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Nucleic Acids Research, 2003, Vol. 31, No. 7 1853-1858
© 2003 Oxford University Press

Conditional gene expression by controlling translation with tetracycline-binding aptamers

Beatrix Suess, Shane Hanson, Christian Berens1, Barbara Fink, Renée Schroeder1 and Wolfgang Hillen

Lehrstuhl für Mikrobiologie, Friedrich-Alexander-Universität Erlangen-Nürnberg, Staudtstrasse 5, 91058 Erlangen, Germany and 1 Lehrstuhl für Genetik, Universität Wien, Dr Bohrgasse 9/4, A-1030 Wien, Austria

*To whom correspondence should be addressed. Tel: +49 9131 8528085; Fax: +49 9131 8528082; Email: bsuess{at}biologie.uni-erlangen.de

We present a conditional gene expression system in Saccharomyces cerevisiae which exploits direct RNA–metabolite interactions as a mechanism of genetic control. We inserted preselected tetracycline (tc) binding aptamers into the 5'-UTR of a GFP encoding mRNA. While aptamer insertion generally reduces GFP expression, one group of aptamers displayed an additional, up to 6-fold, decrease in fluorescence upon tc addition. Regulation is observed for aptamers inserted cap-proximal or near the start codon, but is more pronounced from the latter position. Increasing the thermodynamic stability of the aptamer augments regulation but reduces expression of GFP. Decreasing the stability leads to the opposite effect. We defined nucleotides which influence the regulatory properties of the aptamer. Exchanging a nucleotide probably involved in tc binding only influences regulation, while mutations at another position alter expression in the absence of tc, without affecting regulation. Thus, we have developed and characterized a regulatory system which is easy to establish and controlled by a non-toxic, small ligand with good cell permeability.


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