imprint:
Springer Science and Business Media LLC, 2017
Published in:Scientific Reports
Language:
English
DOI:
10.1038/s41598-017-07870-w
ISSN:
2045-2322
Origination:
Footnote:
Description:
<jats:title>Abstract</jats:title><jats:p>Riboswitches are bacterial RNA elements that regulate gene expression in response to metabolite or ion abundance and are considered as potential drug targets. In recent years a number of methods to find non-natural riboswitch ligands have been described. Here we report a high-throughput <jats:italic>in vivo</jats:italic> screening system that allows identifying OFF-riboswitch modulators in a 384 well bioluminescence assay format. We use a reverse reporter gene setup in <jats:italic>Bacillus subtilis</jats:italic>, consisting of a primary screening assay, a secondary assay as well as counter assays to detect compounds in a library of 1,280 molecules that act on the guanine-responsive <jats:italic>xpt</jats:italic> riboswitch from <jats:italic>B. anthracis</jats:italic>. With this <jats:italic>in vivo</jats:italic> high-throughput approach we identified several hit compounds and could validate the impact of one of them on riboswitch-mediated gene regulation, albeit this might not be due to direct binding to the riboswitch. However, our data demonstrate the capability of our screening assay for bigger high-throughput screening campaigns. Furthermore, the screening system described here can not only be generally employed to detect non-natural ligands or compounds influencing riboswitches acting as genetic OFF switches, but it can also be used to investigate natural ligands of orphan OFF-riboswitches.</jats:p>