title: A general design strategy for protein-responsive riboswitches in mammalian cells.
abstract: RNAs are ideal for the design of gene switches that can monitor and program cellular behavior because of their high modularity and predictable structure-function relationship. We have assembled an expression platform with an embedded modular ribozyme scaffold that correlates self-cleavage activity of designer ribozymes with transgene translation in bacteria and mammalian cells. A design approach devised to screen ribozyme libraries in bacteria and validate variants with functional tertiary stem-loop structures in mammalian cells resulted in a designer ribozyme with a protein-binding nutR-boxB stem II and a selected matching stem I. In a mammalian expression context, this designer ribozyme exhibited dose-dependent translation control by the N-peptide, had rapid induction kinetics and could be combined with classic small molecule-responsive transcription control modalities to construct complex, programmable genetic circuits.
authors.affiliation: Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland.
authors.forename: Simon
authors.initials: S
authors.lastname: Ausländer
authors.affiliation: Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland.
authors.forename: Pascal
authors.initials: P
authors.lastname: Stücheli
authors.affiliation: Department of Chemistry, University of Konstanz, Konstanz, Germany.
authors.forename: Charlotte
authors.initials: C
authors.lastname: Rehm
authors.affiliation: Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland.
authors.forename: David
authors.initials: D
authors.lastname: Ausländer
authors.affiliation: Department of Chemistry, University of Konstanz, Konstanz, Germany.
authors.forename: Jörg S
authors.initials: JS
authors.lastname: Hartig
authors.affiliation: 1] Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland. [2] Faculty of Science, University of Basel, Basel, Switzerland.
authors.forename: Martin
authors.initials: M
authors.lastname: Fussenegger
mesh_terms: D000469:Alkaline Phosphatase / Q000096:biosynthesis* / Q000235:genetics*; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D003412:Cricetulus; D058851:GPI-Linked Proteins / Q000096:biosynthesis / Q000235:genetics; D015870:Gene Expression; D053263:Gene Regulatory Networks; D049452:Green Fluorescent Proteins / Q000235:genetics; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000096:biosynthesis* / Q000235:genetics*; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D014176:Protein Biosynthesis; D016337:RNA, Catalytic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D058928:Riboswitch; D013329:Structure-Activity Relationship; D019076:Transgenes; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D058851:GPI-Linked Proteins; D007527:Isoenzymes; C119762:NutR protein, bacteriophage lambda; D016337:RNA, Catalytic; D012333:RNA, Messenger; D058928:Riboswitch; D014764:Viral Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins; D000469:Alkaline Phosphatase; C019839:alkaline phosphatase, placental