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Open Access Methodology article

Targeted protein depletion in Saccharomyces cerevisiae by activation of a bidirectional degron

Marc Jungbluth, Christian Renicke and Christof Taxis*

Author Affiliations

Department of Genetics, Philipps-Universität Marburg, Karl-von-Frisch-Str. 8, 35043 Marburg, Germany

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BMC Systems Biology 2010, 4:176  doi:10.1186/1752-0509-4-176

Published: 29 December 2010

Abstract

Background

Tools for in vivo manipulation of protein abundance or activity are highly beneficial for life science research. Protein stability can be efficiently controlled by conditional degrons, which induce target protein degradation at restrictive conditions.

Results

We used the yeast Saccharomyces cerevisiae for development of a conditional, bidirectional degron to control protein stability, which can be fused to the target protein N-terminally, C-terminally or placed internally. Activation of the degron is achieved by cleavage with the tobacco etch virus (TEV) protease, resulting in quick proteolysis of the target protein. We found similar degradation rates of soluble substrates using destabilization by the N- or C-degron. C-terminal tagging of essential yeast proteins with the bidirectional degron resulted in deletion-like phenotypes at non-permissive conditions. Developmental process-specific mutants were created by N- or C-terminal tagging of essential proteins with the bidirectional degron in combination with sporulation-specific production of the TEV protease.

Conclusions

We developed a system to influence protein abundance and activity genetically, which can be used to create conditional mutants, to regulate the fate of single protein domains or to design artificial regulatory circuits. Thus, this method enhances the toolbox to manipulate proteins in systems biology approaches considerably.