tet-regulation in bacteria
The minimal requirements for tet-regulation in bacteria consist of a viable target cell, a target gene placed downstream of a promoter amenable to tet-control, the regulator, encoded by tetR, and a suitable effector molecule. The figure below illustrates the basic principle.
Left: Repression of a target gene (grey arrow) downstream of a tet-sensitive promoter occurs, when TetR is bound to its cognate DNA sequence tet operator (tetO), hampering initiation of transcription; the target gene is in the OFF-state. Right: Target gene expression is initiated upon administration of tetracycline (Tc, orange) or a derivative thereof (such as anhydro-Tc, Degenkolb et al. 1991). Upon binding of the inducer to TetR, the repressor undergoes a conformational change, resulting in detachment from tetO (square) included in the promoter (bent arrow). The mechanisms underlying tet-regulation have been reviewed by Berens and Hillen 1994.
Characteristics of the tet-system for inducible gene expression in bacteria include
- particularly tight target gene repression
- an inert inducer not influencing the target organism's metabolome
- graded target gene expression, dependent on inducer concentration
- regulation of bacterial genes during infection
More than 13 different naturally occuring TetR classes have been identified to date (Review: Berens and Hillen 2004). In addition, artificially altered TetR variants with enhanced and altered properties have been developed within the last years and decades. These include mutant regulators, which
- bind to mutated tetO sequences (Helbl and Hillen 1998, Helbl et al. 1998, Krüger et al. 2007),
- interact with and get induced by different Tc derivatives (Scholz et al. 2003, Henssler et al. 2004), peptides (Klotzsche et al. 2005), or RNA aptamers (Hunsicker et al. 2009),
- show reverse behavior (i.e. bind to tetO in the presence of Tc molecules, Scholz et al. 2004) and thus enable to rapidly shut down transcription of a target gene.
These new TetR variants allow the independent regulation of two genes in one bacterial cell (Kamionka et al. 2004). Furthermore, the phenotypic alterations can be combined to e.g. obtain reverse TetR variants that bind to mutated operators (Bertram et al. 2004), or that can be co-repressed with Tc derivatives (Henssler et al. 2005).