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Nucleic Acids Research 2006 34(5):e37; doi:10.1093/nar/gkl034
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Published online 6 March 2006

© The Author 2006. Published by Oxford University Press. All rights reserved
The online version of this article has been published under an open access model. Users are entitled to use, reproduce, disseminate, or display the open access version of this article for non-commercial purposes provided that: the original authorship is properly and fully attributed; the Journal and Oxford University Press are attributed as the original place of publication with the correct citation details given; if an article is subsequently reproduced or disseminated not in its entirety but only in part or as a derivative work this must be clearly indicated. For commercial re-use, please contact journals.permissions@oxfordjournals.org

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Control of small inhibitory RNA levels and RNA interference by doxycycline induced activation of a minimal RNA polymerase III promoter

Lahouari Amar, Mathieu Desclaux, Nicole Faucon-Biguet, Jacques Mallet* and Roland Vogel

Laboratoire de la Génétique Moléculaire de la Neurotransmission et des Processus Neurodégénératifs (LGN), CNRS-UMR 7091, Bâtiment CERVI, Hôpital de la Pitié Salpêtrière, 83 Boulevard de l'Hôpital 75013 Paris, France

*To whom correspondence should be addressed. Tel: +33 1 42 17 75 32; Fax: +33 1 42 17 75 33; Email: mallet{at}chups.jussieu.fr

Received September 21, 2005. Revised November 21, 2005. Accepted February 21, 2006.

RNA interference (RNAi) mediated by expression of short hairpin RNAs (shRNAs) is a powerful tool for efficiently suppressing target genes. The approach allows studies of the function of individual genes and may also be applied to human therapy. However, in many instances regulation of RNAi by administration of a small inducer molecule will be required. To date, the development of appropriate regulatory systems has been hampered by the few possibilities for modification within RNA polymerase III promoters capable of driving efficient expression of shRNAs. We have developed an inducible minimal RNA polymerase III promoter that is activated by a novel recombinant transactivator in the presence of doxycycline (Dox). The recombinant transactivator and the engineered promoter together form a system permitting regulation of RNAi by Dox-induced expression of shRNAs. Regulated RNAi was mediated by one single lentiviral vector, blocked the expression of green fluorescent protein (GFP) in a GFP-expressing HEK 293T derived cell line and suppressed endogenous p53 in wild-type HEK 293T, MCF-7 and A549 cells. RNA interference was induced in a dose- and time-dependent manner by administration of Dox, silenced the expression of both target genes by 90% and was in particular reversible after withdrawal of Dox.


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