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Nucleic Acids Research, 2001, Vol. 29, No. 3 622-628
© 2001 Oxford University Press

Triplex forming oligonucleotide targeted to 3'UTR downregulates the expression of the bcl-2 proto-oncogene in HeLa cells

Changxian Shen, Andreas Buck, Gerhard Mehrke, Bülent Polat, Hans-Jügen Gross1, Max Bachem1 and Sven Reske*

Department of Nuclear Medicine and 1Department of Clinical Chemistry, University of Ulm, Germany

The bcl-2 proto-oncogene is overexpressed in a variety of human cancers and plays an important role in programmed cell death. Recent reports implied that the 3'-untranslated region (3'UTR) functions effectively in the regulation of gene expression. Here, we attempt to assay the ability of triplex forming oligonucleotides (TFOs) to inhibit expression of a target gene in vivo and to examine the potential of the 3'UTR of the bcl-2 proto-oncogene in the regulation of bcl-2 gene expression. To do this, we have developed a novel cellular system that involves transfection of a Doxycyclin inducible expression plasmid containing the bcl-2 ORF and the 3'UTR together with a TFO targeted to the 3'UTR of the bcl-2 proto-oncogene. Phosphorothioate-modified TFO targeted to the 3'UTR of the bcl-2 gene significantly downregulated the expression of the bcl-2 gene in HeLa cells as demonstrated by western blotting. Our results indicate that blocking the functions of the 3'UTR using the TFO can downregulate the expression of the targeted gene, and suggest that triplex strategy is a promising approach for oligonucleotide-based gene therapy. In addition, triplex-based sequence targeting may provide a useful tool for studying the regulation of gene expression.

* To whom correspondence should be addressed at: Department of Nuclear Medicine, University of Ulm, Robert-Koch-Straße 8, D-89070 Ulm, Germany. Tel: +49 731 5024500; Fax: +49 731 5024503; Email: sven.reske{at}medizin.uni-ulm.de


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