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An Ecdysone Receptor-based Singular Gene Switch for Deliberate Expression of Transgene with Robustness, Reversibility, and Negligible Leakiness
Published on: May 7, 2018
A dual switch controls bacterial enhancer-dependent transcription.
Simone C Wiesler1, Patricia C Burrows, Martin Buck
1Department of Life Sciences, Imperial College London, Sir Alexander Fleming Building, London SW7 2AZ, UK. s.wiesler@imperial.ac.uk
Nucleic Acids Research
|September 12, 2012
Summary
Bacterial RNA polymerases (RNAPs) are antibiotic targets. Enhancer-dependent RNAPs and enhancer-binding proteins (bEBPs) resist myxopyronin inhibition by regulating transcription initiation through dual DNA opening barriers.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Bacterial RNA polymerases (RNAPs) are crucial for gene transcription and are primary targets for antibiotic development.
- Myxopyronin is an antibiotic that inhibits RNAP activity by binding to switch regions, preventing essential structural changes for transcription initiation.
- The σ(54) factor, in conjunction with enhancer-binding proteins (bEBPs), regulates transcription of genes involved in bacterial pathogenicity and stress responses.
Purpose of the Study:
- To investigate the role of enhancer-dependent RNAPs in bacterial survival against myxopyronin.
- To elucidate the mechanism by which bEBPs mediate resistance to myxopyronin inhibition.
- To understand the structural basis for regulated transcription initiation at enhancer-dependent promoters.
Main Methods:
- Comparative analysis of bacterial survival in the presence of myxopyronin with and without enhancer-dependent RNAPs.
- Biochemical assays to assess the inhibitory effects of myxopyronin on enhancer-dependent RNAP activity.
- Enzyme kinetic studies to determine the role of ATP hydrolysis by bEBPs in RNAP-promoter interactions.
Main Results:
- Escherichia coli strains utilizing enhancer-dependent RNAPs exhibit increased survival in the presence of myxopyronin.
- Enhancer-dependent RNAPs demonstrate partial resistance to myxopyronin-induced inhibition.
- ATP hydrolysis by bEBPs is essential for the proper interaction between RNAP switch regions and the transcription start site.
Conclusions:
- Enhancer-dependent RNAPs provide a survival advantage to bacteria under antibiotic pressure.
- The mechanism of resistance involves bEBPs facilitating a dual DNA opening process at promoters, creating regulatory barriers.
- This intricate regulation ensures precise control over transcription initiation, offering a target for novel antibiotic strategies.
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