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Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression

CRISPR-Cas systems have shown tremendous promise as heterologous tools for genome editing and transcriptional regulation. Because these RNA-directed immune systems are found in most prokaryotes, an opportunity exists to harness the endogenous systems as convenient tools in these organisms. Here, we...

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Autores principales: Luo, Michelle L., Mullis, Adam S., Leenay, Ryan T., Beisel, Chase L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4288209/
https://www.ncbi.nlm.nih.gov/pubmed/25326321
http://dx.doi.org/10.1093/nar/gku971
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author Luo, Michelle L.
Mullis, Adam S.
Leenay, Ryan T.
Beisel, Chase L.
author_facet Luo, Michelle L.
Mullis, Adam S.
Leenay, Ryan T.
Beisel, Chase L.
author_sort Luo, Michelle L.
collection PubMed
description CRISPR-Cas systems have shown tremendous promise as heterologous tools for genome editing and transcriptional regulation. Because these RNA-directed immune systems are found in most prokaryotes, an opportunity exists to harness the endogenous systems as convenient tools in these organisms. Here, we report that the Type I-E CRISPR-Cas system in Escherichia coli can be co-opted for programmable transcriptional repression. We found that deletion of the signature cas3 gene converted this immune system into a programmable gene regulator capable of reversible gene silencing of heterologous and endogenous genes. Targeting promoter regions yielded the strongest repression, whereas targeting coding regions showed consistent strand bias. Furthermore, multi-targeting CRISPR arrays could generate complex phenotypes. This strategy offers a simple approach to convert many endogenous Type I systems into transcriptional regulators, thereby expanding the available toolkit for CRISPR-mediated genetic control while creating new opportunities for genome-wide screens and pathway engineering.
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spelling pubmed-42882092015-02-19 Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression Luo, Michelle L. Mullis, Adam S. Leenay, Ryan T. Beisel, Chase L. Nucleic Acids Res Synthetic Biology and Bioengineering CRISPR-Cas systems have shown tremendous promise as heterologous tools for genome editing and transcriptional regulation. Because these RNA-directed immune systems are found in most prokaryotes, an opportunity exists to harness the endogenous systems as convenient tools in these organisms. Here, we report that the Type I-E CRISPR-Cas system in Escherichia coli can be co-opted for programmable transcriptional repression. We found that deletion of the signature cas3 gene converted this immune system into a programmable gene regulator capable of reversible gene silencing of heterologous and endogenous genes. Targeting promoter regions yielded the strongest repression, whereas targeting coding regions showed consistent strand bias. Furthermore, multi-targeting CRISPR arrays could generate complex phenotypes. This strategy offers a simple approach to convert many endogenous Type I systems into transcriptional regulators, thereby expanding the available toolkit for CRISPR-mediated genetic control while creating new opportunities for genome-wide screens and pathway engineering. Oxford University Press 2015-01-09 2014-10-17 /pmc/articles/PMC4288209/ /pubmed/25326321 http://dx.doi.org/10.1093/nar/gku971 Text en © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Synthetic Biology and Bioengineering
Luo, Michelle L.
Mullis, Adam S.
Leenay, Ryan T.
Beisel, Chase L.
Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title_full Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title_fullStr Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title_full_unstemmed Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title_short Repurposing endogenous type I CRISPR-Cas systems for programmable gene repression
title_sort repurposing endogenous type i crispr-cas systems for programmable gene repression
topic Synthetic Biology and Bioengineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4288209/
https://www.ncbi.nlm.nih.gov/pubmed/25326321
http://dx.doi.org/10.1093/nar/gku971
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