Gene expression profile changes after short-activating RNA-mediated induction of endogenous pluripotency factors in human mesenchymal stem cells

It is now recognized that small noncoding RNA sequences have the ability to mediate transcriptional activation of specific target genes in human cells. Using bioinformatics analysis and functional screening, we screened short-activating RNA (saRNA) oligonucleotides designed to target the promoter regions of the pluripotency reprogramming factors, Kruppel-like factor 4 (KLF4) and c-MYC. We identified KLF4 and c-MYC promoter-targeted saRNA sequences that consistently induced increases in their respective levels of nascent mRNA and protein expression in a time- and dose-dependent manner, as compared with scrambled sequence control oligonucleotides. The functional consequences of saRNA-induced activation of each targeted reprogramming factor were then characterized by comprehensively profiling changes in gene expression by microarray analysis, which revealed significant increases in mRNA levels of their respective downstream pathway genes. Notably, the microarray profile after saRNA-mediated induction of endogenous KLF4 and c-MYC showed similar gene expression patterns for stem cell- and cell cycle-related genes as compared with lentiviral vector-mediated overexpression of exogenous KLF4 and c-MYC transgenes, while divergent gene expression patterns common to viral vector-mediated transgene delivery were also noted. The use of promoter-targeted saRNAs for the activation of pluripotency reprogramming factors could have broad implications for stem cell research.publishedVersionCopyright © 2012 American Society of Gene & Cell Therapy. Published by Elsevier Inc. Attribution-NonCommercial-NoDerivs 3.0 Unported (CC BY-NC-ND 3.0

A role for human Dicer in pre-RISC loading of siRNAs

RNA interference is a powerful mechanism for sequence-specific inhibition of gene expression. It is widely known that small interfering RNAs (siRNAs) targeting the same region of a target-messenger RNA can have widely different efficacies. In efforts to better understand the siRNA features that influence knockdown efficiency, we analyzed siRNA interactions with a high-molecular weight complex in whole cell extracts prepared from two different cell lines. Using biochemical tools to study the nature of the complex, our results demonstrate that the primary siRNA-binding protein in the whole cell extracts is Dicer. We find that Dicer is capable of discriminating highly functional versus poorly functional siRNAs by recognizing the presence of 2-nt 3′ overhangs and the thermodynamic properties of 2–4 bp on both ends of effective siRNAs. Our results suggest a role for Dicer in pre-selection of effective siRNAs for handoff to Ago2. This initial selection is reflective of the overall silencing potential of an siRNA

Gene expression profile changes after short-activating RNA-mediated induction of endogenous pluripotency factors in human mesenchymal stem cells

It is now recognized that small noncoding RNA sequences have the ability to mediate transcriptional activation of specific target genes in human cells. Using bioinformatics analysis and functional screening, we screened short-activating RNA (saRNA) oligonucleotides designed to target the promoter regions of the pluripotency reprogramming factors, Kruppel-like factor 4 (KLF4) and c-MYC. We identified KLF4 and c-MYC promoter-targeted saRNA sequences that consistently induced increases in their respective levels of nascent mRNA and protein expression in a time- and dose-dependent manner, as compared with scrambled sequence control oligonucleotides. The functional consequences of saRNA-induced activation of each targeted reprogramming factor were then characterized by comprehensively profiling changes in gene expression by microarray analysis, which revealed significant increases in mRNA levels of their respective downstream pathway genes. Notably, the microarray profile after saRNA-mediated induction of endogenous KLF4 and c-MYC showed similar gene expression patterns for stem cell- and cell cycle-related genes as compared with lentiviral vector-mediated overexpression of exogenous KLF4 and c-MYC transgenes, while divergent gene expression patterns common to viral vector-mediated transgene delivery were also noted. The use of promoter-targeted saRNAs for the activation of pluripotency reprogramming factors could have broad implications for stem cell research

Studies on W.B. Yeats

Dans ce recueil convergent différents regards sur la poésie de W.B. Yeats. Ces pages le situent par rapport à d’autres poètes comme MacNeice. Elles nous promènent aussi des premiers volumes, où l’espace de l’écriture devient l’écriture de l’espace, à Responsabilities, volume qui témoigne d’une intensité qu’Ezra Pound qualifie de « robustesse nouvelle », puis à travers les thèmes sexuels, politiques et esthétiques de Michael Robartes and the Dancer. Nous abordons ensuite les grands poèmes de la maturité avec The Tower. Cet ouvrage envisage également des sujets généraux, comme l’importance de la tradition pour Yeats, sa conception de l’au-delà avec la dette envers l’Inde en particulier, son attitude face à la dégénérescence, sa vision de l’Apocalypse avec le symbole de la spirale. D’autres études se concentrent sur certains grands poèmes tels que « The Circus Animais Desertion », qui tisse ensemble et la vie et l’art, ou « Lapis Lazuli » et la notion de « joie tragique ».In this book, different perspectives on W. B. Yeats' poetry converge. These pages place him in relation to other poets such as MacNeice. They also take us from the first volumes, where the space of writing becomes the writing of space, to Responsabilities, a volume that reflects an intensity that Ezra Pound calls "new robustness", and then through the sexual, political and aesthetic themes of Michael Robartes and the Dancer. We then discuss the great poems of maturity with The Tower. This book also considers general topics, such as the importance of tradition for Yeats, his conception of the afterlife with the debt to India in particular, his attitude towards degeneration, his vision of the Apocalypse with the symbol of the spiral. Other studies focus on some major poems such as "The Circus Animais Desertion", which weaves together life and art, or "Lapis Lazuli" and the notion of "tragic joy"