Aptazyme-Mediated Regulation of 16S Ribosomal RNA

SummaryDeveloping artificial genetic switches in order to control gene expression via an external stimulus is an important aim in chemical and synthetic biology. Here, we expand the application range of RNA switches to the regulation of 16S rRNA function in Escherichia coli. For this purpose, we incorporated hammerhead ribozymes at several positions into orthogonalized 16S rRNA. We observed that ribosomal function is remarkably tolerant toward the incorporation of large additional RNA fragments at certain sites of the 16S rRNA. However, ribozyme-mediated cleavage results in severe reduction of 16S rRNA stability. We carried out an in vivo screen for the identification of sequences acting as ligand-responsive RNA switches, enabling thiamine-dependent switching of 16S rRNA function. In addition to expanding the regulatory toolbox, the presented artificial riboswitches should prove valuable to study aspects of rRNA folding and stability in bacteria.

Graphical AbstractFigure optionsDownload full-size imageDownload high-quality image (126 K)Download as PowerPoint slideHighlights► The bacterial 16S rRNA is tolerant toward insertion of large sequences ► Hammerhead ribozymes cleave very efficiently in the 16S rRNA context ► 16S rRNA can be inactivated by in cis-cleavage of a ribozyme ► Using aptazymes, SSU activity can be controlled on demand by thiamine