Selenoprotein N is dynamically expressed during mouse development and detected early in muscle precursors

<p>Abstract</p> <p>Background</p> <p>In humans, mutations in the <it>SEPN1 </it>gene, encoding selenoprotein N (SelN), are involved in early onset recessive neuromuscular disorders, referred to as <it>SEPN1</it>-related-myopathies. The mechanisms behind these pathologies are poorly understood since the function of SelN remains elusive. However, previous results obtained in humans and more recently in zebrafish pointed to a potential role for SelN during embryogenesis. Using qRT-PCR, Western blot and whole mount <it>in situ </it>hybridization, we characterized in detail the spatio-temporal expression pattern of the murine <it>Sepn1 </it>gene during development, focusing particularly on skeletal muscles.</p> <p>Results</p> <p>In whole embryos, <it>Sepn1 </it>transcripts were detected as early as E5.5, with expression levels peaking at E12.5, and then strongly decreasing until birth. In isolated tissues, only mild transcriptional variations were observed during development, whereas a striking reduction of the protein expression was detected during the perinatal period. Furthermore, we demonstrated that <it>Sepn1 </it>is expressed early in somites and restricted to the myotome, the sub-ectodermal mesenchyme and the dorsal root ganglia at mid-gestation stages. Interestingly, <it>Sepn1 </it>deficiency did not alter somitogenesis in embryos, suggesting that SelN is dispensable for these processes in mouse.</p> <p>Conclusion</p> <p>We characterized for the first time the expression pattern of <it>Sepn1 </it>during mammalian embryogenesis and we demonstrated that its differential expression is most likely dependent on major post-transcriptional regulations. Overall, our data strongly suggest a potential role for selenoprotein N from mid-gestation stages to the perinatal period. Interestingly, its specific expression pattern could be related to the current hypothesis that selenoprotein N may regulate the activity of the ryanodine receptors.</p

Small ncRNA transcriptome analysis from Aspergillus fumigatus suggests a novel mechanism for regulation of protein synthesis

Small non-protein-coding RNAs (ncRNAs) have systematically been studied in various model organisms from Escherichia coli to Homo sapiens. Here, we analyse the small ncRNA transcriptome from the pathogenic filamentous fungus Aspergillus fumigatus. To that aim, we experimentally screened for ncRNAs, expressed under various growth conditions or during specific developmental stages, by generating a specialized cDNA library from size-selected small RNA species. Our screen revealed 30 novel ncRNA candidates from known ncRNA classes such as small nuclear RNAs (snRNAs) and C/D box-type small nucleolar RNAs (C/D box snoRNAs). Additionally, several candidates for H/ACA box snoRNAs could be predicted by a bioinformatical screen. We also identified 15 candidates for ncRNAs, which could not be assigned to any known ncRNA class. Some of these ncRNA species are developmentally regulated implying a possible novel function in A. fumigatus development. Surprisingly, in addition to full-length tRNAs, we also identified 5′- or 3′-halves of tRNAs, only, which are likely generated by tRNA cleavage within the anti-codon loop. We show that conidiation induces tRNA cleavage resulting in tRNA depletion within conidia. Since conidia represent the resting state of A. fumigatus we propose that conidial tRNA depletion might be a novel mechanism to down-regulate protein synthesis in a filamentous fungus

Direct cloning of double-stranded RNAs from RNase protection analysis reveals processing patterns of C/D box snoRNAs and provides evidence for widespread antisense transcript expression

We describe a new method that allows cloning of double-stranded RNAs (dsRNAs) that are generated in RNase protection experiments. We demonstrate that the mouse C/D box snoRNA MBII-85 (SNORD116) is processed into at least five shorter RNAs using processing sites near known functional elements of C/D box snoRNAs. Surprisingly, the majority of cloned RNAs from RNase protection experiments were derived from endogenous cellular RNA, indicating widespread antisense expression. The cloned dsRNAs could be mapped to genome areas that show RNA expression on both DNA strands and partially overlapped with experimentally determined argonaute-binding sites. The data suggest a conserved processing pattern for some C/D box snoRNAs and abundant expression of longer, non-coding RNAs in the cell that can potentially form dsRNAs

Increased Muscle Stress-Sensitivity Induced by Selenoprotein N Inactivation in Mouse: A Mammalian Model for SEPN1-Related Myopathy

Selenium is an essential trace element and selenoprotein N (SelN) was the first selenium-containing protein shown to be directly involved in human inherited diseases. Mutations in the SEPN1 gene, encoding SelN, cause a group of muscular disorders characterized by predominant affection of axial muscles. SelN has been shown to participate in calcium and redox homeostasis, but its pathophysiological role in skeletal muscle remains largely unknown. To address SelN function in vivo, we generated a Sepn1-null mouse model by gene targeting. The Sepn1−/− mice had normal growth and lifespan, and were macroscopically indistinguishable from wild-type littermates. Only minor defects were observed in muscle morphology and contractile properties in SelN-deficient mice in basal conditions. However, when subjected to challenging physical exercise and stress conditions (forced swimming test), Sepn1−/− mice developed an obvious phenotype, characterized by limited motility and body rigidity during the swimming session, as well as a progressive curvature of the spine and predominant alteration of paravertebral muscles. This induced phenotype recapitulates the distribution of muscle involvement in patients with SEPN1-Related Myopathy, hence positioning this new animal model as a valuable tool to dissect the role of SelN in muscle function and to characterize the pathophysiological process

RNPomics: Defining the ncRNA transcriptome by cDNA library generation from ribonucleo-protein particles

Up to 450 000 non-coding RNAs (ncRNAs) have been predicted to be transcribed from the human genome. However, it still has to be elucidated which of these transcripts represent functional ncRNAs. Since all functional ncRNAs in Eukarya form ribonucleo-protein particles (RNPs), we generated specialized cDNA libraries from size-fractionated RNPs and validated the presence of selected ncRNAs within RNPs by glycerol gradient centrifugation. As a proof of concept, we applied the RNP method to human Hela cells or total mouse brain, and subjected cDNA libraries, generated from the two model systems, to deep-sequencing. Bioinformatical analysis of cDNA sequences revealed several hundred ncRNP candidates. Thereby, ncRNAs candidates were mainly located in intergenic as well as intronic regions of the genome, with a significant overrepresentation of intron-derived ncRNA sequences. Additionally, a number of ncRNAs mapped to repetitive sequences. Thus, our RNP approach provides an efficient way to identify new functional small ncRNA candidates, involved in RNP formation

Une approche originale de sélection de nouveaux ARN non codants

Deux principales familles d’acides ribonucléiques ou ARN ont été identifiées jusqu’à présent dans tous les organismes vivants (voir Glossaire) [

Une mutation ponctuelle dans la protéine Rrp9 de la particule U3 entraîne un découplage des clivages précoces de l’ARN pré-ribosomique

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Toward understanding selenium and selenoprotein N functions in muscular disorders

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Toward understanding selenium and selenoprotein N functions in muscular disorders

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Generation of cDNA Libraries from RNP-Derived Regulatory Noncoding RNAs

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