A Genetic Tool to Quantify trans-Translation Activity in Vivo.

Autor: Macé K; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Demay F; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Guyomar C; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Georgeault S; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Giudice E; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Goude R; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Trautwetter A; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Ermel G; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France., Blanco C; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France. Electronic address: carlos.blanco@univ-rennes1.fr., Gillet R; Univ. Rennes, CNRS, Institut de Génétique et Développement de Rennes (IGDR) UMR6290, 35000 Rennes, France. Electronic address: reynald.gillet@univ-rennes1.fr.
Jazyk: angličtina
Zdroj: Journal of molecular biology [J Mol Biol] 2017 Nov 24; Vol. 429 (23), pp. 3617-3625. Date of Electronic Publication: 2017 Oct 13.
DOI: 10.1016/j.jmb.2017.10.007
Abstrakt: In bacteria, trans-translation is the main quality control mechanism for rescuing ribosomes arrested during translation. This key process is universally conserved and plays a critical role in the viability and virulence of many pathogens. We developed a reliable in vivo double-fluorescence reporter system for the simultaneous quantification of both trans-translation and the associated proteolysis activities in bacteria. The assay was validated using mutant bacteria lacking tmRNA, SmpB, and the ClpP protease. Both antisense tmRNA-binding RNA and a peptide mimicking the SmpB C-terminal tail proved to be potent inhibitors of trans-translation in vivo. The double-fluorescent reporter was also tested with KKL-35, an oxadiazole derivative that is supposed to be a promising trans-translation inhibitor, and it surprisingly turns out that trans-translation is not the only target of KKL-35 in vivo.
(Copyright © 2017 Elsevier Ltd. All rights reserved.)
Databáze: MEDLINE
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