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.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Alkaline Phosphatase / biosynthesis*
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Alkaline Phosphatase / genetics*
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Animals
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Binding Sites / genetics
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CHO Cells
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Cricetulus
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GPI-Linked Proteins / biosynthesis
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GPI-Linked Proteins / genetics
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Gene Expression
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Gene Regulatory Networks*
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Green Fluorescent Proteins / genetics
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HEK293 Cells
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HeLa Cells
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Humans
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Isoenzymes / biosynthesis*
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Isoenzymes / genetics*
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Molecular Sequence Data
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Nucleic Acid Conformation
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Protein Biosynthesis*
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RNA, Catalytic / chemistry
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RNA, Catalytic / genetics
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RNA, Catalytic / metabolism*
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RNA, Messenger / biosynthesis
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RNA, Messenger / genetics
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Riboswitch*
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Structure-Activity Relationship
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Transgenes*
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Viral Proteins / genetics
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Viral Proteins / metabolism
Substances
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GPI-Linked Proteins
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Isoenzymes
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NutR protein, bacteriophage lambda
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RNA, Catalytic
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RNA, Messenger
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Riboswitch
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Viral Proteins
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enhanced green fluorescent protein
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Green Fluorescent Proteins
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Alkaline Phosphatase
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alkaline phosphatase, placental
Associated data
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GENBANK/KM403566
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GENBANK/KM403567
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GENBANK/KM403568
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GENBANK/KM403569