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TECHNOLOGY DEVELOPMENT

A Cassette System to Study Embryonic Stem Cell Differentiation by Inducible RNA Interference

^aInstitute for Medical Microbiology, Department of Biological and Clinical Sciences, University of Basel, Basel, Switzerland;
^bFriedrich Miescher Institute for Biomedical Research, Basel, Switzerland

Key Words. Embryonic stem cells • Cardiac development • RNA interference • Tetracycline • Zfp36L1 • mRNA turnover

Correspondence: Christoph Moroni, M.D., University of Basel, Institute for Medical Microbiology, Petersplatz 10, 4051 Basel, Switzerland. Telephone: +41 61 2673262; Fax: +41 61 267 32 83; e-mail: Christoph.Moroni{at}unibas.ch

Received February 24, 2006; accepted for publication January 2, 2007.
First published online in STEM CELLS EXPRESS   January 11, 2007.



Although differentiation of pluripotent embryonic stem cells is restricted by a hierarchy of transcription factors, little is known about whether post-transcriptional mechanisms similarly regulate early embryoid differentiation. We developed a system where small hairpin (sh)RNAs can be induced in embryonic stem (ES) cells from a defined locus following integration by Flp recombinase-mediated DNA recombination. To verify the system, the key transcription factor Stat3, which maintains pluripotency, was downregulated by shRNA, and the expected morphological and biochemical markers of differentiation were observed. Induction of shRNA specific for the post-transcriptional regulator Brf1 (Zfp36L1) amplified the cardiac markers with strong stimulation of cardiomyocyte formation within embryoid bodies. These findings identify Brf1 as a novel potential regulator of cardiomyocyte formation and suggest that post-transcriptional mechanisms are of importance to early development and, possibly, to regenerative medicine. The inducible RNA interference system presented here should also allow assignment of function for candidate genes with suspected roles in ES cell development.

Disclosure of potential conflicts of interest is found at the end of this article.

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				J. Wang, T. W. Theunissen, and S. H. Orkin Site-directed, virus-free, and inducible RNAi in embryonic stem cells PNAS, December 26, 2007; 104(52): 20850 - 20855. [Abstract] [Full Text] [PDF]