A high-throughput screen for synthetic riboswitches reveals mechanistic insights into their function.
Riboswitches are RNA-based genetic control elements that regulate gene expression in a ligand-dependent fashion without the need for proteins. The ability to create synthetic riboswitches that control gene expression in response to any desired small-molecule ligand will enable the development of sensitive genetic screens that can detect the presence of small molecules, as well as designer genetic control elements to conditionally modulate cellular behavior. Herein, we present an automated high-throughput screening method that identifies synthetic riboswitches that display extremely low background levels of gene expression in the absence of the desired ligand and robust increases in expression in its presence. Mechanistic studies reveal how these riboswitches function and suggest design principles for creating new synthetic riboswitches. We anticipate that the screening method and design principles will be generally useful for creating functional synthetic riboswitches.
Lynch
Sean A
SA
Department of Chemistry and Center for Fundamental and Applied Molecular Evolution, Emory University, Atlanta, GA 30322, USA.
Desai
Shawn K
SK
Sajja
Hari Krishna
HK
Gallivan
Justin P
JP
D052157:Aptamers, Nucleotide / Q000235:genetics; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D015723:Gene Library; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009690:Nucleic Acid Conformation; D012329:RNA, Bacterial / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D038621:Regulatory Sequences, Ribonucleic Acid / Q000235:genetics / Q000502:physiology*; D013806:Theophylline / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
D052157:Aptamers, Nucleotide; D012329:RNA, Bacterial; D012333:RNA, Messenger; D038621:Regulatory Sequences, Ribonucleic Acid; D013806:Theophylline; D001616:beta-Galactosidase