Safe and easy in vitro evaluation of tmRNA-SmpB-mediated trans-translation from ESKAPE pathogenic bacteria.

Fiche publication

Date publication

août 2021

Journal

RNA (New York, N.Y.)

Auteurs

Membres identifiés du Cancéropôle Est :
Dr ENNIFAR Eric

Tous les auteurs :
Thepaut M, Campos-Silva R, Renard E, Barloy-Hubler F, Ennifar E, Boujard D, Gillet R

Lien Pubmed

https://www.ncbi.nlm.nih.gov/pubmed/34353925

Résumé

In bacteria, trans-translation is the major quality control system for rescuing stalled ribosomes. It is mediated by tmRNA, a hybrid RNA with properties of both a tRNA and a mRNA, and the small protein SmpB. Because trans-translation is absent in eukaryotes but necessary for bacterial fitness or survival, it is a promising target for the development of novel antibiotics. To facilitate screening of chemical libraries, various reliable in vitro and in vivo systems have been created for assessing trans-translational activity. However, the aim of the current work was to permit the safe and easy in vitro evaluation of trans-translation from pathogenic bacteria, which are obviously the ones we should be targeting. Based on green fluorescent protein (GFP) reassembly during active trans-translation, we have created a cell-free assay adapted to the rapid evaluation of trans-translation in ESKAPE bacteria, with 24 different possible combinations. It can be used for easy high-throughput screening of chemical compounds as well as for exploring the mechanism of trans-translation in these pathogens.