Reprogramming Bacteria to Seek and Destroy a Herbicide

A major goal of synthetic biology is to reprogram cells to perform complex tasks. Here we show how a combination of in vitro and in vivo selection rapidly identifies a synthetic riboswitch that activates protein translation in response to the herbicide atrazine. We further demonstrate that this riboswitch can reprogram bacteria to migrate in the presence of atrazine. Finally, we show that incorporating a gene from an atrazine catabolic pathway allows these cells to seek and destroy atrazine.

[1]  H. Berg,et al.  Complex patterns formed by motile cells of Escherichia coli , 1991, Nature.

[2]  Günter Mayer,et al.  An RNA aptamer that induces transcription. , 2009, Chemistry & biology.

[3]  S. K. Desai,et al.  Genetic screens and selections for small molecules based on a synthetic riboswitch that activates protein translation. , 2004, Journal of the American Chemical Society.

[4]  L. Gold,et al.  Systematic evolution of ligands by exponential enrichment: RNA ligands to bacteriophage T4 DNA polymerase. , 1990, Science.

[5]  Y Wang,et al.  RNA molecules that specifically and stoichiometrically bind aminoglycoside antibiotics with high affinities. , 1996, Biochemistry.

[6]  B. Suess,et al.  A designed RNA shuts down transcription. , 2007, Chemistry & biology.

[7]  Koichi Abe,et al.  Mechanism‐Guided Library Design and Dual Genetic Selection of Synthetic OFF Riboswitches , 2009, Chembiochem : a European journal of chemical biology.

[8]  A D Ellington,et al.  In vitro evolution of beta-glucuronidase into a beta-galactosidase proceeds through non-specific intermediates. , 2001, Journal of molecular biology.

[9]  Shana Topp,et al.  Riboswitches in unexpected places--a synthetic riboswitch in a protein coding region. , 2008, RNA.

[10]  Yohei Yokobayashi,et al.  An efficient platform for genetic selection and screening of gene switches in Escherichia coli , 2009, Nucleic acids research.

[11]  S. Clay,et al.  Adsorption and Desorption of Atrazine, Hydroxyatrazine, and S-Glutathione Atrazine on Two Soils , 1990, Weed Science.

[12]  L. Wackett,et al.  Biodegradation of atrazine in transgenic plants expressing a modified bacterial atrazine chlorohydrolase (atzA) gene. , 2005, Plant biotechnology journal.

[13]  Karl-Dieter Entian,et al.  A fast and efficient translational control system for conditional expression of yeast genes , 2009, Nucleic acids research.

[14]  L. Wackett,et al.  The Atrazine Catabolism Genes atzABC Are Widespread and Highly Conserved , 1998, Journal of bacteriology.

[15]  J. Gallivan,et al.  A flow cytometry-based screen for synthetic riboswitches , 2008, Nucleic acids research.

[16]  J. S. Parkinson,et al.  Collaborative signaling by bacterial chemoreceptors. , 2005, Current opinion in microbiology.

[17]  B. Suess,et al.  A theophylline responsive riboswitch based on helix slipping controls gene expression in vivo. , 2004, Nucleic acids research.

[18]  David R. Liu,et al.  Engineering a ligand-dependent RNA transcriptional activator. , 2004, Chemistry & biology.

[19]  L. Wackett,et al.  Molecular Basis of a Bacterial Consortium: Interspecies Catabolism of Atrazine , 2000, Applied and Environmental Microbiology.

[20]  M. Famulok,et al.  A novel RNA motif for neomycin recognition. , 1995, Chemistry & biology.

[21]  L. Wackett,et al.  Isolation and Characterization of a Pseudomonas sp. That Mineralizes the s-Triazine Herbicide Atrazine , 1995, Applied and environmental microbiology.

[22]  R. Breaker,et al.  In-line probing analysis of riboswitches. , 2008, Methods in molecular biology.

[23]  Yohei Yokobayashi,et al.  Engineering complex riboswitch regulation by dual genetic selection. , 2008, Journal of the American Chemical Society.

[24]  L. Wackett,et al.  Atrazine chlorohydrolase from Pseudomonas sp. strain ADP: gene sequence, enzyme purification, and protein characterization , 1996, Journal of bacteriology.

[25]  L. Wackett,et al.  Biodegradation of atrazine and related s-triazine compounds: from enzymes to field studies , 2001, Applied Microbiology and Biotechnology.

[26]  Beatrix Suess,et al.  Screening for engineered neomycin riboswitches that control translation initiation. , 2007, RNA.

[27]  M. Goulian,et al.  Changing the specificity of a bacterial chemoreceptor. , 2006, Journal of molecular biology.

[28]  R R Breaker,et al.  Relationship between internucleotide linkage geometry and the stability of RNA. , 1999, RNA.

[29]  G. Wadhams,et al.  Making sense of it all: bacterial chemotaxis , 2004, Nature Reviews Molecular Cell Biology.

[30]  J. Sabina,et al.  Expanded sequence dependence of thermodynamic parameters improves prediction of RNA secondary structure. , 1999, Journal of molecular biology.

[31]  S. K. Desai,et al.  A high-throughput screen for synthetic riboswitches reveals mechanistic insights into their function. , 2007, Chemistry & biology.

[32]  J. Oakeshott,et al.  Catalytic Improvement and Evolution of Atrazine Chlorohydrolase , 2009, Applied and Environmental Microbiology.

[33]  M. Win,et al.  A modular and extensible RNA-based gene-regulatory platform for engineering cellular function , 2007, Proceedings of the National Academy of Sciences.

[34]  J. Adler Chemotaxis in Bacteria , 1966, Science.

[35]  C. Berens,et al.  A tetracycline-binding RNA aptamer. , 2001, Bioorganic & medicinal chemistry.

[36]  A. Pardi,et al.  High-resolution molecular discrimination by RNA. , 1994, Science.

[37]  Andrew D Ellington,et al.  Group I aptazymes as genetic regulatory switches , 2002, BMC biotechnology.

[38]  H. Mori,et al.  Construction of Escherichia coli K-12 in-frame, single-gene knockout mutants: the Keio collection , 2006, Molecular systems biology.

[39]  Yohei Yokobayashi,et al.  Dual selection of a genetic switch by a single selection marker , 2007, Biosyst..

[40]  M. Green,et al.  Controlling gene expression in living cells through small molecule-RNA interactions. , 1998, Science.

[41]  Dan S. Tawfik,et al.  The 'evolvability' of promiscuous protein functions , 2005, Nature Genetics.

[42]  Shana Topp,et al.  Random Walks to Synthetic Riboswitches—A High‐Throughput Selection Based on Cell Motility , 2008, Chembiochem : a European journal of chemical biology.

[43]  Michael Zuker,et al.  Mfold web server for nucleic acid folding and hybridization prediction , 2003, Nucleic Acids Res..

[44]  J. Gallivan,et al.  Guiding bacteria with small molecules and RNA. , 2007, Journal of the American Chemical Society.

[45]  B. Suess,et al.  A novel mechanism for translation initiation operates in haloarchaea , 2009, Molecular microbiology.

[46]  Mark Goulian,et al.  Engineered single- and multi-cell chemotaxis pathways in E. coli , 2009, Molecular systems biology.

[47]  Barbara Fink,et al.  Molecular analysis of a synthetic tetracycline-binding riboswitch. , 2005, RNA.

[48]  J. Szostak,et al.  In vitro selection of RNA molecules that bind specific ligands , 1990, Nature.