rad) allowed pre-fractionation of proteins extracted from HeLa cells. Rotofor protein fractions were further separated by SDS/PAGE and then transferred to a PVDF membrane. pADPr-binding proteins were analysed by autoradiography of the protein blot after incubation with (32)P-labelled automodified pADPr polymerase-1 (PARP-1). Peptide mass fingerprinting of selected bands identified the most abundant pADPr-binding proteins as hnRNPs, a family of proteins that bind pre-mRNA into functional complexes involved in mRNA maturation and transport to the cytoplasm. Sequence homology database searching against a previously reported pADPr-binding sequence motif revealed that the hnRNPs contain a putative pADPr-binding sequence pattern [Pleschke, Kleczkowska, Strohm and Althaus (2000) J. Biol. Chem. 275, 40974-40980]. pADPr-binding assays performed with synthetic peptides by the dot-blot technique and with nitrocellulose-transferred recombinant hnRNPs confirmed the pADPr-binding protein identification and the specificity of the interaction. These results could establish a link between increased levels of pADPr in DNA damaged cells and the modified protein expression pattern resulting from altered mRNA trafficking

Deux fermes voisines des XVIe- début XVIIIe siècles

International audienc

Role of the splicing factor SRSF4 in cisplatin-induced modifications of pre-mRNA splicing and apoptosis.

BACKGROUND: Modification of splicing by chemotherapeutic drugs has usually been evaluated on a limited number of pre-mRNAs selected for their recognized or potential importance in cell proliferation or apoptosis. However, the pathways linking splicing alterations to the efficiency of cancer therapy remain unclear. METHODS: Next-generation sequencing was used to analyse the transcriptome of breast carcinoma cells treated by cisplatin. Pharmacological inhibitors, RNA interference, cells deficient in specific signalling pathways, RT-PCR and FACS analysis were used to investigate how the anti-cancer drug cisplatin affected alternative splicing and the cell death pathway. RESULTS: We identified 717 splicing events affected by cisplatin, including 245 events involving cassette exons. Gene ontology analysis indicates that cell cycle, mRNA processing and pre-mRNA splicing were the main pathways affected. Importantly, the cisplatin–induced splicing alterations required class I PI3Ks P110β but not components such as ATM, ATR and p53 that are involved in the DNA damage response. The siRNA-mediated depletion of the splicing regulator SRSF4, but not SRSF6, expression abrogated many of the splicing alterations as well as cell death induced by cisplatin. CONCLUSION: Many of the splicing alterations induced by cisplatin are caused by SRSF4 and they contribute to apoptosis in a process requires class I PI3K. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12885-015-1259-0) contains supplementary material, which is available to authorized users

Tumor microenvironment–associated modifications of alternative splicing

Abstract: Pre-mRNA alternative splicing is modified in cancer, but the origin and specificity of these changes remain unclear. Here, we probed ovarian tumors to identify cancer-associated splicing isoforms and define the mechanism by which splicing is modified in cancer cells. Using high-throughput quantitative PCR, we monitored the expression of splice variants in laser-dissected tissues from ovarian tumors. Surprisingly, changes in alternative splicing were not limited to the tumor tissues but were also found in the tumor microenvironment. Changes in the tumor-associated splicing events were found to be regulated by splicing factors that are differentially expressed in cancer tissues. Overall, ∼20% of the alternative splicing events affected by the down-regulation of the splicing factors QKI and RBFOX2 were altered in the microenvironment of ovarian tumors. Together, our results indicate that the tumor microenvironment undergoes specific changes in alternative splicing orchestrated by a limited number of splicing factors

Les fermes d’Herneuse à Verberie (Oise). Deux fermes voisines des XVIe-début XVIIIe siècles

Im Norden von Verberie, zwischen der Oise und dem königlichen Forst von Compiègne wurden zwei Bauernhöfe des 16. und Anfang des 18. Jh. ausgegraben. Obwohl sie geschliffen worden waren, ist die Organisation der Höfe zu erkennen, zudem können das Keramik-und Metallmobiliar (Geschirr und Werkzeuge) sowie Spuren von Viehzucht und Fleischnahrung identifiziert werden. Anhand von Urkunden aus den Archiven des Departements wurden die Hypothesen bezüglich der Natur und der Funktion der bei der Grabung erkannten Spuren bestätigt. Zudem konnten diese Höfe in ihrem geschichtlichen Umfeld erfasst werden. Sie besaßen unterschiedliche rechtliche Stellungen, bei dem einen handelte es sich um kirchlichen, bei dem anderen um weltlichen Grundbesitz. Sie waren Gegenstand von Pachtverträgen, von denen bis heute nur wenige Erwähnungen gefunden wurden. Archäologische Beispiele dieser Art sind selten. Sie veranschaulichen die neuzeitlichen Bauernhöfe des Valois aus der Periode zwischen der Krisenzeit des 15. Jh. und der Phase der Konzentration des Grundbesitzes im späten 17. Jh.Two farms in activity from the 16th to the early 18th century have been excavated north of Verberie, between the river Oise and the royal forest of Compiègne. Although levelled off, the remains have yielded information about the organization of the farms, their ceramic and metallic utensils (crockery and tools), and some evidence concerning the cattle and the consumption of meat. Limited documentary evidence from the district Archives tends to support the hypotheses concerning the nature and the function of the remains discovered in the excavations and provides the basis for a preliminary historical overview of the environment of these settlements. The two farms did not have the same status, the one being ecclesiastical and the other secular, and they were the subject of successive leases to which only a few references have so far been found in our documentary evidence. Such archaeological examples are rare ; they appear to be, on their own scale, typical of the farms in the Valois district during the Modern era, between the crisis of the 15th century and the phase of concentration of the landed property at the end of the 17th century.Deux fermes, des XVIe siècle-début XVIIIe siècle, ont été fouillées au nord de Verberie, entre l’Oise et la forêt royale de Compiègne. Bien qu’arasés, les vestiges permettent la reconnaissance de l’organisation des fermes, de leur mobilier céramique et métallique (vaisselle et outils) et des témoignages liés au bétail et à l’alimentation carnée. Quelques éléments documentaires, recueillis aux archives départementales, ont permis d’étayer les hypothèses quant à la nature et la fonction des vestiges reconnus à la fouille et d’initier un panorama historique de l’environnement de ces établissements. De statuts différents, propriété ecclésiastique pour l’une et civile pour l’autre, elles ont fait l’objet de baux successifs dont seules de rares mentions ont été pour l’heure retrouvées. Ces exemples archéologiques sont rares, ils apparaissent, à leur échelle, comme des illustrations des fermes du Valois de l’époque moderne, compris entre la période de crise du XVe siècle et la phase de concentration foncière de la fin du XVIIe siècle.Bruley-Chabot Gaëlle, Clavel Benoît, Jouanin Gaëtan, Facquez Dorothée, Legros Vincent, Bernard Jean-Louis. Les fermes d’Herneuse à Verberie (Oise). Deux fermes voisines des XVIe-début XVIIIe siècles. In: Revue archéologique de Picardie, n°3-4, 2012. pp. 207-295

Effect of hypoxia on cod bioenergetics

International audienc