The wing of the ToxR winged helix-turn-helix domain is required for DNA binding and activation of toxT and ompU

English version

Autoři: Sarah J. Morgan ^aff001; Emily L. French ^aff002; Sarah C. Plecha ^aff002; Eric S. Krukonis ^aff001
Působiště autorů: Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, United States of America ^aff001; Division of Integrated Biomedical Sciences, University of Detroit Mercy School of Dentistry, Detroit, MI, United States of America ^aff002; Department of Biochemistry, Microbiology and Immunology, Wayne State University, Detroit, MI, United States of America ^aff003
Vyšlo v časopise: PLoS ONE 14(9)
Kategorie: Research Article
doi: https://doi.org/10.1371/journal.pone.0221936

Souhrn

ToxR and TcpP, two winged helix-turn-helix (w-HTH) family transcription factors, co-activate expression of the toxT promoter in Vibrio cholerae. ToxT then directly regulates a number of genes required for virulence. In addition to co-activation of toxT, ToxR can directly activate the ompU promoter and repress the ompT promoter. Based on a previous study suggesting that certain wing residues of ToxR are preferentially involved in toxT co-activation compared to direct ompU activation, we employed alanine-scanning mutagenesis to determine which residues in the wing of ToxR are required for activation of each promoter. All of the ToxR wing residues tested that were critical for transcriptional activation of toxT and/or ompU were also critical for DNA binding. While some ToxR wing mutants had reduced interaction with TcpP, that reduced interaction did not correlate with a specific defect in toxT activation. Rather, such mutants also affected ompU activation and DNA binding. Based on these findings we conclude that the primary role of the wing of ToxR is to bind DNA, along with the DNA recognition helix of ToxR, and this function is required both for direct activation of ompU and co-activation of toxT.

Klíčová slova:

Biology and life sciences – Genetics – Gene expression – DNA transcription – Biochemistry – Proteins – DNA-binding proteins – Cell biology – Cellular structures and organelles – Cell membranes – Membrane proteins – Outer membrane proteins – Microbiology – Medical microbiology – Microbial pathogens – Bacterial pathogens – Vibrio cholerae – Organisms – Bacteria – Vibrio – Research and analysis methods – Chemical characterization – Binding analysis – DNA binding assay – Immunologic techniques – Immunoassays – Enzyme-linked immunoassays – Extraction techniques – Protein extraction – Medicine and health sciences – Pathology and laboratory medicine – Pathogens – Physical sciences – Physics – Condensed matter physics – Solid state physics – Crystallography – Crystal structure

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The wing of the ToxR winged helix-turn-helix domain is required for DNA binding and activation of toxT and ompU

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