Sequence-specific inhibition of RNA polymerase III-dependent transcription using Zorro locked nucleic acid (LNA)
✍ Scribed by Rongbin Ge; Mathias G. Svahn; Oscar E. Simonson; Abdalla J. Mohamed; Karin E. Lundin; C. I. Edvard Smith
- Publisher
- John Wiley and Sons
- Year
- 2007
- Tongue
- English
- Weight
- 534 KB
- Volume
- 10
- Category
- Article
- ISSN
- 1099-498X
- DOI
- 10.1002/jgm.1124
No coin nor oath required. For personal study only.
✦ Synopsis
Abstract
Background
RNA polymerase III (pol III)‐dependent transcripts are involved in many fundamental activities in a cell, such as splicing and protein synthesis. They also regulate cell growth and influence tumor formation. During recent years vector‐based systems for expression of short hairpin (sh) RNA under the control of a pol III promoter have been developed as gene‐based medicines. Therefore, there is an increasing interest in means to regulate pol III‐dependent transcription. Recently, we have developed a novel anti‐gene molecule ‘Zorro LNA (Locked Nucleic Acid)’, which simultaneously hybridizes to both strands of super‐coiled DNA and potently inhibits RNA polymerase II‐derived transcription. We have now applied Zorro LNA in an attempt to also control U6 promoter‐driven expression of shRNA.
Methods
In this study, we constructed pshluc and pshluc2BS plasmids, in which U6 promoter‐driven small hairpin RNA specific for luciferase gene (shluc) was without or with Zorro LNA binding sites, respectively. After hybridization of Zorro LNA to pshluc2BS, the LNA‐bound plasmid was cotransfected with pEGFPluc into mammalian cells and into a mouse model. In cellular experiments, cotransfection of unhybridized pshluc2BS, Zorro LNA and pEGFPluc was also performed.
Results
The results showed that the Zorro LNA construct efficiently inhibited pol III‐dependent transcription as an anti‐gene reagent in a cellular context, including in vivo in a mouse model.
Conclusions
Thus, this new form of gene silencer ‘Zorro LNA’ could potentially serve as a versatile regulator of pol III‐dependent transcription, including various forms of shRNAs. Copyright © 2007 John Wiley & Sons, Ltd.