An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system
Abstract 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 in...
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Nature Portfolio
2017
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oai:doaj.org-article:e1d0fcb6ce724c0993b9bc64d13571bf2021-12-02T15:06:21ZAn in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system10.1038/s41598-017-07870-w2045-2322https://doaj.org/article/e1d0fcb6ce724c0993b9bc64d13571bf2017-08-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-07870-whttps://doaj.org/toc/2045-2322Abstract 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 in vivo screening system that allows identifying OFF-riboswitch modulators in a 384 well bioluminescence assay format. We use a reverse reporter gene setup in Bacillus subtilis, 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 xpt riboswitch from B. anthracis. With this in vivo 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.Marion KirchnerKenji SchorppKamyar HadianSabine SchneiderNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-11 (2017) |
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DOAJ |
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Medicine R Science Q |
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Medicine R Science Q Marion Kirchner Kenji Schorpp Kamyar Hadian Sabine Schneider An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| description |
Abstract 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 in vivo screening system that allows identifying OFF-riboswitch modulators in a 384 well bioluminescence assay format. We use a reverse reporter gene setup in Bacillus subtilis, 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 xpt riboswitch from B. anthracis. With this in vivo 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. |
| format |
article |
| author |
Marion Kirchner Kenji Schorpp Kamyar Hadian Sabine Schneider |
| author_facet |
Marion Kirchner Kenji Schorpp Kamyar Hadian Sabine Schneider |
| author_sort |
Marion Kirchner |
| title |
An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| title_short |
An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| title_full |
An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| title_fullStr |
An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| title_full_unstemmed |
An in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| title_sort |
in vivo high-throughput screening for riboswitch ligands using a reverse reporter gene system |
| publisher |
Nature Portfolio |
| publishDate |
2017 |
| url |
https://doaj.org/article/e1d0fcb6ce724c0993b9bc64d13571bf |
| work_keys_str_mv |
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1718388484500094976 |