Molecular basis of differential target regulation by miR-96 and miR-182: the Glypican-3 as a model

Besides the fact that miR-96 and miR-182 belong to the miR-182/183 cluster, their seed region (UUGGCA, nucleotides 2–7) is identical suggesting potential common properties in mRNA target recognition and cellular functions. Here, we used the mRNA encoding Glypican-3, a heparan-sulfate proteoglycan, as a model target as its short 3′ untranslated region is predicted to contain one miR-96/182 site, and assessed whether it is post-transcriptionally regulated by these two microRNAs. We found that miR-96 downregulated GPC3 expression by targeting its mRNA 3′-untranslated region and interacting with the predicted site. This downregulatory effect was due to an increased mRNA degradation and depended on Argonaute-2. Despite its seed similarity with miR-96, miR-182 was unable to regulate GPC3. This differential regulation was confirmed on two other targets, FOXO1 and FN1. By site-directed mutagenesis, we demonstrated that the miRNA nucleotide 8, immediately downstream the UUGGCA seed, plays a critical role in target recognition by miR-96 and miR-182. Our data suggest that because of a base difference at miRNA position 8, these two microRNAs control a completely different set of genes and therefore are functionally independent

Expression and Function of Osteopontin in Vascular Adventitial Fibroblasts and Pathological Vascular Remodeling

Osteopontin is known to play important roles in various diseases including vascular disorders. However, little is known about its expression and function in vascular adventitial fibroblasts. Adventitial fibroblasts have been shown to play a key role in pathological vascular remodeling associating with various vascular disorders. In this study, we measured activation of Osteopontin and its biological functions in cultured adventitial fibroblasts and injured rat carotid injury arteries induced by balloon angioplasty. Our results showed that angiotensin II and aldosterone increased Osteopontin expression in adventitial fibroblasts in a time- and concentration-dependent manner. MAPKs and AP-1 pathways were involved in Osteopontin upregulation. In addition, Adventitial fibroblast migration stimulated by Angiotensin II and aldosterone required OPN expression. Perivascular delivery of antisense oligonucleotide for Osteopontin suppressed neointimal formation post-injury. We concluded that upregulation of Osteopontin expression in adventitial fibroblasts might be important in the pathogenesis of vascular remodeling after arterial injury

Transcriptional and Post-Transcriptional Regulation of SPAST, the Gene Most Frequently Mutated in Hereditary Spastic Paraplegia

Hereditary spastic paraplegias (HSPs) comprise a group of neurodegenerative disorders that are characterized by progressive spasticity of the lower extremities, due to axonal degeneration in the corticospinal motor tracts. HSPs are genetically heterogeneous and show autosomal dominant inheritance in ∼70–80% of cases, with additional cases being recessive or X-linked. The most common type of HSP is SPG4 with mutations in the SPAST gene, encoding spastin, which occurs in 40% of dominantly inherited cases and in ∼10% of sporadic cases. Both loss-of-function and dominant-negative mutation mechanisms have been described for SPG4, suggesting that precise or stoichiometric levels of spastin are necessary for biological function. Therefore, we hypothesized that regulatory mechanisms controlling expression of SPAST are important determinants of spastin biology, and if altered, could contribute to the development and progression of the disease. To examine the transcriptional and post-transcriptional regulation of SPAST, we used molecular phylogenetic methods to identify conserved sequences for putative transcription factor binding sites and miRNA targeting motifs in the SPAST promoter and 3′-UTR, respectively. By a variety of molecular methods, we demonstrate that SPAST transcription is positively regulated by NRF1 and SOX11. Furthermore, we show that miR-96 and miR-182 negatively regulate SPAST by effects on mRNA stability and protein level. These transcriptional and miRNA regulatory mechanisms provide new functional targets for mutation screening and therapeutic targeting in HSP

La migration des cellules musculaires lisses artérielles (rôle et régulation transcriptionnelle de l'ostéopontine)

La migration des cellules musculaires lisses (CML) artérielles tient une place prépondérante dans le développement des pathologies athéroscléreuses et la maturation des néovaisseaux. Afin de comprendre les mécanismes régulant la migration des CML, nous nous sommes proposés 1) de déterminer le rôle de l'ostéopontine (OPN) dans cette migration induite par différents agents chimiotactiques et 2) d'approfondir les mécanismes qui régulent son expression. Nous avons démontré que la production autocrine d'OPN est nécessaire à la migration des CML induite par l'UTP ou le PDGF. Ces facteurs agissent essentiellement au niveau de la transcription du gène de l'OPN. L'étude du promoteur de l'OPN a révélé que le facteur de transcriptiion CREB agit, selon l'agoniste, au niveau des sites CRE ou AP-1, probablement en formant un multicomplexe avec c-Fos. Les facteurs AP-1, USF et NF-kB sont aussi impliqués dans la régulation de l'OPN, de façon différentielle pour l'UTP, l'AII ou le PDGF. L'effet inhibiteur des oestrogènes sur la migration des CML est en partie dû à l'inhibition de l'expression d'OPN, en bloquant l'interaction entre AP-1 et USF.Arterial smooth muscle cells (SMC) migration is determinant in the development of atherosclerosis and in the maturation of newly formed vessels. In order to understand the mechanisms regulating SMC migration, we aimed at 1) determining the role of osteopontin (OPN) in SMC migration induced by different chemotactic factors and 2) understanding the mechanisms of OPN expression regulation. In this work, we showed that autocrine production of OPN is involved in UTP- or PDGF-induced SMC migration. These agonists regulate OPN transcription. We showed, by using promoter analysis methods, that CREB transcription factors acts, depending on the agonist, on CRE or AP-1 sites, probably by forming a multicomplex with c-Fos. Moreover, AP-1, USF and NF-kB factors are differentially involved in UTP, AII- or PDGF-induced OPN expression. Finally, we showed that the inhibitory effect of estrogens on SMC migration can be partially explained by OPN inhibition due to abolition of AP-1 /USF interaction.BORDEAUX2-BU Santé (330632101) / SudocSudocFranceF

ExAO ? De quoi s'agit-il ?

Sommaire du numéro :http://archive-edutice.ccsd.cnrs.fr/edutice-00000857Le matériel ExAO comprend un ordinateur associé à une interface et différents capteurs. Il permet de mesurer en temps réel des variations de divers paramètres sur des individus, des cellules, des organites cellulaires. L'expérimentation assistée par ordinateur (ExAO) entre de plus en plus dans l'enseignement de la Biologie. Les établissements s'équipent le plus souvent d'abord avec un poste ; cette étape est nécessaire pour démystifier l'ordinateur auprès des professeurs de Biologie. Très vite les enseignants s'aperçoivent qu'ils ne font pas de l'informatique mais qu'ils disposent d'un outil aux possibilités particulièrement intéressantes pour les élèves et pour la conduite de la classe.Les élèves aussi ont un avis sur l'ExAO ; une consultation réalisée durant l'année scolaire 1989-1990 avec deux classes de 1re S souligne un accueil très favorable, une perception aiguë des aspects positifs introduits par cette innovation ; les citations en caractère italique dans le texte représentent les réponses écrites des élèves à la question posée

Analysis of post-transcriptional regulation using the FunREG method

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