Competitive dCas9 binding as a mechanism for transcriptional control

Abstract Catalytically dead Cas9 (dCas9) is a programmable transcription factor that can be targeted to promoters through the design of small guide RNAs (sgRNAs), where it can function as an activator or repressor. Natural promoters use overlapping binding sites as a mechanism for signal integration...

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Autores principales:	Daniel A Anderson, Christopher A Voigt
Formato:	article
Lenguaje:	EN
Publicado:	Wiley 2021
Materias:	analog circuit CRISPRi ratio sensing synthetic biology Biology (General) QH301-705.5 Medicine (General) R5-920
Acceso en línea:	https://doaj.org/article/731a42a837d54e9783bf363e115c67f8
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spelling	oai:doaj.org-article:731a42a837d54e9783bf363e115c67f82021-11-29T08:21:36ZCompetitive dCas9 binding as a mechanism for transcriptional control1744-429210.15252/msb.202110512https://doaj.org/article/731a42a837d54e9783bf363e115c67f82021-11-01T00:00:00Zhttps://doi.org/10.15252/msb.202110512https://doaj.org/toc/1744-4292Abstract Catalytically dead Cas9 (dCas9) is a programmable transcription factor that can be targeted to promoters through the design of small guide RNAs (sgRNAs), where it can function as an activator or repressor. Natural promoters use overlapping binding sites as a mechanism for signal integration, where the binding of one can block, displace, or augment the activity of the other. Here, we implemented this strategy in Escherichia coli using pairs of sgRNAs designed to repress and then derepress transcription through competitive binding. When designed to target a promoter, this led to 27‐fold repression and complete derepression. This system was also capable of ratiometric input comparison over two orders of magnitude. Additionally, we used this mechanism for promoter sequence‐independent control by adopting it for elongation control, achieving 8‐fold repression and 4‐fold derepression. This work demonstrates a new genetic control mechanism that could be used to build analog circuit or implement cis‐regulatory logic on CRISPRi‐targeted native genes.Daniel A AndersonChristopher A VoigtWileyarticleanalog circuitCRISPRiratio sensingsynthetic biologyBiology (General)QH301-705.5Medicine (General)R5-920ENMolecular Systems Biology, Vol 17, Iss 11, Pp n/a-n/a (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	analog circuit CRISPRi ratio sensing synthetic biology Biology (General) QH301-705.5 Medicine (General) R5-920
spellingShingle	analog circuit CRISPRi ratio sensing synthetic biology Biology (General) QH301-705.5 Medicine (General) R5-920 Daniel A Anderson Christopher A Voigt Competitive dCas9 binding as a mechanism for transcriptional control
description	Abstract Catalytically dead Cas9 (dCas9) is a programmable transcription factor that can be targeted to promoters through the design of small guide RNAs (sgRNAs), where it can function as an activator or repressor. Natural promoters use overlapping binding sites as a mechanism for signal integration, where the binding of one can block, displace, or augment the activity of the other. Here, we implemented this strategy in Escherichia coli using pairs of sgRNAs designed to repress and then derepress transcription through competitive binding. When designed to target a promoter, this led to 27‐fold repression and complete derepression. This system was also capable of ratiometric input comparison over two orders of magnitude. Additionally, we used this mechanism for promoter sequence‐independent control by adopting it for elongation control, achieving 8‐fold repression and 4‐fold derepression. This work demonstrates a new genetic control mechanism that could be used to build analog circuit or implement cis‐regulatory logic on CRISPRi‐targeted native genes.
format	article
author	Daniel A Anderson Christopher A Voigt
author_facet	Daniel A Anderson Christopher A Voigt
author_sort	Daniel A Anderson
title	Competitive dCas9 binding as a mechanism for transcriptional control
title_short	Competitive dCas9 binding as a mechanism for transcriptional control
title_full	Competitive dCas9 binding as a mechanism for transcriptional control
title_fullStr	Competitive dCas9 binding as a mechanism for transcriptional control
title_full_unstemmed	Competitive dCas9 binding as a mechanism for transcriptional control
title_sort	competitive dcas9 binding as a mechanism for transcriptional control
publisher	Wiley
publishDate	2021
url	https://doaj.org/article/731a42a837d54e9783bf363e115c67f8
work_keys_str_mv	AT danielaanderson competitivedcas9bindingasamechanismfortranscriptionalcontrol AT christopheravoigt competitivedcas9bindingasamechanismfortranscriptionalcontrol
_version_	1718407468993740800

Competitive dCas9 binding as a mechanism for transcriptional control

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