Engineering synthetic TAL effectors with orthogonal target sites.

Nucleic Acids Research
Abhishek GargThomas Z Armel

Abstract

The ability to engineer biological circuits that process and respond to complex cellular signals has the potential to impact many areas of biology and medicine. Transcriptional activator-like effectors (TALEs) have emerged as an attractive component for engineering these circuits, as TALEs can be designed de novo to target a given DNA sequence. Currently, however, the use of TALEs is limited by degeneracy in the site-specific manner by which they recognize DNA. Here, we propose an algorithm to computationally address this problem. We apply our algorithm to design 180 TALEs targeting 20 bp cognate binding sites that are at least 3 nt mismatches away from all 20 bp sequences in putative 2 kb human promoter regions. We generated eight of these synthetic TALE activators and showed that each is able to activate transcription from a targeted reporter. Importantly, we show that these proteins do not activate synthetic reporters containing mismatches similar to those present in the genome nor a set of endogenous genes predicted to be the most likely targets in vivo. Finally, we generated and characterized TALE repressors comprised of our orthogonal DNA binding domains and further combined them with shRNAs to accomplish near complete re...Continue Reading

References

Jul 24, 1998·Molecular Plant-microbe Interactions : MPMI·W ZhuF F White
Feb 16, 2002·Methods : a Companion to Methods in Enzymology·K J Livak, T D Schmittgen
Sep 6, 2005·Proceedings of the National Academy of Sciences of the United States of America·Frank StegmeierStephen J Elledge
Apr 29, 2006·Nature Genetics·Piero CarninciYoshihide Hayashizaki
Jun 2, 2006·Molecular Systems Biology·Ernesto AndrianantoandroRon Weiss
Oct 28, 2008·Nucleic Acids Research·Mark Koudritsky, Eytan Domany
Apr 30, 2009·Annual Review of Phytopathology·Jens Boch, Ulla Bonas
May 16, 2009·Proceedings of the National Academy of Sciences of the United States of America·Wenlin An, Jason W Chin
May 29, 2009·The Plant Journal : for Cell and Molecular Biology·Sabine KayUlla Bonas
Jul 1, 2009·Cell·Jeffrey J TaborAndrew D Ellington
Nov 17, 2009·Proceedings of the National Academy of Sciences of the United States of America·Patrick RömerThomas Lahaye
Nov 26, 2009·Science·Matthew J Moscou, Adam J Bogdanove
Dec 2, 2009·The Journal of Cell Biology·Karmella A Haynes, Pamela A Silver
Dec 17, 2009·Nature Biotechnology·Timothy K LuJames J Collins
Feb 27, 2010·Nucleic Acids Research·Oliver P T Barrett, Jason W Chin
Apr 17, 2010·Nature Reviews. Genetics·Ahmad S Khalil, James J Collins
May 6, 2010·PLoS Computational Biology·Kenzie D MacIsaacErnest Fraenkel
Jul 14, 2010·Nature Nanotechnology·Madeleine LeisnerYaakov Benenson
Jul 28, 2010·Genetics·Michelle ChristianDaniel F Voytas
Nov 26, 2010·Proceedings of the National Academy of Sciences of the United States of America·Robert MorbitzerThomas Lahaye
Dec 24, 2010·Virulence·Heidi Scholze, Jens Boch
Dec 24, 2010·Nature Biotechnology·Jeffrey C MillerEdward J Rebar
Jan 11, 2011·Current Opinion in Microbiology·Heidi Scholze, Jens Boch
Mar 23, 2011·Nucleic Acids Research·Robert MorbitzerThomas Lahaye
Jun 1, 2011·PloS One·Ernst WeberSylvestre Marillonnet
Dec 15, 2011·Plant Molecular Biology·Magdy M MahfouzJian-Kang Zhu
Jan 10, 2012·Science·Amanda Nga-Sze MakBarry L Stoddard
Feb 11, 2012·Nucleic Acids Research·Jason J LohmuellerPamela A Silver

❮ Previous
Next ❯

Citations

Sep 5, 2013·Molecular Therapy. Nucleic Acids·Pierre ChapdelaineJacques P Tremblay
Feb 5, 2013·Nature Methods·Pablo Perez-PineraCharles A Gersbach
Feb 12, 2013·Nature Methods·Morgan L MaederJ Keith Joung
Jul 3, 2013·Nature Methods·Justin Crocker, David L Stern
Nov 22, 2012·Nature Reviews. Molecular Cell Biology·J Keith Joung, Jeffry D Sander
Sep 29, 2012·Nucleic Acids Research·Andrew C MercerCarlos F Barbas
Feb 15, 2013·Nucleic Acids Research·Joshua F MecklerEnoch P Baldwin
Aug 29, 2013·Nucleic Acids Research·Florian LienertPamela A Silver
Jun 4, 2013·Annual Review of Phytopathology·Sebastian SchornackDiana M Horvath
Jan 16, 2013·Biological Procedures Online·Claudia Uhde-StoneBiao Lu
Mar 26, 2013·PLoS Computational Biology·Jan GrauJens Boch
Nov 15, 2013·PloS One·Junjiao YangWensheng Wei
Nov 28, 2013·PloS One·Basile I M WickyMatteo Dal Peraro
Jan 18, 2014·Nature Reviews. Molecular Cell Biology·Florian LienertPamela A Silver
Mar 13, 2014·Nature Communications·Annekatrin RichterJens Boch
Jan 15, 2014·Nature Chemical Biology·Rok GaberRoman Jerala
Jul 5, 2013·Chaos·Avi Robinson-MosherJeffrey Way
Feb 4, 2014·Journal of Biological Engineering·Mario A Marchisio
Dec 3, 2013·Progress in Biophysics and Molecular Biology·Mingjie ZhangXueqing Xu
May 2, 2014·Nature Methods·Jennifer A N Brophy, Christopher A Voigt
Feb 27, 2014·Nucleic Acids Research·Alexandre JuilleratPhilippe Duchateau
Jun 8, 2014·Briefings in Functional Genomics·Radek Jankele, Petr Svoboda
Sep 30, 2014·Nature Communications·Tina LebarRoman Jerala
Dec 10, 2013·Nature Chemical Biology·Brynne C StantonChristopher A Voigt
Oct 19, 2013·Nature Protocols·Matthew H LarsonLei S Qi
Feb 9, 2016·Nature Chemical Biology·Matthew F CopelandBrian F Pfleger
Oct 31, 2015·Biotechnology Advances·Gert PetersMarjan De Mey
Jun 26, 2013·Journal of Genetics and Genomics = Yi Chuan Xue Bao·Chuanxian WeiRenjie Jiao
Nov 28, 2012·Trends in Biotechnology·Joerg MampelJuergen Eck
Nov 2, 2014·ACS Synthetic Biology·Brynne C StantonChristopher A Voigt
Nov 21, 2013·ACS Synthetic Biology·Andrew C MercerCarlos F Barbas
Jun 17, 2014·ACS Synthetic Biology·Richard MooreLeonidas Bleris
Nov 14, 2014·ACS Synthetic Biology·Andreja MajerleRoman Jerala
Apr 4, 2014·ACS Synthetic Biology·Michael Guinn, Leonidas Bleris
Mar 19, 2014·Current Opinion in Biotechnology·Matthew F CopelandBrian F Pfleger
Jan 31, 2015·Scientific Reports·Alexandre JuilleratPhilippe Duchateau
May 30, 2015·Scientific Reports·Navneet RaiIlias Tagkopoulos
Oct 16, 2015·Journal of Molecular Biology·Robert W BradleyBaojun Wang
Mar 25, 2014·Protein Science : a Publication of the Protein Society·Rachael ParkerTijana Z Grove

❮ Previous
Next ❯

Methods Mentioned

BETA
transfections
Assay
transfection
fluorescence
flow cytometry
fluorescence microscopy

Software Mentioned

TALEs
Relative Expression Software Tool ( REST )
BioBrick
Metamorph

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.